The Ran GTPase plays a central function in control of nucleo-cytoplasmic transport in interphase.... more The Ran GTPase plays a central function in control of nucleo-cytoplasmic transport in interphase. Mitotic roles of Ran have also been firmly established in Xenopus oocyte extracts. In this system, Ran-GTP, or the RCC1 exchange factor for Ran, drive spindle assembly by regulating the availability of `aster-promoting activities'. In previous studies to assess whether the Ran network also influences mitosis in mammalian cells, we found that overexpression of Ran-binding protein 1 (RanBP1), a major effector of Ran, induces multipolar spindles. We now show that these abnormal spindles are generated through loss of cohesion in mitotic centrosomes. Specifically,RanBP1 excess induces splitting of mother and daughter centrioles at spindle poles; the resulting split centrioles can individually organize functional microtubule arrays, giving rise to functional spindle poles. RanBP1-dependent centrosome splitting is specifically induced in mitosis and requires microtubule integrity and Eg5 a...
Human lymphoblastoid cells of normal origin and from genetic instability syndromes, i.e. Fanconi ... more Human lymphoblastoid cells of normal origin and from genetic instability syndromes, i.e. Fanconi anemia (FA) group C and ataxia telangectasia, were continuously exposed to extremely low frequency magnetic field (ELF‐MF). We report that ELF‐MF, though not perturbing cell cycle progression, increases the rate of cell death in normal cell lines. In contrast, cell death is not affected in cells from genetic instability syndromes; this reflects a specific failure of the apoptotic response. Reintroduction of complementation group C in FA cells re‐established the apoptotic response to ELF‐MF. Thus, genes implicated in genetic instability syndromes are relevant in modulating the response of cells to ELF‐MF.
In this work, we have tried to establish whether human memory T cells may be protected from Fas (... more In this work, we have tried to establish whether human memory T cells may be protected from Fas (CD95)-induced apoptosis when correctly activated by Ag, and not protected when nonspecifically or incorrectly activated. In particular, we wanted to investigate the molecular mechanisms that regulate the fate of memory T cells following an antigenic challenge. To address this issue, we chose an experimental system that closely mimics physiological T cell activation such as human T cell lines and clones specific for viral peptides or alloantigens. We demonstrate that memory T cells acquire an activation-induced cell death (AICD)-resistant phenotype when TCRs are properly engaged by specific Ag bound to MHC molecules. Ag concentration and costimulation are critical parameters in regulating the protective effect. The analysis of the mechanisms involved in the block of CD95 signal transduction pathways revealed that the crucial events are the inhibition of CD95-associated IL-1beta-converting...
In this work, we have tried to establish whether human memory T cells may be protected from Fas (... more In this work, we have tried to establish whether human memory T cells may be protected from Fas (CD95)-induced apoptosis when correctly activated by Ag, and not protected when nonspecifically or incorrectly activated. In particular, we wanted to investigate the molecular mechanisms that regulate the fate of memory T cells following an antigenic challenge. To address this issue, we chose an experimental system that closely mimics physiological T cell activation such as human T cell lines and clones specific for viral peptides or alloantigens. We demonstrate that memory T cells acquire an activation-induced cell death (AICD)-resistant phenotype when TCRs are properly engaged by specific Ag bound to MHC molecules. Ag concentration and costimulation are critical parameters in regulating the protective effect. The analysis of the mechanisms involved in the block of CD95 signal transduction pathways revealed that the crucial events are the inhibition of CD95-associated IL-1beta-converting...
By means of the yeast two-hybrid system, we have discovered a novel physical interaction between ... more By means of the yeast two-hybrid system, we have discovered a novel physical interaction between the adenovirus E1A oncoprotein and Ran, a small GTPase which regulates nucleocytoplasmic transport, cell cycle progression, and mitotic spindle organization. Expression of E1A elicits induction of S phase and centrosome amplification in a variety of rodent cell lines. The induction of supernumerary centrosomes requires functional RCC1, the nucleotide exchange factor for Ran and, hence, a functional Ran network. The E1A portion responsible for the interaction with Ran is the extreme NH(2)-terminal region (amino acids 1-36), which is also required for the induction of centrosome amplification. In an in vitro assay with recombinant proteins, wild-type E1A interferes with nucleotide exchange on Ran, whereas an E1A mutant, deleted from the extreme NH(2)-terminal region, does not. In addition, we detected an in vitro interaction between Ran and HPV-16 E7 and SV40 large T antigen, two oncoprote...
