Natascia Tiso
Università degli Studi di Padova, Biology, Faculty Member
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Research Interests: Biochemistry, Chromatography, Catalysis, Comparative Genomics, Biology, and 15 moreBiological Chemistry, Biological Sciences, Cercopithecus aethiops, Animals, Archaea, Biochemical, Biomed, CHEMICAL SCIENCES, Applied Microbiology and Biotechnology, Biochimie, Aged, Amino Acid Sequence, Base Sequence, Biochemistry and cell biology, and Beta Lactamases
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Evaluation of Aβ(25-35) in 3D spheroids and zebrafish larvae
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Evaluation of caspae 3 staining after treatment with Aβ(25-35) in mice
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CD271 (NGFR) is a neurotrophin receptor that belongs to the tumor necrosis receptor (TNFR) family. Upon ligand binding, CD271 can mediate either survival or cell death. Although the role of CD271 as a marker of tumor-initiating cells is... more
CD271 (NGFR) is a neurotrophin receptor that belongs to the tumor necrosis receptor (TNFR) family. Upon ligand binding, CD271 can mediate either survival or cell death. Although the role of CD271 as a marker of tumor-initiating cells is still a matter of debate, its role in melanoma progression has been well documented. Moreover, CD271 has been shown to be upregulated after exposure to both chemotherapy and targeted therapy. In this study, we demonstrate that activation of CD271 by a short β-amyloid–derived peptide (Aβ(25–35)) in combination with either chemotherapy or MAPK inhibitors induces apoptosis in 2D and 3D cultures of eight melanoma cell lines. This combinatorial treatment significantly reduced metastasis in a zebrafish xenograft model and led to significantly decreased tumor volume in mice. Administration of Aβ(25–35) in ex vivo tumors from immunotherapy- and targeted therapy–resistant patients significantly reduced proliferation of melanoma cells, showing that activation ...
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The mitochondrial protein IF1 binds to the catalytic domain of the ATP synthase and inhibits ATP hydrolysis in ischemic tissues. Moreover, IF1 is overexpressed in many tumors and has been shown to act as a pro-oncogenic protein, although... more
The mitochondrial protein IF1 binds to the catalytic domain of the ATP synthase and inhibits ATP hydrolysis in ischemic tissues. Moreover, IF1 is overexpressed in many tumors and has been shown to act as a pro-oncogenic protein, although its mechanism of action is still debated. Here, we show that ATP5IF1 gene disruption in HeLa cells decreases colony formation in soft agar and tumor mass development in xenografts, underlining the role of IF1 in cancer. Notably, the lack of IF1 does not affect proliferation or oligomycin-sensitive mitochondrial respiration, but it sensitizes the cells to the opening of the permeability transition pore (PTP). Immunoprecipitation and proximity ligation analysis show that IF1 binds to the ATP synthase OSCP subunit in HeLa cells under oxidative phosphorylation conditions. The IF1–OSCP interaction is confirmed by NMR spectroscopy analysis of the recombinant soluble proteins. Overall, our results suggest that the IF1-OSCP interaction protects cancer cells...
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The transcription factor STAT3 is required for proliferation and pluripotency of embryonic stem cells; we have prepared and characterized fluorescent STAT3-reporter zebrafish based on repeats of minimal responsive elements. These... more
The transcription factor STAT3 is required for proliferation and pluripotency of embryonic stem cells; we have prepared and characterized fluorescent STAT3-reporter zebrafish based on repeats of minimal responsive elements. These transgenic lines mimic in vivo STAT3 expression patterns and are responsive to exogenous STAT3; notably, fluorescence is inhibited by both stat3 knock-out and IL6/JAK/STAT inhibitors. At larval stages, STAT3 reporter activity correlates with proliferating regions of the brain, haematopoietic tissue and intestine. In the adult gut the reporter is active in sparse proliferating cells, located at the base of intestinal folds, expressing the stemness marker sox9b and having the mammalian Crypt Base Columnar cells morphology; noteworthy, zebrafish stat3 mutants show defects in intestinal folding. The STAT3 reporter activity in the gut is abolished in mutants of Tcf7l2, the intestinal mediator of Wnt/β-catenin-dependent transcription, and the Wnt/β-catenin depend...
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Foetal Growth Restriction (FGR), previously known as Intrauterine Growth Restriction (IUGR), is an obstetrical condition due to placental insufficiency, affecting yearly about 30 million newborns worldwide. In this work, we aimed to... more
Foetal Growth Restriction (FGR), previously known as Intrauterine Growth Restriction (IUGR), is an obstetrical condition due to placental insufficiency, affecting yearly about 30 million newborns worldwide. In this work, we aimed to identify and pharmacologically target signalling pathways specifically involved in the FGR condition, focusing on FGR-related cardiovascular phenotypes. The transcriptional profile of human umbilical cords from FGR and control cases was compared with the response to hypoxia of zebrafish (Danio rerio) transgenic lines reporting in vivo the activity of twelve signalling pathways involved in embryonic development. Wnt/β-catenin and Jak/Stat3 were found as key pathways significantly dysregulated in both human and zebrafish samples. This information was used in a chemical-genetic analysis to test drugs targeting Wnt/β-catenin and Jak/Stat3 pathways to rescue a set of FGR phenotypes, including growth restriction and cardiovascular modifications. Treatments wit...
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Additional file 5 : Fig.S5. Tbar clusters in responsive neurons. Distributions of neuronal spectral responses classified with Tbar and shown as a function of their overall response to stimuli (quantified by T4D). Comparison of emergence... more
Additional file 5 : Fig.S5. Tbar clusters in responsive neurons. Distributions of neuronal spectral responses classified with Tbar and shown as a function of their overall response to stimuli (quantified by T4D). Comparison of emergence of the different Tbar classes for two different choices of T thresholds.
Additional file 8 : Fig.S8. T distributions at different stimulus intensities for stimulated and control larvae at 5 dpf. Comparison of neuronal response distributions for the four different wavelengths at three different powers.
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Background Visually guided behaviors such as optomotor and optokinetic responses, phototaxis, and prey capture are crucial for survival in zebrafish and become apparent after just a few days of development. Color vision, which in... more
Background Visually guided behaviors such as optomotor and optokinetic responses, phototaxis, and prey capture are crucial for survival in zebrafish and become apparent after just a few days of development. Color vision, which in zebrafish is based on a spatially anisotropic tetrachromatic retina, provides an additional important component of world representation driving fundamental larval behaviors. However, little is known about the central nervous system (CNS) circuitry underlying color vision processing downstream of the retina, and its activity correlates with behavior. Here, we used the transparent larva of zebrafish to image CNS neurons and their activity in response to colored visual stimuli. Results To investigate the processing of chromatic information in the zebrafish larva brain, we mapped with cellular resolution, spectrally responsive neurons in the larva encephalon and spinal cord. We employed the genetically encoded calcium indicator GCaMP6s and two-photon microscopy...
