In the past decade, RNA fragments derived from full length small nucleolar RNAs (snoRNAs) have be... more In the past decade, RNA fragments derived from full length small nucleolar RNAs (snoRNAs) have been shown to be specifically excised and functional. These sno-derived RNAs (sdRNAs) have been implicated as gene regulators in a multitude of cancers, controlling a variety of genes post-transcriptionally via association with the RNA-induced silencing complex (RISC). In this review, we have summarized the literature connecting sdRNAs to cancer gene regulation. SdRNAs possess miRNA-like functions, and are able to fill the role of tumor-suppressor or tumor-promoter in a tissue context-dependent manner. Indeed, there are many miRNAs that are actually derived from snoRNA transcripts, meaning that they are truly sdRNAs and as such are included in this review. As sdRNAs are frequently discarded from ncRNA analyses, we emphasize that sdRNAs are functionally relevant gene regulators and likely represent an overlooked subclass of miRNAs. Based on the evidence provided by the papers reviewed here,...
ABSTRACTWe have identified 38 specifically excised, differentially expressed snoRNA fragments (sd... more ABSTRACTWe have identified 38 specifically excised, differentially expressed snoRNA fragments (sdRNAs) in TCGA prostate cancer (PCa) patient samples as compared to normal prostate controls. SnoRNA-derived fragments sdRNA-D19b and -A24 emerged among the most differentially expressed and were selected for further experimentation. We found that overexpression of either sdRNA significantly increased PC3 (a well-established model of castration-resistant prostate cancer (CRPC)) cell proliferation, and that sdRNA-D19b overexpression also markedly increased the rate of PC3 cell migration. In addition, both sdRNAs provided drug-specific resistances with sdRNA-D19b levels correlating with paclitaxel resistance and sdRNA-24A conferring dasatinib resistance. In silico and in vitro analyses revealed that two established PCa tumor suppressor genes, CD44 and CDK12, represent targets for sdRNA-D19b and sdRNA-A24 respectively. This outlines a biologically coherent mechanism by which sdRNAs downregul...
ABSTRACTThe widespread utilization of high-throughput sequencing technologies has unequivocally d... more ABSTRACTThe widespread utilization of high-throughput sequencing technologies has unequivocally demonstrated that eukaryotic transcriptomes consist primarily (>98%) of non-coding RNA (ncRNA) transcripts significantly more diverse than their protein-coding counterparts.ncRNAs are typically divided into two categories based on their length. (1) ncRNAs less than 200 nucleotides (nt) long are referred as small non-coding RNAs (sncRNAs) and include microRNAs (miRNAs), piwi-interacting RNAs (piRNAs), small nucleolar RNAs (snoRNAs), transfer ribonucleic RNAs (tRNAs), etc., and the majority of these are thought to function primarily in controlling gene expression. That said, the full repertoire of sncRNAs remains fairly poorly defined as evidenced by two entirely new classes of sncRNAs only recently being reported, i.e., snoRNA-derived RNAs (sdRNAs) and tRNA-derived fragments (tRFs). (2) ncRNAs longer than 200 nt long are known as long ncRNAs (lncRNAs). lncRNAs represent the 2nd largest ...
ABSTRACTSalmonella Outer Membrane Vesicles (OMVs) were recently shown to inhibit P22 bacteriophag... more ABSTRACTSalmonella Outer Membrane Vesicles (OMVs) were recently shown to inhibit P22 bacteriophage infection. Furthermore, despite there being several published reports now independently describing (1) the marked prevalence of tRFs within secreted vesicle transcriptomes and (2) roles for specific tRFs in facilitating/inhibiting viral replication, there have been no examinations of the effects of vesicle-secreted tRFs on viral infection reported to date. Notably, while specific tRFs have been reported in a number of bacteria, the tRFs expressed by salmonellae have not been previously characterized. As such, we recently screened small RNA-seq datasets for the presence of recurrent, specifically excised tRFs and identified 31 recurrent, relatively abundant tRFs expressed by Salmonella enterica serovar Typhimurium (SL1344). What’s more, we find S. Typhimurium OMVs contain significant levels of tRFs highly complementary to known Salmonella enterica-infecting bacteriophage with 17 of 31 t...