Journal of immunology (Baltimore, Md. : 1950), 1999
In this work, we have tried to establish whether human memory T cells may be protected from Fas (... more In this work, we have tried to establish whether human memory T cells may be protected from Fas (CD95)-induced apoptosis when correctly activated by Ag, and not protected when nonspecifically or incorrectly activated. In particular, we wanted to investigate the molecular mechanisms that regulate the fate of memory T cells following an antigenic challenge. To address this issue, we chose an experimental system that closely mimics physiological T cell activation such as human T cell lines and clones specific for viral peptides or alloantigens. We demonstrate that memory T cells acquire an activation-induced cell death (AICD)-resistant phenotype when TCRs are properly engaged by specific Ag bound to MHC molecules. Ag concentration and costimulation are critical parameters in regulating the protective effect. The analysis of the mechanisms involved in the block of CD95 signal transduction pathways revealed that the crucial events are the inhibition of CD95-associated IL-1beta-converting...
Mitosis is the most potentially dangerous event in the life of a cell, during which the cell gene... more Mitosis is the most potentially dangerous event in the life of a cell, during which the cell genetic identity is transmitted to daughters; errors at this stage may yield aneuploid cells that can initiate a genetically unstable clone. The small GTPase Ran is the central element of a conserved signaling network that has a prominent role in mitotic regulation. Pioneering studies with amphibian oocytes indicated that Ran, in the GTP-bound form, activates factors that regulate spindle assembly and dynamics. An increasing body of data indicate higher specificity and complexity in mitotic control operated by Ran in somatic cells. Newly identified target factors of Ran operate with different specificity, and it is emerging that mitotic progression requires the precise positioning of Ran network components and effectors at specific sites of the mitotic apparatus according to a highly regulated schedule in space and time. In this review we summarize our current understanding of Ran control of mitosis and highlight the specificity of mechanisms operating in mammalian somatic cells.
The Ran GTPase plays a central function in control of nucleo-cytoplasmic transport in interphase.... more The Ran GTPase plays a central function in control of nucleo-cytoplasmic transport in interphase. Mitotic roles of Ran have also been firmly established in Xenopus oocyte extracts. In this system, Ran-GTP, or the RCC1 exchange factor for Ran, drive spindle assembly by regulating the availability of `aster-promoting activities'. In previous studies to assess whether the Ran network also influences mitosis in mammalian cells, we found that overexpression of Ran-binding protein 1 (RanBP1), a major effector of Ran, induces multipolar spindles. We now show that these abnormal spindles are generated through loss of cohesion in mitotic centrosomes. Specifically,RanBP1 excess induces splitting of mother and daughter centrioles at spindle poles; the resulting split centrioles can individually organize functional microtubule arrays, giving rise to functional spindle poles. RanBP1-dependent centrosome splitting is specifically induced in mitosis and requires microtubule integrity and Eg5 a...
Human lymphoblastoid cells of normal origin and from genetic instability syndromes, i.e. Fanconi ... more Human lymphoblastoid cells of normal origin and from genetic instability syndromes, i.e. Fanconi anemia (FA) group C and ataxia telangectasia, were continuously exposed to extremely low frequency magnetic field (ELF‐MF). We report that ELF‐MF, though not perturbing cell cycle progression, increases the rate of cell death in normal cell lines. In contrast, cell death is not affected in cells from genetic instability syndromes; this reflects a specific failure of the apoptotic response. Reintroduction of complementation group C in FA cells re‐established the apoptotic response to ELF‐MF. Thus, genes implicated in genetic instability syndromes are relevant in modulating the response of cells to ELF‐MF.