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<p>A) Integrating our data from this study with previous work we suggest how the initial DRG consisting of bipotent DRG progenitors (black rectangles) become specified (second row) and the resultant cell-fates adopted (bottom row).... more
<p>A) Integrating our data from this study with previous work we suggest how the initial DRG consisting of bipotent DRG progenitors (black rectangles) become specified (second row) and the resultant cell-fates adopted (bottom row). We outline the process in WT (neuron (N, cyan) and glial fates (G, magenta); first column), <i>baz1</i> (neuron only; second column), <i>m618</i> (failure to undergo fate specification; third column) and <i>neurog1</i> mutants (all adopt glia; fourth column). The model summarises at each stage the relative levels of inputs (large font is high activity, small font is weak activity) from each of Sox10, Neurog1, Notch signaling and a Sox10-dependent Factor X, proposed to provide a repressive input on <i>neurog1</i> expression. Colour coding of text indicates function of each input (i.e. Neurog1 drives sensory neuron, Notch signaling and Factor X drive glial specification), but Sox10 (which has independent roles in specification of both fates) is shown in both colours where WT, and in blue in <i>baz1</i> and in black in <i>m618</i> to reflect the neural fate specification activity retained in these mutants. In the case of Sox10 input, note that also font size in neighbouring cells refers to activity with respect to fate being adopted by each cell, since <i>baz1</i> mutant reveals that these roles can be separated. The underlying gene regulatory network is outlined in panel B). Asterisk in <i>m618</i> cells denotes that sensory neuron specification in <i>m618</i> mutants occurs in the absence of Sox10 activity, as a result of Sox9b-mediated transcription of <i>neurog1</i>[<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0172947#pone.0172947.ref013" target="_blank">13</a>]. See text for further details. We note that the data from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0172947#pone.0172947.g008" target="_blank">Fig 8</a> indicates the possibility that Sox10(baz1) may have unique non-autonomous effects on <u><i>neurog1</i> expression, which, if confirmed, would require modification of the model proposed.</u></p
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Research Interests: Biology, Cell Biology, Medicine, In Situ Hybridization, Bone Morphogenetic Proteins, and 15 moreBiological Sciences, Cell Differentiation, Mutation, Pancreas, Animals, Gastrulation, Mechanisms, Glycoproteins, Body Patterning, Embryos, Embryo Development, High mobility group proteins, endoderm, Medical and Health Sciences, and Expression pattern
doi: 10.1152/physiolgenomics.00227.2005 You might find this additional info useful... Supplementary material for this article can be found at:
Zebrafish reporter lines reveal in vivo signaling pathway activities involved in pancreatic cancer
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3653 Poster Board III-589 Introduction Human AF9 (alias MLLT3) gene was initially identified as one of the most common translocation partners of MLL associated with acute myeloid leukemia. More recently it has been demonstrated that AF9... more
3653 Poster Board III-589 Introduction Human AF9 (alias MLLT3) gene was initially identified as one of the most common translocation partners of MLL associated with acute myeloid leukemia. More recently it has been demonstrated that AF9 can be a positive regulator of early erythroid/megakaryocytic lineage decisions. However, many biological functions of AF9 and its role in leukemia remain unclear. Here, we aimed to identify the zebrafish homologue of human AF9 and to analyze its function using the zebrafish model system. Methods and Results A single zebrafish af9 gene was identified by homology searching at zebrafish genome database (www.ncbi.nlm.nih.gov/). The putative af9 protein showed 60% identity and 73% similarity to human counterpart. These results from the perspective of phylogenetic and synteny analyses indicated that zebrafish af9 was the likely homologue of human AF9. We used whole-mount in situ hybridization to analyzed its spatial and temporal expression patterns during...
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2600 Background AF9 is a transcription factor that plays an essential role in hematopoiesis and embryonic development. The alteration of AF9 is principally associated in acute myeloid leukemia as fusion partner of human MLL (mixed-lineage... more
2600 Background AF9 is a transcription factor that plays an essential role in hematopoiesis and embryonic development. The alteration of AF9 is principally associated in acute myeloid leukemia as fusion partner of human MLL (mixed-lineage leukemia) gene rearrangements. Zebrafish is an excellent model organism to study embryonic development and hematopoiesis. We have previously shown that zebrafish af9 is expressed within the intermediate cell mass (ICM), a site of primitive hematopoiesis in zebrafish. Here we study the loss of af9 in zebrafish development to further understand how af9 modulates early hematopoietic and embryonic development. Methods and results Two morpholino antisense oligos (MOs), designed to block translation and inhibit pre-mRNA splicing of af9, were co-injected in embryos at 1–2 cell stage. To control for off-target effects, two morpholino mismatch oligos were designed and co-injected. Efficacy of MOs was demonstrated by Western blot analysis and RT-PCR in contr...
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Research Interests: Biochemistry, Endocrinology, Computational Biology, Biology, Membrane Proteins, and 15 moreClinical Biochemistry, Medicine, Humans, Child, Female, Animals, Male, Clinical Sciences, Fluorescent Antibody Technique, Adult, Zebrafish, Calcium Binding Proteins, Alagille syndrome, Child preschool, and Paediatrics and reproductive medicine
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ABSTRACT In the last decades, the tropical teleost zebrafish (Danio rerio) has proved to be an excellent vertebrate system to model mammalian molecular events occurring during embryonic development, organ formation and adult physiology,... more
ABSTRACT In the last decades, the tropical teleost zebrafish (Danio rerio) has proved to be an excellent vertebrate system to model mammalian molecular events occurring during embryonic development, organ formation and adult physiology, either under normal or pathological conditions. Low costs and small dimensions, external fertilization and high fecundity, tissue transparency, rapid development, availability of mutant and transgenic lines, easy manipulability for gene perturbation and pharmacological screening, are some examples of the advantages characterizing this model organism. In the past 15 years, the use of zebrafish in the endocrinology field has been mainly focused on the analysis of endocrine organ development. Our team and other research groups have elucidated the main steps leading to the formation of endocrine glands such as pancreatic islets and thyroid, hypothalamus and pituitary, interrenal gland and gonads. Comparison of the zebrafish endocrine system to that of mammals has demonstrated that the systems are sufficiently similar for zebrafish to be employed as a model for endocrine research. In more recent years, new zebrafish-based tools have been generated to elucidate in vivo the molecular cross-talks occurring among cells, tissues and organs. Signalling pathway reporter lines represent an interesting implementation of classical transgenesis to visualize in vivo, in an intact organism, the anatomical regions that, in a given time interval, are responding to a specific signalling pathway. To generate these transgenic fish lines, signal-specific responsive sequences, identified at the genomic level, are multimerized and placed upstream of reporter genes, typically encoding for fluorescent proteins such as GFP or mCherry. At present, a series of pathway reporter lines are already available, among which Bmp, Shh, FGF, Notch, TGFβ, Wnt, hypoxia and glucocorticoid signalling. Our group is currently generating and validating additional lines, among which the cAMP/CREB pathway, while others, such as thyroid hormone and Foxo signalling, are in the planning phase. Preliminary results and envisaged applications will be presented and discussed.
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olig genes encode a previously unrecognized group of vertebrate-specific basic helix–loop–helix transcription factors. As shown in mice, chickens, and zebrafish, two members of this group, olig1 and olig2 , are involved in the... more
olig genes encode a previously unrecognized group of vertebrate-specific basic helix–loop–helix transcription factors. As shown in mice, chickens, and zebrafish, two members of this group, olig1 and olig2 , are involved in the differentiation of motoneurons and oligodendrocytes, but nothing is known about the role of the third member, olig3 . Here, we show that olig3 plays an essential role in the establishment of the neural crest–lateral neural plate boundary. In zebrafish embryos, morpholino-induced olig3 inactivation dramatically increases the number of neural crest cells, but lateral neural plate fates (interneurons and astrocytes) are missing. Zebrafish swirl mutants that have impaired bone morphogenetic protein signaling and lack neural crest cells display an expanded olig3 expression domain. Moreover, olig3 is up-regulated in mindbomb mutants lacking the neural crest because of an impaired notch signaling, and olig3 repression in such mutants rescues the neural crest. In addi...
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Solute carrier 15 (SLC15) membrane proteins PEPT1 (SLC15A1) and PEPT2 (SLC15A2) have been described in great detail in mammals. In contrast, information in lower vertebrates is limited. We characterized the functional properties of a... more
Solute carrier 15 (SLC15) membrane proteins PEPT1 (SLC15A1) and PEPT2 (SLC15A2) have been described in great detail in mammals. In contrast, information in lower vertebrates is limited. We characterized the functional properties of a novel zebrafish peptide transporter orthologous to mammalian and avian PEPT2, described its gene ( pept2) structure, and determined mRNA tissue distribution. An expressed sequence tag (EST) cDNA (Integrated Molecular Analysis of Gene Expression; IMAGE) corresponding to zebrafish pept2 was completed by inserting a stretch of 75 missing nucleotides in the coding sequence to obtain a 3,238-bp functional clone. The complete open reading frame (ORF) was 2,160 bp and encoded a 719-amino acid protein. Electrophysiological analysis after cRNA injection in Xenopus laevis oocytes suggested that zebrafish PEPT2 is a high-affinity/low-capacity transporter ( K0.5for glycyl-l-glutamine ∼18 μM at −120 mV and pH 7.5). Zebrafish pept2 gene was 19,435 kb, thus being the ...