An increasingly apparent role of noncoding RNA (ncRNAs) is to coordinate gene expression during e... more An increasingly apparent role of noncoding RNA (ncRNAs) is to coordinate gene expression during environmental stress. A mounting body of evidence implicates small RNAs (sRNAs) as key drivers of Salmonella stress survival. Generally thought to be 50–500 nucleotides in length and to occur in intergenic regions, sRNAs typically regulate protein expression through base pairing with mRNA targets. In this work, through employing a refined definition of sRNAs allowing for shorter sequences and sRNA loci to overlap with annotated protein-coding gene loci, we have identified 475 previously unannotated sRNAs that are significantly differentially expressed during carbon starvation (C-starvation). Northern blotting and quantitative RT-PCRs confirm the expressions and identities of several of these novel sRNAs, and our computational analyses find the majority to be highly conserved and structurally related to known sRNAs. Importantly, we show that deletion of one of the sRNAs dynamically express...
RNA editing by RNA specific adenosine deaminase acting on RNA (ADAR) is increasingly being found ... more RNA editing by RNA specific adenosine deaminase acting on RNA (ADAR) is increasingly being found to alter microRNA (miRNA) regulation. Editing of miRNA transcripts can affect their processing, as well as which messenger RNAs (mRNAs) they target. Further, editing of target mRNAs can also affect their complementarity to miRNAs. Notably, ADAR editing is often increased in malignancy with the effect of these RNA changes being largely unclear. In addition, numerous reports have now identified an array of miRNAs that directly contribute to various malignancies although the majority of their targets remain largely undefined. Here we propose that modulating the targets of miRNAs via mRNA editing is a frequent occurrence in cancer and an underappreciated participant in pathology. In order to more accurately characterize the relationship between these two regulatory processes, this study examined RNA editing events within mRNA sequences of two breast cancer cell lines (MCF-7 and MDA-MB-231) a...
Methods in molecular biology (Clifton, N.J.), 2017
MicroRNA (miRNA) mediated silencing and repression of mRNA molecules requires complementary base ... more MicroRNA (miRNA) mediated silencing and repression of mRNA molecules requires complementary base pairing between the "seed" region of the miRNA and the "seed match" region of target mRNAs. While this mechanism is fairly well understood, accurate prediction of valid miRNA targets remains challenging due to factors such as imperfect sequence specificity, target site availability, and the thermodynamic stability of the mRNA structure itself. As knowledge of what genes are being targeted by each miRNA is arguably the most important facet of miRNA biology, many approaches have been developed to address the need for reliable prediction and ranking of putative targets, with most using a combination of various strategies such as evolutionary conservation, statistical inference, and distinct features of the target sequences themselves. This chapter reviews the pros and cons of a number of different prediction algorithms, showcases some databases that store experimentally ...
Genetic searches for tumor suppressors have recently linked small nucleolar RNA misregulations wi... more Genetic searches for tumor suppressors have recently linked small nucleolar RNA misregulations with tumorigenesis. In addition to their classically defined functions, several small nucleolar RNAs are now known to be processed into short microRNA-like fragments called small nucleolar RNA-derived RNAs. To determine if any small nucleolar RNA-derived RNAs contribute to breast malignancy, we recently performed a RNA-seq-based comparison of the small nucleolar RNA-derived RNAs of two breast cancer cell lines (MCF-7 and MDA-MB-231) and identified small nucleolar RNA-derived RNAs derived from 13 small nucleolar RNAs overexpressed in MDA-MB-231s. Importantly, we find that inhibiting the most differentially expressed of these small nucleolar RNA-derived RNAs (sdRNA-93) in MDA-MB-231 cells results primarily in a loss of invasiveness, whereas increased sdRNA-93 expression in either cell line conversely results in strikingly enhanced invasion. Excitingly, we recently determined sdRNA-93 express...
In recent years, sequencing technologies have enabled the identification of a wide range of non-c... more In recent years, sequencing technologies have enabled the identification of a wide range of non-coding RNAs (ncRNAs). Unfortunately, annotation and integration of ncRNA data has lagged behind their identification. Given the large quantity of information being obtained in this area, there emerges an urgent need to integrate what is being discovered by a broad range of relevant communities. To this end, the Non-Coding RNA Ontology (NCRO) is being developed to provide a systematically structured and precisely defined controlled vocabulary for the domain of ncRNAs, thereby facilitating the discovery, curation, analysis, exchange, and reasoning of data about structures of ncRNAs, their molecular and cellular functions, and their impacts upon phenotypes. The goal of NCRO is to serve as a common resource for annotations of diverse research in a way that will significantly enhance integrative and comparative analysis of the myriad resources currently housed in disparate sources. It is our b...