In this work, we have tried to establish whether human memory T cells may be protected from Fas (... more In this work, we have tried to establish whether human memory T cells may be protected from Fas (CD95)-induced apoptosis when correctly activated by Ag, and not protected when nonspecifically or incorrectly activated. In particular, we wanted to investigate the molecular mechanisms that regulate the fate of memory T cells following an antigenic challenge. To address this issue, we chose an experimental system that closely mimics physiological T cell activation such as human T cell lines and clones specific for viral peptides or alloantigens. We demonstrate that memory T cells acquire an activation-induced cell death (AICD)-resistant phenotype when TCRs are properly engaged by specific Ag bound to MHC molecules. Ag concentration and costimulation are critical parameters in regulating the protective effect. The analysis of the mechanisms involved in the block of CD95 signal transduction pathways revealed that the crucial events are the inhibition of CD95-associated IL-1beta-converting...
In this work, we have tried to establish whether human memory T cells may be protected from Fas (... more In this work, we have tried to establish whether human memory T cells may be protected from Fas (CD95)-induced apoptosis when correctly activated by Ag, and not protected when nonspecifically or incorrectly activated. In particular, we wanted to investigate the molecular mechanisms that regulate the fate of memory T cells following an antigenic challenge. To address this issue, we chose an experimental system that closely mimics physiological T cell activation such as human T cell lines and clones specific for viral peptides or alloantigens. We demonstrate that memory T cells acquire an activation-induced cell death (AICD)-resistant phenotype when TCRs are properly engaged by specific Ag bound to MHC molecules. Ag concentration and costimulation are critical parameters in regulating the protective effect. The analysis of the mechanisms involved in the block of CD95 signal transduction pathways revealed that the crucial events are the inhibition of CD95-associated IL-1beta-converting...
By means of the yeast two-hybrid system, we have discovered a novel physical interaction between ... more By means of the yeast two-hybrid system, we have discovered a novel physical interaction between the adenovirus E1A oncoprotein and Ran, a small GTPase which regulates nucleocytoplasmic transport, cell cycle progression, and mitotic spindle organization. Expression of E1A elicits induction of S phase and centrosome amplification in a variety of rodent cell lines. The induction of supernumerary centrosomes requires functional RCC1, the nucleotide exchange factor for Ran and, hence, a functional Ran network. The E1A portion responsible for the interaction with Ran is the extreme NH(2)-terminal region (amino acids 1-36), which is also required for the induction of centrosome amplification. In an in vitro assay with recombinant proteins, wild-type E1A interferes with nucleotide exchange on Ran, whereas an E1A mutant, deleted from the extreme NH(2)-terminal region, does not. In addition, we detected an in vitro interaction between Ran and HPV-16 E7 and SV40 large T antigen, two oncoprote...
Journal of immunology (Baltimore, Md. : 1950), 1999
In this work, we have tried to establish whether human memory T cells may be protected from Fas (... more In this work, we have tried to establish whether human memory T cells may be protected from Fas (CD95)-induced apoptosis when correctly activated by Ag, and not protected when nonspecifically or incorrectly activated. In particular, we wanted to investigate the molecular mechanisms that regulate the fate of memory T cells following an antigenic challenge. To address this issue, we chose an experimental system that closely mimics physiological T cell activation such as human T cell lines and clones specific for viral peptides or alloantigens. We demonstrate that memory T cells acquire an activation-induced cell death (AICD)-resistant phenotype when TCRs are properly engaged by specific Ag bound to MHC molecules. Ag concentration and costimulation are critical parameters in regulating the protective effect. The analysis of the mechanisms involved in the block of CD95 signal transduction pathways revealed that the crucial events are the inhibition of CD95-associated IL-1beta-converting...
Mitosis is the most potentially dangerous event in the life of a cell, during which the cell gene... more Mitosis is the most potentially dangerous event in the life of a cell, during which the cell genetic identity is transmitted to daughters; errors at this stage may yield aneuploid cells that can initiate a genetically unstable clone. The small GTPase Ran is the central element of a conserved signaling network that has a prominent role in mitotic regulation. Pioneering studies with amphibian oocytes indicated that Ran, in the GTP-bound form, activates factors that regulate spindle assembly and dynamics. An increasing body of data indicate higher specificity and complexity in mitotic control operated by Ran in somatic cells. Newly identified target factors of Ran operate with different specificity, and it is emerging that mitotic progression requires the precise positioning of Ran network components and effectors at specific sites of the mitotic apparatus according to a highly regulated schedule in space and time. In this review we summarize our current understanding of Ran control of mitosis and highlight the specificity of mechanisms operating in mammalian somatic cells.
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Papers by R. Mangiacasale