Research Interests: Electrophysiology, Biology, Medicine, Danio rerio, In Situ Hybridization, and 15 morePhylogeny, Female, Animals, Male, Medical Physiology, Inner Ear, Membrane transport proteins, Genome Organization, Larva, Amino Acid Sequence, Base Sequence, Oocytes, Expression analysis, Functional Properties, and Molecular Sequence Data
Research Interests: Computational Biology, Biotechnology, Biology, Membrane Proteins, Medicine, and 15 moreDanio rerio, Humans, Animals, Developmental disabilities, Animal Model, Medical Physiology, Clinical Sciences, Axonal Degeneration, Embryonic Development, Body Patterning, CHarcot Marie Tooth, Charcot Marie Tooth Disease, Motor activity, Mitochondrial Proteins, and MFN
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Research Interests: Genetics, Genomics, Biology, Public Health, Morphogenesis, and 15 moreMedicine, Pancreatic Cancer, Functional Genomics, Danio rerio, Biological Sciences, Cell Differentiation, Pancreas, Animals, Early development, Animal Model, Differentiation, Behavioral Animal Models, Gene Function, Large Scale, and Medical and Health Sciences
Research Interests: Extracellular Matrix, Signal Transduction, Biological Sciences, Humans, Animals, and 14 moreEarly development, Glycosaminoglycan, Spectrum, Body Patterning, Transforming Growth Factor Beta, Zebrafish, Metabolic Disorder, Growth Factor, Matrix Biology, Lysosomal Storage Disorder, Transforming Growth Factor, Molecular Sequence Data, Cell Membrane, and Craniofacial Abnormalities
The mechanisms underlying the early steps of thyroid development are largely unknown. In search for novel candidate genes implicated in thyroid function, we performed a gene expression analysis on thyroid cells revealing that TSH... more
The mechanisms underlying the early steps of thyroid development are largely unknown. In search for novel candidate genes implicated in thyroid function, we performed a gene expression analysis on thyroid cells revealing that TSH regulates the expression of several elements of the Notch pathway, including the ligand Jagged1. Because the Notch pathway is involved in cell-fate determination of several foregut-derived endocrine tissues, we tested its contribution in thyroid development using the zebrafish, a teleost model recapitulating the mammalian molecular events during thyroid development. Perturbing the Notch signaling (e.g. mib mutants, γ-secretase inhibition, or Notch intracellular domain overexpression), we obtained evidence that this pathway has a biological role during the earlier phases of thyroid primordium induction, limiting the number of cells that proceed to a specialized fate and probably involving actions from surrounding tissues. Moreover, we were able to confirm th...
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Emilins are a family of extracellular matrix proteins with common structural organization and containing a characteristic N-terminal cysteine-rich domain. The prototype of this family, Emilin-1, is found in human and murine organs in... more
Emilins are a family of extracellular matrix proteins with common structural organization and containing a characteristic N-terminal cysteine-rich domain. The prototype of this family, Emilin-1, is found in human and murine organs in association with elastic fibers, and other emilins were recently isolated in mammals. To gain insight into these proteins in lower vertebrates, we investigated the expression of emilins in the fish Danio rerio. Using sequence similarity tools, we identified eight members of this family in zebrafish. Each emilin gene has two paralogs in zebrafish, showing conserved structure with the human ortholog. In situ hybridization revealed that expression of zebrafish emilin genes is regulated in a spatiotemporal manner during embryonic development, with overlapping and site-specific patterns mostly including mesenchymal structures. Expression of certain emilin genes in peculiar areas, such as the central nervous system or the posterior notochord, suggests that they may play a role in key morphogenetic processes.
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Research Interests: Developmental Biology, Biology, Membrane Proteins, Cell Biology, Medicine, and 15 moreIn Situ Hybridization, Embryo, Biological Sciences, Phylogeny, Pancreas, Animals, Notochord, Vertebrate, Nervous System, Calcium Binding Proteins, Notch Signaling, Embryos, Expression analysis, Pronephros, and Medical and Health Sciences
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Research Interests: Developmental Biology, Biology, Cell Biology, Transcription Factors, Medicine, and 15 moreBiological Sciences, Anatomy, Neural Crest, Animals, Hox Genes, DNA sequence design, Transcription Factor, Developmental, Positive Feedback, Embryonic Development, Body Patterning, Retinoic Acid, DNA sequence, Medical and Health Sciences, and Expression pattern
In this study we analysed the function of the Meinox gene prep1.1during zebrafish development. Meinox proteins form heterotrimeric complexes with Hox and Pbx members, increasing the DNA binding specificity of Hox proteins in vitro and in... more
In this study we analysed the function of the Meinox gene prep1.1during zebrafish development. Meinox proteins form heterotrimeric complexes with Hox and Pbx members, increasing the DNA binding specificity of Hox proteins in vitro and in vivo. However, a role for a specific Meinox protein in the regulation of Hox activity in vivo has not been demonstrated. In situ hybridization showed that prep1.1 is expressed maternally and ubiquitously up to 24 hours post-fertilization (hpf), and restricted to the head from 48 hpf onwards. Morpholino-induced prep1.1 loss-of-function caused significant apoptosis in the CNS. Hindbrain segmentation and patterning was affected severely, as revealed by either loss or defective expression of several hindbrain markers (foxb1.2/mariposa, krox20, pax2.1 and pax6.1), including anteriorly expressed Hox genes(hoxb1a, hoxa2 and hoxb2), the impaired migration of facial nerve motor neurons, and the lack of reticulospinal neurons (RSNs)except Mauthner cells. Furt...
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Arrhythmogenic right ventricular dysplasia/cardiomyopathy type 2 (ARVD2, OMIM 600996) and stress-induced polymorphic ventricular tachycardia (VTSIP, OMIM 604772) are two cardiac diseases causing juvenile sudden death, both associated with... more
Arrhythmogenic right ventricular dysplasia/cardiomyopathy type 2 (ARVD2, OMIM 600996) and stress-induced polymorphic ventricular tachycardia (VTSIP, OMIM 604772) are two cardiac diseases causing juvenile sudden death, both associated with mutations in the RyR2 calcium channel. By using a quantitative yeast two-hybrid system, we show that VTSIP- and ARVD2-associated point mutations influence positively and negatively, respectively, the binding of RyR2 to its gating protein FKBP12.6. These findings suggest that ARVD2 mutations increase RyR2-mediated calcium release to cytoplasm, while VTSIP mutations do not affect significantly cytosolic calcium levels, thereby explaining the clinical differences between the two diseases. The present two-hybrid system appears to be an efficient molecular tool to assay the binding of FKBP12s proteins to both cardiac RyR2 and skeletal muscle RyR1 isoforms, circumventing the full-length expression of this class of giant channels. We also provide evidence of the suitability of this system to test new drugs that target RyRs-FKBP12s interactions and do not affect yeast growth.
Research Interests: Biophysics, Chemistry, Polymorphism, Biology, Calcium, and 15 moreMedicine, Clinical and Molecular Human Genetics, Sudden Death, Humans, Calcium Channel, Biochemical, Skeletal Muscle, Gating, Protein isoforms, Protein Binding, Cardiac Disease, Full Length Movies, Biochemistry and cell biology, Hybrid System, and Medical biochemistry and metabolomics
Based on the sequence of the human endothelin-1-converting enzyme (hECE-1) cDNA, we cloned a 1982 bp cDNA fragment and we amplified by PCR a 3&#39; fragment located on the last exon (exon 19). Human metaphase chromosomes were studied... more
Based on the sequence of the human endothelin-1-converting enzyme (hECE-1) cDNA, we cloned a 1982 bp cDNA fragment and we amplified by PCR a 3&#39; fragment located on the last exon (exon 19). Human metaphase chromosomes were studied by Fluorescent in Situ Hybridization (FISH) using the 1982 digoxigenated hECE-1 fragment as a probe and chromosome 1-specific probes. Twin spot signals on each of the two homologous chromosomes 1 were found by FISH on band p36. The results of monochromosomal hybrids confirmed that hECE-1 is on chromosome 1. Radiation hybrid mapping localized the hECE-1 gene 3.15 cR far from D1S2436 (WI-3177), at about 25 cM from the telomere of the short arm, possibly at the border between 1p36.3 and 1p36.2.