Small RNAs (sRNAs) are short (˜50-200 nucleotides) noncoding RNAs that regulate cellular activiti... more Small RNAs (sRNAs) are short (˜50-200 nucleotides) noncoding RNAs that regulate cellular activities across bacteria. Salmonella enterica starved of a carbon-energy (C) source experience a host of genetic and physiological changes broadly referred to as the starvation-stress response (SSR). In an attempt to identify novel sRNAs contributing to SSR control, we grew log-phase, 5-h C-starved and 24-h C-starved cultures of the virulent Salmonella enterica subspecies enterica serovar Typhimurium strain SL1344 and comprehensively sequenced their small RNA transcriptomes. Strikingly, after employing a novel strategy for sRNA discovery based on identifying dynamic transcripts arising from "gene-empty" regions, we identify 58 wholly undescribed Salmonella sRNA genes potentially regulating SSR averaging an ˜1,000-fold change in expression between log-phase and C-starved cells. Importantly, the expressions of individual sRNA loci were confirmed by both comprehensive transcriptome anal...
American journal of physiology. Lung cellular and molecular physiology, Jan 2, 2015
In hypoxia, mitochondria-generated reactive oxygen species (ROS) not only stimulate accumulation ... more In hypoxia, mitochondria-generated reactive oxygen species (ROS) not only stimulate accumulation of the transcriptional regulator of hypoxic gene expression, Hif-1, but also cause oxidative base modifications in hypoxic response elements (HREs) of hypoxia-inducible genes. When the hypoxia-induced base modifications are suppressed, Hif-1 fails to associate with the HRE of the VEGF promoter and VEGF mRNA accumulation is blunted. The mechanism linking base modifications to transcription is unknown. Here we determined if recruitment of base excision DNA repair (BER) enzymes in response to hypoxia-induced promoter modifications was required for transcription complex assembly and VEGF mRNA expression. Using ChIP analyses in pulmonary artery endothelial cells, we found that hypoxia-mediated formation of the base oxidation product 8-oxoguanine (8-oxoG) in VEGF HREs was temporally associated with binding of Hif-1α and the BER enzymes Ogg1 and Ref-1/Ape1 and introduction of DNA strand breaks....
MicroRNAs (miRs) are small noncoding RNAs that typically act as regulators of gene expression by ... more MicroRNAs (miRs) are small noncoding RNAs that typically act as regulators of gene expression by base pairing with the 3' UTR of messenger RNAs (mRNAs) and either repressing their translation of initiating degradation. Descriptions of the molecular origins of independent miR molecules currently support the hypothesis that miR hairpin generation is based on the adjacent insertion of two related transposable elements (TEs) at one genomic locus. Thus transcription across such TE interfaces establishes many, if not the majority of functional miRs. We have recently completed an in depth analysis of the genomic origins of all currently annotated miR loci, detailing the formations of 3,605 distinct miR loci primarily from TEs. The implications of these findings are substantial for understanding how TEs confer increased genomic fitness, describing miR transcriptional regulations, and making accurate miR target predictions. As such, hypothesizing that a miR and its mRNA target sites migh...
In the past decade, RNA fragments derived from full length small nucleolar RNAs (snoRNAs) have be... more In the past decade, RNA fragments derived from full length small nucleolar RNAs (snoRNAs) have been shown to be specifically excised and functional. These sno-derived RNAs (sdRNAs) have been implicated as gene regulators in a multitude of cancers, controlling a variety of genes post-transcriptionally via association with the RNA-induced silencing complex (RISC). In this review, we have summarized the literature connecting sdRNAs to cancer gene regulation. SdRNAs possess miRNA-like functions, and are able to fill the role of tumor-suppressor or tumor-promoter in a tissue context-dependent manner. Indeed, there are many miRNAs that are actually derived from snoRNA transcripts, meaning that they are truly sdRNAs and as such are included in this review. As sdRNAs are frequently discarded from ncRNA analyses, we emphasize that sdRNAs are functionally relevant gene regulators and likely represent an overlooked subclass of miRNAs. Based on the evidence provided by the papers reviewed here,...
ABSTRACTWe have identified 38 specifically excised, differentially expressed snoRNA fragments (sd... more ABSTRACTWe have identified 38 specifically excised, differentially expressed snoRNA fragments (sdRNAs) in TCGA prostate cancer (PCa) patient samples as compared to normal prostate controls. SnoRNA-derived fragments sdRNA-D19b and -A24 emerged among the most differentially expressed and were selected for further experimentation. We found that overexpression of either sdRNA significantly increased PC3 (a well-established model of castration-resistant prostate cancer (CRPC)) cell proliferation, and that sdRNA-D19b overexpression also markedly increased the rate of PC3 cell migration. In addition, both sdRNAs provided drug-specific resistances with sdRNA-D19b levels correlating with paclitaxel resistance and sdRNA-24A conferring dasatinib resistance. In silico and in vitro analyses revealed that two established PCa tumor suppressor genes, CD44 and CDK12, represent targets for sdRNA-D19b and sdRNA-A24 respectively. This outlines a biologically coherent mechanism by which sdRNAs downregul...
ABSTRACTThe widespread utilization of high-throughput sequencing technologies has unequivocally d... more ABSTRACTThe widespread utilization of high-throughput sequencing technologies has unequivocally demonstrated that eukaryotic transcriptomes consist primarily (>98%) of non-coding RNA (ncRNA) transcripts significantly more diverse than their protein-coding counterparts.ncRNAs are typically divided into two categories based on their length. (1) ncRNAs less than 200 nucleotides (nt) long are referred as small non-coding RNAs (sncRNAs) and include microRNAs (miRNAs), piwi-interacting RNAs (piRNAs), small nucleolar RNAs (snoRNAs), transfer ribonucleic RNAs (tRNAs), etc., and the majority of these are thought to function primarily in controlling gene expression. That said, the full repertoire of sncRNAs remains fairly poorly defined as evidenced by two entirely new classes of sncRNAs only recently being reported, i.e., snoRNA-derived RNAs (sdRNAs) and tRNA-derived fragments (tRFs). (2) ncRNAs longer than 200 nt long are known as long ncRNAs (lncRNAs). lncRNAs represent the 2nd largest ...
ABSTRACTSalmonella Outer Membrane Vesicles (OMVs) were recently shown to inhibit P22 bacteriophag... more ABSTRACTSalmonella Outer Membrane Vesicles (OMVs) were recently shown to inhibit P22 bacteriophage infection. Furthermore, despite there being several published reports now independently describing (1) the marked prevalence of tRFs within secreted vesicle transcriptomes and (2) roles for specific tRFs in facilitating/inhibiting viral replication, there have been no examinations of the effects of vesicle-secreted tRFs on viral infection reported to date. Notably, while specific tRFs have been reported in a number of bacteria, the tRFs expressed by salmonellae have not been previously characterized. As such, we recently screened small RNA-seq datasets for the presence of recurrent, specifically excised tRFs and identified 31 recurrent, relatively abundant tRFs expressed by Salmonella enterica serovar Typhimurium (SL1344). What’s more, we find S. Typhimurium OMVs contain significant levels of tRFs highly complementary to known Salmonella enterica-infecting bacteriophage with 17 of 31 t...
An increasingly apparent role of noncoding RNA (ncRNAs) is to coordinate gene expression during e... more An increasingly apparent role of noncoding RNA (ncRNAs) is to coordinate gene expression during environmental stress. A mounting body of evidence implicates small RNAs (sRNAs) as key drivers of Salmonella stress survival. Generally thought to be 50–500 nucleotides in length and to occur in intergenic regions, sRNAs typically regulate protein expression through base pairing with mRNA targets. In this work, through employing a refined definition of sRNAs allowing for shorter sequences and sRNA loci to overlap with annotated protein-coding gene loci, we have identified 475 previously unannotated sRNAs that are significantly differentially expressed during carbon starvation (C-starvation). Northern blotting and quantitative RT-PCRs confirm the expressions and identities of several of these novel sRNAs, and our computational analyses find the majority to be highly conserved and structurally related to known sRNAs. Importantly, we show that deletion of one of the sRNAs dynamically express...
RNA editing by RNA specific adenosine deaminase acting on RNA (ADAR) is increasingly being found ... more RNA editing by RNA specific adenosine deaminase acting on RNA (ADAR) is increasingly being found to alter microRNA (miRNA) regulation. Editing of miRNA transcripts can affect their processing, as well as which messenger RNAs (mRNAs) they target. Further, editing of target mRNAs can also affect their complementarity to miRNAs. Notably, ADAR editing is often increased in malignancy with the effect of these RNA changes being largely unclear. In addition, numerous reports have now identified an array of miRNAs that directly contribute to various malignancies although the majority of their targets remain largely undefined. Here we propose that modulating the targets of miRNAs via mRNA editing is a frequent occurrence in cancer and an underappreciated participant in pathology. In order to more accurately characterize the relationship between these two regulatory processes, this study examined RNA editing events within mRNA sequences of two breast cancer cell lines (MCF-7 and MDA-MB-231) a...