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Research Interests: Developmental Biology, Biology, Cell Cycle, Immunohistochemistry, Medicine, and 14 moreSignal Transduction, Transcription, Biological Sciences, Smad 3, Animals, Spinal Cord, Neurons, Phenotype, Reporter, Transforming Growth Factor Beta, Zebrafish, Transgenic, Cell Proliferation, and Medical and Health Sciences
Research Interests: Genetics, Developmental Biology, Molecular Evolution, Immunohistochemistry, Transcription Factors, and 14 moreSignal Transduction, Biological Sciences, Cell Differentiation, Humans, Mice, Pancreas, Animals, Spinal Cord, Central Nervous System, Developmental, Zebrafish, Base Sequence, Embryos, and Medical and Health Sciences
The Helicobacter pylori Neutrophil Activating Protein (HP-NAP) is endowed with immunomodulatory properties that make it a potential candidate for anticancer therapeutic applications. By activating cytotoxic Th1 responses, HP-NAP inhibits... more
The Helicobacter pylori Neutrophil Activating Protein (HP-NAP) is endowed with immunomodulatory properties that make it a potential candidate for anticancer therapeutic applications. By activating cytotoxic Th1 responses, HP-NAP inhibits the growth of bladder cancer and enhances the anti-tumor activity of oncolytic viruses in the treatment of metastatic breast cancer and neuroendocrine tumors. The possibility that HP-NAP exerts its anti-tumor effect also by modulating the activity of innate immune cells has not yet been explored. Taking advantage of the zebrafish model, we examined the therapeutic efficacy of HP-NAP against metastatic human melanoma, limiting the observational window to 9 days post-fertilization, well before the maturation of the adaptive immunity. Human melanoma cells were xenotransplanted into zebrafish embryos and tracked in the presence or absence of HP-NAP. The behavior and phenotype of macrophages and the impact of their drug-induced depletion were analyzed ex...
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The mortality rate for malignant melanoma (MM) is very high, since it is highly invasive and resistant to chemotherapeutic treatments. The modulation of some transcription factors affects cellular processes in MM. In particular, a higher... more
The mortality rate for malignant melanoma (MM) is very high, since it is highly invasive and resistant to chemotherapeutic treatments. The modulation of some transcription factors affects cellular processes in MM. In particular, a higher expression of the osteogenic master gene RUNX2 has been reported in melanoma cells, compared to normal melanocytes. By analyzing public databases for recurrent RUNX2 genetic and epigenetic modifications in melanoma, we found that the most common RUNX2 genetic alteration that exists in transcription upregulation is, followed by genomic amplification, nucleotide substitution and multiple changes. Additionally, altered RUNX2 is involved in unchecked pathways promoting tumor progression, Epithelial Mesenchymal Transition (EMT), and metastasis. In order to investigate further the role of RUNX2 in melanoma development and to identify a therapeutic target, we applied the CRISPR/Cas9 technique to explore the role of the RUNT domain of RUNX2 in a melanoma ce...
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1 University of Florence, Department of Neuroscience, Psychology, Drug Research and Child Health, Viale Pieraccini 6, Florence, Italy, 50139 2 European Laboratory for Non-Linear Spectroscopy, Via Nello Carrara 1, Sesto Fiorentino, Italy,... more
1 University of Florence, Department of Neuroscience, Psychology, Drug Research and Child Health, Viale Pieraccini 6, Florence, Italy, 50139 2 European Laboratory for Non-Linear Spectroscopy, Via Nello Carrara 1, Sesto Fiorentino, Italy, 50019 3 University of Florence, Department of Physics and Astronomy, Via Sansone 1, Sesto Fiorentino, Italy, 50019 4 School of Physics and Astronomy, Kelvin Building, University of Glasgow, G12 8QQ, Glasgow, UK 5 National Institute of Optics, National Research Council, Via Nello Carrara 1, Sesto Fiorentino, Italy, 50019 6 University of Padova, Department of Biology, Via U. Bassi 58/B, Padua, Italy, 35131 7 University of Florence, Department of Biology, Via Madonna del Piano 6, Sesto Fiorentino, Italy, 50019
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Deciphering the workings of a brain (and its alterations due to pathologies and disorders) requires real-time mapping of the activity of all neurons. Recently, this formidable task has been brought into reach by the implementation of... more
Deciphering the workings of a brain (and its alterations due to pathologies and disorders) requires real-time mapping of the activity of all neurons. Recently, this formidable task has been brought into reach by the implementation of light sheet fluorescence (LSF) microscopy on the transparent zebrafish larva, a simple yet representative model of the vertebrate brain. This application is however hindered by the use of visible light for fluorescence excitation, inducing unwanted visual stimulation that can only be partially overcome resorting to complex excitation geometries. On the other hand, two-photon (2P) LSF microscopy, owing to the use of invisible infra-red illumination, enables volumetric imaging of neuronal activity avoiding undesirable visual stimulation. However, due to the low efficiency of the 2P absorption process, the imaging speed of this technique is typically limited (~ 1 Hz) by the signal to noise ratio achievable without sample photodamage. Here we describe a cus...
Glucocorticoids (GCs) are steroid hormones that contribute to the regulation of many physiological processes, such as inflammation, metabolism and stress response, mainly through binding to their cognate receptor, GR, which works as a... more
Glucocorticoids (GCs) are steroid hormones that contribute to the regulation of many physiological processes, such as inflammation, metabolism and stress response, mainly through binding to their cognate receptor, GR, which works as a ligand-activated transcription factor. Due to their pleiotropy and the common medical use of these steroids to treat patients affected by different pathologies, the investigation of their mechanisms of action is extremely important in biology and clinical research. The evolutionary conservation of GC physiological function, biosynthesis pathways, as well as the sequence and structure of the GC nuclear receptors has stimulated, in the last 20 years, the use of zebrafish (a teleost of Cyprinidae family) as a reliable model organism to investigate this topic. In this review, we wanted to collect many of the most significant findings obtained by the the scientific community using zebrafish to study GCs and their receptors. The paper begins by describing th...
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Drug resistance mechanisms still characterize metastatic melanoma, despite new treatments have been recently developed. Targeting of the cGMP/protein kinase G (PKG) pathway is emerging as a therapeutic approach in cancer research. In this... more
Drug resistance mechanisms still characterize metastatic melanoma, despite new treatments have been recently developed. Targeting of the cGMP/protein kinase G (PKG) pathway is emerging as a therapeutic approach in cancer research. In this study, we evaluated the anticancer effects of two polymeric linked dimeric (PDL) cGMP analogues able to bind and activate PKG, called PKG-activator (PA) 4 and 5. PA5 was identified as the most effective compound on melanoma cell lines as well as on patient-derived metastatic melanoma cells cultured as three-dimensional spheroids and in a zebrafish melanoma model. PA5 was able to significantly reduce cell viability, size, and invasion of melanoma spheroids. Importantly, PA5 showed a tumor specific outcome because no toxic effect was observed in healthy melanocytes exposed to the cGMP analogue. We defined that, by triggering PKG, PA5 interfered with the EGF pathway as shown by lower EGFR phosphorylation and reduction of activated, phosphorylated forms of AKT and ERK1/2 in melanoma cells. Finally, PA5 significantly reduced the metastatic process in zebrafish. These studies open future perspectives for the cGMP analogue PA5 as a potential therapeutic strategy for melanoma.
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We present the development of a custom-made two-photon light-sheet microscope optimized for high-speed (5 Hz) volumetric imaging of zebrafish larval brain for the analysis of neuronal physiological and pathological activity. High-speed... more
We present the development of a custom-made two-photon light-sheet microscope optimized for high-speed (5 Hz) volumetric imaging of zebrafish larval brain for the analysis of neuronal physiological and pathological activity. High-speed volumetric two-photon light-sheet microscopy is challenging to achieve, due to constrains on the signal-to-noise ratio. To maximize this parameter, we optimized our setup for high peak power of excitation light, while finely controlling its polarization, and we implemented remote scanning of the focal plane to record without disturbing the sample. Two-photon illumination is advantageous for zebrafish larva studies since infra-red excitation does not induce a visual response, that otherwise would affect the neuronal activity. In particular, we were able to record whole-brain neuronal activity of the larva with high temporal- and spatial-resolution during the nocturnal period without affecting the circadian rhythm. Analyzing the spatially resolved power spectra of GCaMP signal, we found significant differences for several frequency bands between the day/night phases in various brain regions. Moreover, we studied the fast dynamics that characterize the acutely induced pathological epileptic activity of the larvae, identifying the brain structures that are more susceptible to the action of the epileptogenic drug. In conclusion, the high speed two-photon light-sheet microscope that we developed is proving to be an important tool to study both the physiological and the pathological activity of the zebrafish larval brain without undesired visual stimulation.
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While endothelial cell (EC) function is influenced by mitochondrial metabolism, the role of mitochondrial dynamics in angiogenesis, the formation of new blood vessels from existing vasculature, is unknown. Here we show that the inner... more
While endothelial cell (EC) function is influenced by mitochondrial metabolism, the role of mitochondrial dynamics in angiogenesis, the formation of new blood vessels from existing vasculature, is unknown. Here we show that the inner mitochondrial membrane mitochondrial fusion protein optic atrophy 1 (OPA1) is required for angiogenesis. In response to angiogenic stimuli, OPA1 levels rapidly increase to limit nuclear factor kappa-light-chain-enhancer of activated B cell (NFκB) signaling, ultimately allowing angiogenic genes expression and angiogenesis. Endothelial Opa1 is indeed required in an NFκB-dependent pathway essential for developmental and tumor angiogenesis, impacting tumor growth and metastatization. A first-in-class small molecule-specific OPA1 inhibitor confirms that EC Opa1 can be pharmacologically targeted to curtail tumor growth. Our data identify Opa1 as a crucial component of physiological and tumor angiogenesis.
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Precise three-dimensional mapping of a large number of gene expression patterns, neuronal types and connections to an anatomical reference helps us to understand the vertebrate brain and its development. Zebrafish has evolved as a model... more
Precise three-dimensional mapping of a large number of gene expression patterns, neuronal types and connections to an anatomical reference helps us to understand the vertebrate brain and its development. Zebrafish has evolved as a model organism for such study. In this paper, we propose a novel non-rigid registration algorithm for volumetric zebrafish larval image datasets. A coarse affine registration using the L-BFGS algorithm is applied first on the moving dataset. We then divide this coarsely registered moving image and the reference image into a union of overlapping patches. Minimum weight bipartite graph matching algorithm is employed to find the correspondence between the two sets of patches. The corresponding patches are then registered using the diffeomorphic demons method with proper intra-patch regularization. For each voxel lying in the overlapping regions, we impose inter-patch regularization through a composite transformation obtained from the adjacent transformation f...
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The DNA polymerase gamma (Polg) is a nuclear-encoded enzyme involved in DNA replication in animal mitochondria. In humans, mutations in the POLG gene underlie a set of mitochondrial diseases characterized by mitochondrial DNA (mtDNA)... more
The DNA polymerase gamma (Polg) is a nuclear-encoded enzyme involved in DNA replication in animal mitochondria. In humans, mutations in the POLG gene underlie a set of mitochondrial diseases characterized by mitochondrial DNA (mtDNA) depletion or deletion and multiorgan defects, named POLG disorders, for which an effective therapy is still needed. By applying antisense strategies, ENU- and CRISPR/Cas9-based mutagenesis, we have generated embryonic, larval-lethal and adult-viable zebrafish Polg models. Morphological and functional characterizations detected a set of phenotypes remarkably associated to POLG disorders, including cardiac, skeletal muscle, hepatic and gonadal defects, as well as mitochondrial dysfunctions and, notably, a perturbed mitochondria-to-nucleus retrograde signaling (CREB and Hypoxia pathways). Next, taking advantage of preliminary evidence on the candidate molecule Clofilium tosylate (CLO), we tested CLO toxicity and then its efficacy in our zebrafish lines. In...
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RUNX2, a master osteogenic transcript ion factor, is overexpressed in several cancer cells; in melanoma it promotes cells migration and invasion as well as neoangiogenesis. The annual mortality rates related to metastatic melanoma are... more
RUNX2, a master osteogenic transcript ion factor, is overexpressed in several cancer cells; in melanoma it promotes cells migration and invasion as well as neoangiogenesis. The annual mortality rates related to metastatic melanoma are high and novel agents are needed to improve melanoma patients’ survival. It has been shown that lectins specifically target malignant cells since they present the Thomsen–Friedenreich antigen. This disaccharide is hidden in normal cells, while it allows selective lectins binding in transformed cells. Recently, an edible lectin named BEL β-trefoil has been obtained from the wild mushroom Boletus edulis. Our previous study showed BEL β-trefoil effects on transcription factor RUNX2 downregulation as well as on the migration ability in melanoma cells treated in vitro. Therefore, to better understand the role of this lectin, we investigated the BEL β-trefoil effects in a zebrafish in vivo model, transplanted with human melanoma cells expressing RUNX2. Our d...
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An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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The mortality rate for malignant melanoma (MM) is very high, since it is highly invasive and resistant to chemotherapeutic treatments. The modulation of some transcription factors affects cellular processes in MM. In particular, a higher... more
The mortality rate for malignant melanoma (MM) is very high, since it is highly invasive and resistant to chemotherapeutic treatments. The modulation of some transcription factors affects cellular processes in MM. In particular, a higher expression of the osteogenic master gene RUNX2 has been reported in melanoma cells, compared to normal melanocytes. By analyzing public databases for recurrent RUNX2 genetic and epigenetic modifications in melanoma, we found that the most common RUNX2 genetic alteration that exists in transcription upregulation is, followed by genomic amplification, nucleotide substitution and multiple changes. Additionally, altered RUNX2 is involved in unchecked pathways promoting tumor progression, Epithelial Mesenchymal Transition (EMT), and metastasis. In order to investigate further the role of RUNX2 in melanoma development and to identify a therapeutic target, we applied the CRISPR/Cas9 technique to explore the role of the RUNT domain of RUNX2 in a melanoma ce...
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Familial aggregation is a significant risk factor for the development of thyroid cancer and familial non-medullary thyroid cancer (FNMTC) accounts for 5-7% of all NMTC. Whole exome sequencing analysis in the family affected by FNMTC with... more
Familial aggregation is a significant risk factor for the development of thyroid cancer and familial non-medullary thyroid cancer (FNMTC) accounts for 5-7% of all NMTC. Whole exome sequencing analysis in the family affected by FNMTC with oncocytic features where our group previously identified a predisposing locus on chromosome 19p13.2, revealed a novel heterozygous mutation (c.400G > A, NM_012335; p.Gly134Ser) in exon 5 of MYO1F, mapping to the linkage locus. In the thyroid FRTL-5 cell model stably expressing the mutant MYO1F p.Gly134Ser protein, we observed an altered mitochondrial network, with increased mitochondrial mass and a significant increase in both intracellular and extracellular reactive oxygen species, compared to cells expressing the wild-type (wt) protein or carrying the empty vector. The mutation conferred a significant advantage in colony formation, invasion and anchorage-independent growth. These data were corroborated by in vivo studies in zebrafish, since we ...
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Type 2 diabetes (T2D) is a disease characterized by impaired insulin secretion. The Wnt signaling transcription factor Tcf7l2 is to date the T2D-associated gene with the largest effect on disease susceptibility. However, the mechanisms by... more
Type 2 diabetes (T2D) is a disease characterized by impaired insulin secretion. The Wnt signaling transcription factor Tcf7l2 is to date the T2D-associated gene with the largest effect on disease susceptibility. However, the mechanisms by which TCF7L2 variants affect insulin release from β-cells are not yet fully understood. By taking advantage of a tcf7l2 zebrafish mutant line, we first show that these animals are characterized by hyperglycemia and impaired islet development. Moreover, we demonstrate that the zebrafish tcf7l2 gene is highly expressed in the exocrine pancreas, suggesting potential bystander effects on β-cell growth, differentiation and regeneration. Finally, we describe a peculiar vascular phenotype in tcf7l2 mutant larvae, characterized by significant reduction in the average number and diameter of pancreatic islet capillaries. Overall, the zebrafish Tcf7l2 mutant, characterized by hyperglycemia, pancreatic and vascular defects, and reduced regeneration proves to b...
The development of functional peripheral ganglia requires a balance of specification of both neuronal and glial components. In the developing dorsal root ganglia (DRGs), these components form from partially-restricted bipotent neuroglial... more
The development of functional peripheral ganglia requires a balance of specification of both neuronal and glial components. In the developing dorsal root ganglia (DRGs), these components form from partially-restricted bipotent neuroglial precursors derived from the neural crest. Work in mouse and chick has identified several factors, including Delta/Notch signaling, required for specification of a balance of these components. We have previously shown in zebrafish that the Sry-related HMG domain transcription factor, Sox10, plays an unexpected, but crucial, role in sensory neuron fate specification in vivo. In the same study we described a novel Sox10 mutant allele, sox10baz1, in which sensory neuron numbers are elevated above those of wild-types. Here we investigate the origin of this neurogenic phenotype. We demonstrate that the supernumerary neurons are sensory neurons, and that enteric and sympathetic neurons are almost absent just as in classical sox10 null alleles; peripheral g...
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Mutations in Pantothenate kinase 2 and Coenzyme A (CoA) synthase (COASY), genes involved in CoA biosynthesis, are associated with rare neurodegenerative disorders with brain iron accumulation. We showed that zebrafish pank2 gene plays an... more
Mutations in Pantothenate kinase 2 and Coenzyme A (CoA) synthase (COASY), genes involved in CoA biosynthesis, are associated with rare neurodegenerative disorders with brain iron accumulation. We showed that zebrafish pank2 gene plays an essential role in brain and vasculature development. Now we extended our study to coasy. The gene has high level of sequence identity with the human ortholog and is ubiquitously expressed from the earliest stages of development. The abrogation of its expression led to strong reduction of CoA content, high lethality and a phenotype resembling to that of dorsalized mutants. Lower doses of morpholino resulted in a milder phenotype, with evident perturbation in neurogenesis and formation of vascular arborization; the dorso-ventral patterning was severely affected, the expression of bone morphogenetic protein (Bmp) receptors and activity were decreased, while cell death increased. These features specifically correlated with the block in CoA biosynthesis ...
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CD271 is a neurotrophin receptor variably expressed in melanoma. While contradictory data are reported on its role as a marker of tumor initiating cells, little is known on its function in tumor progression. CD271 expression was higher in... more
CD271 is a neurotrophin receptor variably expressed in melanoma. While contradictory data are reported on its role as a marker of tumor initiating cells, little is known on its function in tumor progression. CD271 expression was higher in spheroids derived from freshly isolated cells of primary melanomas and in primary WM115 and WM793-B cell lines, while it decreased during progression to advanced stages in cells isolated from metastatic melanomas and in metastatic WM266-4 and 1205Lu cell lines. Moreover, CD271 was scarcely detected in the highly invasive spheroids (SKMEL28 and 1205Lu). CD271, originally expressed in the epidermis of skin reconstructs, disappeared when melanoma started to invade the dermis. SKMEL8 CD271(-) cells showed greater proliferation and invasiveness in vitro, and were associated with a higher number of metastases in zebrafish, as compared to CD271(+) cells. CD271 silencing in WM115 induced a more aggressive phenotype in vitro and in vivo. On the contrary, CD...
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... Beis d , 1 ,; Claudia Wierzbicki e ,; Rodrigo M. Young e ,; Despina Bournele d ,; Alice Domenichini f ,; Leonardo E. Valdivia e ,; Lawrence Lum g ,; ... Weidinger et al., 2005) and Tg(hsp70l:dkk1-GFP) w32 (Stoick-Cooper et al., 2007)... more
... Beis d , 1 ,; Claudia Wierzbicki e ,; Rodrigo M. Young e ,; Despina Bournele d ,; Alice Domenichini f ,; Leonardo E. Valdivia e ,; Lawrence Lum g ,; ... Weidinger et al., 2005) and Tg(hsp70l:dkk1-GFP) w32 (Stoick-Cooper et al., 2007) Tg(fli1a:EGFP) y1 (Lawson and Weinstein, 2002), Tg ...
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The members of the Olig gene family encode for basic helix-loop-helix (bHLH) transcription factors involved in neural cell type specification. Three Olig genes (Olig1, Olig2 and Olig3) have been identified in all known vertebrate models... more
The members of the Olig gene family encode for basic helix-loop-helix (bHLH) transcription factors involved in neural cell type specification. Three Olig genes (Olig1, Olig2 and Olig3) have been identified in all known vertebrate models and a fourth one in anamniotes (olig4). Here we have performed a global analysis of olig genes during zebrafish embryonic development and determined which signaling pathways control their induction and regionalization in the CNS. Interestingly, zebrafish olig3 and olig4 together establish most of the expression domains corresponding to mouse Olig3. According to our data, olig1 is specifically confined to the oligodendrocyte lineage, whereas the other members display stratified expression in diencephalon, hindbrain, and spinal cord. We observed differential expression of olig genes within specific motoneuron and interneuron domains of the spinal cord. olig2, olig3, and olig4 expression appears to be regulated by nodal and FGF signaling during gastrulation and early somitogenesis, by RA signaling in the hindbrain, and by BMP and Hh signals along the dorsoventral axis of the embryonic CNS. Our findings suggest a role for olig genes in CNS patterning as well as in multiple cell fate decisions during neural differentiation.
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Research Interests: Skeletal muscle biology, Immunohistochemistry, Sequence Analysis, Western blotting, Cell line, and 15 moreHumans, Molecular cloning, Fluorescent Antibody Technique, Skeletal Muscle, Myocardium, Western blot, Protein Expression, Amino Acid Profile, Amino Acid Sequence, Base Sequence, Open Reading Frame, Biochemistry and cell biology, Gene Expression Regulation, Molecular Sequence Data, and cDNA cloning
During the development of the central nervous system, the proliferation of neural progenitors and differentiation of neurons and glia are tightly regulated by different transcription factors and signaling cascades, such as the Wnt and Shh... more
During the development of the central nervous system, the proliferation of neural progenitors and differentiation of neurons and glia are tightly regulated by different transcription factors and signaling cascades, such as the Wnt and Shh pathways. This process takes place in cooperation with several microRNAs, some of which evolutionarily conserved in vertebrates, from teleosts to mammals. We focused our attention on miR-7, as its role in the regulation of cell signaling during neural development is still unclear. Specifically, we used human stem cell cultures and whole zebrafish embryos to study, in vitro and in vivo, the role of miR-7 in the development of dopaminergic (DA) neurons, a cell type primarily affected in Parkinson’s disease. We demonstrated that the zebrafish homologue of miR-7 (miR-7a) is expressed in the forebrain during the development of DA neurons. Moreover, we identified 143 target genes downregulated by miR-7, including the neural fate markers TCF4 and TCF12, a...
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During the development of the central nervous system, the proliferation of neural progenitors and differentiation of neurons and glia are tightly regulated by different transcription factors and signaling cascades, such as the Wnt and Shh... more
During the development of the central nervous system, the proliferation of neural progenitors and differentiation of neurons and glia are tightly regulated by different transcription factors and signaling cascades, such as the Wnt and Shh pathways. This process takes place in cooperation with several microRNAs, some of which evolutionarily conserved in vertebrates, from teleosts to mammals. We focused our attention on miR-7, as its role in the regulation of cell signaling during neural development is still unclear. Specifically, we used human stem cell cultures and whole zebrafish embryos to study, in vitro and in vivo, the role of miR-7 in the development of dopaminergic (DA) neurons, a cell type primarily affected in Parkinson’s disease. We demonstrated that the zebrafish homologue of miR-7 (miR-7a) is expressed in the forebrain during the development of DA neurons. Moreover, we identified 143 target genes downregulated by miR-7, including the neural fate markers TCF4 and TCF12, a...
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Research Interests: Electrophysiology, Sequence Analysis, Danio rerio, In Situ Hybridization, Phylogeny, and 18 moreXenopus, Female, Animals, Vertebrates, Male, Physiological, Medical Physiology, Inner Ear, Membrane transport proteins, Genome Organization, Larva, Zebrafish, Amino Acid Sequence, Base Sequence, Oocytes, Expression analysis, Physiological Genomics, and Functional Properties
Research Interests: Electrophysiology, Sequence Analysis, Danio rerio, In Situ Hybridization, Phylogeny, and 18 moreXenopus, Female, Animals, Vertebrates, Male, Physiological, Medical Physiology, Inner Ear, Membrane transport proteins, Genome Organization, Larva, Zebrafish, Amino Acid Sequence, Base Sequence, Oocytes, Expression analysis, Physiological Genomics, and Functional Properties
OBJECTIVES We sought to establish the role of genetic screening for ryanodine receptor type 2 (RyR2) gene mutations in families with effort-induced polymorphic ventricular arrhythmia (PVA), syncope and juvenile sudden death. BACKGROUND... more
OBJECTIVES We sought to establish the role of genetic screening for ryanodine receptor type 2 (RyR2) gene mutations in families with effort-induced polymorphic ventricular arrhythmia (PVA), syncope and juvenile sudden death. BACKGROUND The RyR2 mutations have been associated with PVA, syncope and sudden death in response to physical or emotional stress. METHODS We studied 81 subjects (39 males and 42
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We present the Human Muscle Gene Map (HMGM), the first comprehensive and updated high-resolution expression map of human skeletal muscle. The 1078 entries of the map were obtained by merging data retrieved from UniGene with the RH mapping... more
We present the Human Muscle Gene Map (HMGM), the first comprehensive and updated high-resolution expression map of human skeletal muscle. The 1078 entries of the map were obtained by merging data retrieved from UniGene with the RH mapping information on 46 novel muscle transcripts, which showed no similarity to any known sequence. In the map, distances are expressed in megabase pairs. About one-quarter of the map entries represents putative novel genes. Genes known to be specifically expressed in muscle account for <4% of the total. The genomic distribution of the map entries confirmed the previous finding that muscle genes are selectively concentrated in chromosomes 17, 19, and X. Five chromosomal regions are suspected to have a significant excess of muscle genes. Present data support the hypothesis that the biochemical and functional properties of differentiated muscle cells may result from the transcription of a very limited number of muscle-specific genes along with the activ...
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Research Interests: Genetics, Developmental Biology, Biological Sciences, Cell Differentiation, Molecular and cellular biology, and 14 moreThyroid gland, Endocrine disruptors, Animals, Drug Screening, Thyroid Hormones, Early development, In Vivo Imaging, Behavioral Animal Models, Embryonic Development, Zebrafish, Endocrine Disruptors, Large Scale, Power Modeling, and Embryos
Research Interests: Polymorphism, Adolescent, Clinical and Molecular Human Genetics, Electrocardiography, Echocardiography, and 19 moreSudden Death, Humans, Child, Genetic Testing, Mutation, Haplotypes, Female, Male, Ryanodine Receptor, Genetic Screening, The, Pedigree, Middle Aged, Adult, Public health systems and services research, Exercise Test, Ventricular Arrhythmia, Syncope, and The American
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By sequencing 11,405 individual expressed sequence tags (ESTs) from a cDNA library of a human skeletal muscle, we identified 1945 individual transcripts, 725 of which showed no correspondence with known human genes. We report here the... more
By sequencing 11,405 individual expressed sequence tags (ESTs) from a cDNA library of a human skeletal muscle, we identified 1945 individual transcripts, 725 of which showed no correspondence with known human genes. We report here the chromosomal localization of 267 of these, obtained by radiation hybrid (RH) mapping. The map position of additional 242 ESTs from the same library, corresponding to known human genes, is also reported. The resulting information provides a preliminary genomic transcriptional profile of a human muscle. Several genes occur in clusters on different chromosomes. Moreover, chromosomes 17, 19, 21 and X appear to be significantly rich in muscle ESTs. By analysing several collections of ESTs from different tissues, we observed significant deviations in the distribution of ESTs by chromosome in fetal heart, adult brain and adult retina, supporting the hypothesis that a non-random localization of genes expressed in specific tissues might not be uncommon. The selective concentration of expressed genes in some chromosomes and in specific chromosomal subregions in a given tissue might reflect the existence of batteries of genes under the same control mechanisms, regulating tissue-specific gene expression.
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Autosomal dominant arrhythmogenic right ventricular cardiomyopathy (ARVD, MIM 107970) is one of the major causes of juvenile sudden death. We have previously assigned the disease locus to chromosome 14q23-q24. Here we report on a novel... more
Autosomal dominant arrhythmogenic right ventricular cardiomyopathy (ARVD, MIM 107970) is one of the major causes of juvenile sudden death. We have previously assigned the disease locus to chromosome 14q23-q24. Here we report on a novel variant of ARVD, which is transmitted associated to 1q42-q43 and is characterized by a concealed form, showing effort-induced polymorphic tachycardias. Since both loci ARVD1 and ARVD2 map in proximity of alpha-actinin genes, the possible implication of these myofibrillar proteins in the pathogenesis of ARVD is discussed. Two additional ARVD families, tested with markers of chromosomes 1q42-q43 and 14q23-q24, failed to show linkage, providing evidence of further genetic heterogeneity.
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Autosomal dominant arrhythmogenic right ventricular dysplasia (ARVD; MIM 107970) is a genetically heterogeneous cardiomyopathy, which often causes sudden death in juveniles and athletes. Two disease loci were previously mapped... more
Autosomal dominant arrhythmogenic right ventricular dysplasia (ARVD; MIM 107970) is a genetically heterogeneous cardiomyopathy, which often causes sudden death in juveniles and athletes. Two disease loci were previously mapped respectively to 14q23-q24 (ARVD1) and to 1q42-q43 (ARVD2). A third possible locus was assigned to 14q12-q22. We report here on a linkage study performed on three independent families with recurrence of ARVD characterized by localized involvement of the left ventricle. In these families the disease appears to be transmitted with three polymorphic DNA markers of the chromosome 2 long arm, showing a maximum lod score of 3.46 at theta = 0 for the marker D2S152. The multipoint linkage analysis suggests that the novel ARVD locus, provisionally named ARVD4, maps to 2q32. 1-q32.3, within the chromosomal region including markers D2S152, D2S103, and D2S389.
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The chromosome assignment of 115 expressed sequence tags (ESTs) from human skeletal muscle, 101 of which identify unknown human genes, is reported. The ESTs were selected among over 4,000 obtained from systematic sequencing of a skeletal... more
The chromosome assignment of 115 expressed sequence tags (ESTs) from human skeletal muscle, 101 of which identify unknown human genes, is reported. The ESTs were selected among over 4,000 obtained from systematic sequencing of a skeletal muscle cDNA library containing 3&#39; portions of the mRNAs. Chromosome assignments were obtained by PCR amplification of two panels of human x rodent somatic cell hybrids. Analysis of these preliminary data suggests a nonrandom distribution of muscle ESTs in the human chromosome complement. The unexpected occurrence of multiple chromosome localizations for some ESTs is discussed.
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Research Interests: Genetics, Adolescent, Sudden Death, Cardiac arrest, Humans, and 19 moreChild, Female, Male, Polymerase Chain Reaction, Ryanodine Receptor, Pedigree, Phenotype, Clinical Sciences, Peripheral blood lymphocyte, Adult, Public health systems and services research, Degeneration, Circulation, Arrhythmia, Base Sequence, Catecholamines, Ventricular Tachycardia, Catecholaminergic polymorphic ventricular tachycardia, and Intracellular Calcium
Familial polymorphic ventricular tachycardia is an autosomal-dominant, inherited disease with a relatively early onset and a mortality rate of approximately 30% by the age of 30 years. Phenotypically, it is characterized by salvoes of... more
Familial polymorphic ventricular tachycardia is an autosomal-dominant, inherited disease with a relatively early onset and a mortality rate of approximately 30% by the age of 30 years. Phenotypically, it is characterized by salvoes of bidirectional and polymorphic ventricular tachycardias in response to vigorous exercise, with no structural evidence of myocardial disease. We previously mapped the causative gene to chromosome 1q42-q43. In the present study, we demonstrate that patients with familial polymorphic ventricular tachycardia have missense mutations in the cardiac sarcoplasmic reticulum calcium release channel (ryanodine receptor type 2 [RyR2]). In 3 large families studied, 3 different RyR2 mutations (P2328S, Q4201R, V4653F) were detected and shown to fully cosegregate with the characteristic arrhythmic phenotype. These mutations were absent in the nonaffected family members and in 100 healthy controls. In addition to identifying 3 causative mutations, we identified a number of single nucleotide polymorphisms that span the genomic structure of RyR2 and will be useful for candidate-based association studies for other arrhythmic disorders. Our data illustrate that mutations of the RyR2 gene cause at least one variety of inherited polymorphic tachycardia. These findings define a new entity of disorders of myocardial calcium signaling.
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Members of the ICE/CED-3 protease family appear to play an essential role in programmed cell death process. In this paper the chromosomal localization of the human genes CPP32, Mch2, Mch3 and Ich-1 is reported, obtained by Radiation... more
Members of the ICE/CED-3 protease family appear to play an essential role in programmed cell death process. In this paper the chromosomal localization of the human genes CPP32, Mch2, Mch3 and Ich-1 is reported, obtained by Radiation Hybrid Mapping. CPP32 was assigned to chromosome 4q33-q35.1, Mch2 to chromosome 4q25-q26, Mch3 to chromosome 10q25.1-q25.2 and Ich-1 to chromosome 7q35. Ich-1 was found to map very close to the marker WI-9353. The possible overlapping of the two independent locus assignments is considered. The genomic distribution of these genes is discussed, with particular reference to the co-location with some human genetic diseases all characterized by autosomal dominant inheritance and by similar malformative features.
Research Interests: Human Genetics, Apoptosis, Clinical and Molecular Human Genetics, Caspases, Humans, and 11 moreAnimals, Polymerase Chain Reaction, Programmed cell death, Biochemical, Proteins, Caspase, Hybrid Solar Cells, Cysteine endopeptidases, Base Sequence, Congenital abnormalities, and Biochemistry and cell biology
The present paper reports on the fine mapping of the ACTN2 gene and on the reconstruction of its genomic structure. By radiation hybrid mapping, the gene was located about 912 cR from the 1p-telomere. ACTN2 was placed between the marker... more
The present paper reports on the fine mapping of the ACTN2 gene and on the reconstruction of its genomic structure. By radiation hybrid mapping, the gene was located about 912 cR from the 1p-telomere. ACTN2 was placed between the marker WI-9317 (alias D1S2421) and the marker AFMA045ZC5, within the chromosomal band 1q43. The gene was detected in YAC 955 c 12. This YAC was used as template DNA for long-distance and Alu-PCR, using a set of putative exonic primers, designed on the cDNA sequence of alpha-actinin-2, in order to characterize the ACTN2 intron-exon boundaries. The entire genomic structure of the gene was reconstructed. The ACTN2 gene contained 21 exons, in a segment spanning about 40 kb of genomic DNA. Only the proximal part of the gene shows a high conservation through evolution, whereas in the remaining part a divergence from the genomic organization of C. elegans and D. melanogaster was noticed. A series of intronic primers was specifically designed and produced, to amplify all the exons of ACTN2, directly from genomic DNA. This will enable mutation screening in patients affected with hereditary diseases linked to the marker CA4F/R, a polymorphism in the last intron of the alpha-actinin-2 gene.
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In this paper the chromosomal localization of the human skeletal muscle genes Troponin-I slow-twitch (TNNI1), Troponin-I fast-twitch (TNNI2), and Troponin-C fast (TNNC2) and the refinement of the position for alpha-Tropomyosin (TPM1) and... more
In this paper the chromosomal localization of the human skeletal muscle genes Troponin-I slow-twitch (TNNI1), Troponin-I fast-twitch (TNNI2), and Troponin-C fast (TNNC2) and the refinement of the position for alpha-Tropomyosin (TPM1) and beta-Tropomyosin (TPM2) are reported. By radiation hybrid mapping, TPM1 was assigned to chromosome 15q22.1, TPM2 to chromosome 9p13.2-p13.1, TNNI1 to chromosome 1q31.3, TNNI2 to chromosome 11p15.5, and TNNC2 to chromosome 20q12-q13.11. The genomic distribution of these genes is discussed, with particular emphasis on the cluster organization of the Troponin genes.
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In the present study we report on another cause of an arrhythmia associated with familial arrhythmogenic right ventricular cardiomyopathy (ARVC), which is linked to chromosome 1q42-43. Two families with 48 subjects were studied with... more
In the present study we report on another cause of an arrhythmia associated with familial arrhythmogenic right ventricular cardiomyopathy (ARVC), which is linked to chromosome 1q42-43. Two families with 48 subjects were studied with 12-lead electrocardiography, 24-hour ambulatory electrocardiography, chest x-ray, M-mode and 2-dimensional echocardiography, signal-averaging electrocardiography, and exercise stress testing. Six subjects also underwent right and left ventricular angiography and electrophysiologic study. An endomyocardial biopsy was performed in 1 subject. The genetic study included pedigree reconstruction and linkage analysis with polymorphic DNA markers. Five young subjects died suddenly during exercise; autopsy was performed in 3 and showed segmental fibro-fatty replacement of the right ventricle, mostly at the apex. Two of them experienced syncopal attacks during effort. Sixteen living subjects, without arrhythmias at rest had polymorphic ventricular arrhythmias during effort; ARVC was diagnosed in 15, whereas 1 did not have any demonstrable cardiac abnormality. The remaining family members were healthy and did not have arrhythmias. The linkage study assigned the disease locus to chromosome 1q42-q43, in close proximity to the alpha-actinin 2 locus (maximal lod score was 5.754 at theta = 0) with a 95% penetrance. Thus, these data suggest that effort-induced polymorphic ventricular arrhythmias and juvenile sudden death can be due to adrenergic stimulation in a particular genetic group of ARVC patients. In these cases the pathology was segmental, mostly localized to the right ventricular apex. Ventricular arrhythmias that are present in these families differ from the monomorphic ones that are usually seen in patients with ARVC.
Research Interests: Genetics, Polymorphism, Clinical and Molecular Human Genetics, Electrocardiography, Echocardiography, and 23 moreSudden Death, Humans, Left Ventricular Assist Device, Female, Male, Exercise, Pedigree, Genetic linkage analysis, Dimensional, Stress Testing, Adult, Myocardium, Sudden Cardiac Death, Arrhythmia, Right Ventricle, Exercise Test, Genetic Markers, Ventricular Arrhythmia, Linkage Analysis, Syncope, DNA marker, The American, and Chest X-ray
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Research Interests: Skeletal muscle biology, Immunohistochemistry, Sequence Analysis, Western blotting, Cell line, and 13 moreHumans, Molecular cloning, Fluorescent Antibody Technique, Skeletal Muscle, Myocardium, Western blot, Protein Expression, Amino Acid Profile, Amino Acid Sequence, Base Sequence, Open Reading Frame, Biochemistry and cell biology, and Gene Expression Regulation
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Research Interests: Polymorphism, Calcium, Clinical and Molecular Human Genetics, Sudden Death, Humans, and 11 moreCalcium Channel, Biochemical, Skeletal Muscle, Yeast Two-hybrid System, Protein isoforms, Protein Binding, Ventricular Tachycardia, Cardiac Disease, Full Length Movies, Biochemistry and cell biology, and Hybrid System
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Pancreatic adenocarcinoma, one of the worst malignancies of the exocrine pancreas, is a solid tumor with increasing incidence and mortality in industrialized countries. This condition is usually driven by oncogenic KRAS point mutations... more
Pancreatic adenocarcinoma, one of the worst malignancies of the exocrine pancreas, is a solid tumor with increasing incidence and mortality in industrialized countries. This condition is usually driven by oncogenic KRAS point mutations and evolves into a highly aggressive metastatic carcinoma due to secondary gene mutations and unbalanced expression of genes involved in the specific signaling pathways. To examine in vivo the effects of KRAS(G12D) during pancreatic cancer progression and time correlation with cancer signaling pathway activities, we have generated a zebrafish model of pancreatic adenocarcinoma in which eGFP-KRAS(G12D) expression was specifically driven to the pancreatic tissue by using the GAL4/UAS conditional expression system. Outcrossing the inducible oncogenic KRAS(G12D) line with transgenic zebrafish reporters, harboring specific signaling responsive elements of transcriptional effectors, we were able to follow TGFβ, Notch, Bmp and Shh activities during tumor dev...