Methods in molecular biology (Clifton, N.J.), 2017
MicroRNA (miRNA) mediated silencing and repression of mRNA molecules requires complementary base ... more MicroRNA (miRNA) mediated silencing and repression of mRNA molecules requires complementary base pairing between the "seed" region of the miRNA and the "seed match" region of target mRNAs. While this mechanism is fairly well understood, accurate prediction of valid miRNA targets remains challenging due to factors such as imperfect sequence specificity, target site availability, and the thermodynamic stability of the mRNA structure itself. As knowledge of what genes are being targeted by each miRNA is arguably the most important facet of miRNA biology, many approaches have been developed to address the need for reliable prediction and ranking of putative targets, with most using a combination of various strategies such as evolutionary conservation, statistical inference, and distinct features of the target sequences themselves. This chapter reviews the pros and cons of a number of different prediction algorithms, showcases some databases that store experimentally ...
Genetic searches for tumor suppressors have recently linked small nucleolar RNA misregulations wi... more Genetic searches for tumor suppressors have recently linked small nucleolar RNA misregulations with tumorigenesis. In addition to their classically defined functions, several small nucleolar RNAs are now known to be processed into short microRNA-like fragments called small nucleolar RNA-derived RNAs. To determine if any small nucleolar RNA-derived RNAs contribute to breast malignancy, we recently performed a RNA-seq-based comparison of the small nucleolar RNA-derived RNAs of two breast cancer cell lines (MCF-7 and MDA-MB-231) and identified small nucleolar RNA-derived RNAs derived from 13 small nucleolar RNAs overexpressed in MDA-MB-231s. Importantly, we find that inhibiting the most differentially expressed of these small nucleolar RNA-derived RNAs (sdRNA-93) in MDA-MB-231 cells results primarily in a loss of invasiveness, whereas increased sdRNA-93 expression in either cell line conversely results in strikingly enhanced invasion. Excitingly, we recently determined sdRNA-93 express...
In recent years, sequencing technologies have enabled the identification of a wide range of non-c... more In recent years, sequencing technologies have enabled the identification of a wide range of non-coding RNAs (ncRNAs). Unfortunately, annotation and integration of ncRNA data has lagged behind their identification. Given the large quantity of information being obtained in this area, there emerges an urgent need to integrate what is being discovered by a broad range of relevant communities. To this end, the Non-Coding RNA Ontology (NCRO) is being developed to provide a systematically structured and precisely defined controlled vocabulary for the domain of ncRNAs, thereby facilitating the discovery, curation, analysis, exchange, and reasoning of data about structures of ncRNAs, their molecular and cellular functions, and their impacts upon phenotypes. The goal of NCRO is to serve as a common resource for annotations of diverse research in a way that will significantly enhance integrative and comparative analysis of the myriad resources currently housed in disparate sources. It is our b...
Small RNAs (sRNAs) are short (˜50-200 nucleotides) noncoding RNAs that regulate cellular activiti... more Small RNAs (sRNAs) are short (˜50-200 nucleotides) noncoding RNAs that regulate cellular activities across bacteria. Salmonella enterica starved of a carbon-energy (C) source experience a host of genetic and physiological changes broadly referred to as the starvation-stress response (SSR). In an attempt to identify novel sRNAs contributing to SSR control, we grew log-phase, 5-h C-starved and 24-h C-starved cultures of the virulent Salmonella enterica subspecies enterica serovar Typhimurium strain SL1344 and comprehensively sequenced their small RNA transcriptomes. Strikingly, after employing a novel strategy for sRNA discovery based on identifying dynamic transcripts arising from "gene-empty" regions, we identify 58 wholly undescribed Salmonella sRNA genes potentially regulating SSR averaging an ˜1,000-fold change in expression between log-phase and C-starved cells. Importantly, the expressions of individual sRNA loci were confirmed by both comprehensive transcriptome anal...
American journal of physiology. Lung cellular and molecular physiology, Jan 2, 2015
In hypoxia, mitochondria-generated reactive oxygen species (ROS) not only stimulate accumulation ... more In hypoxia, mitochondria-generated reactive oxygen species (ROS) not only stimulate accumulation of the transcriptional regulator of hypoxic gene expression, Hif-1, but also cause oxidative base modifications in hypoxic response elements (HREs) of hypoxia-inducible genes. When the hypoxia-induced base modifications are suppressed, Hif-1 fails to associate with the HRE of the VEGF promoter and VEGF mRNA accumulation is blunted. The mechanism linking base modifications to transcription is unknown. Here we determined if recruitment of base excision DNA repair (BER) enzymes in response to hypoxia-induced promoter modifications was required for transcription complex assembly and VEGF mRNA expression. Using ChIP analyses in pulmonary artery endothelial cells, we found that hypoxia-mediated formation of the base oxidation product 8-oxoguanine (8-oxoG) in VEGF HREs was temporally associated with binding of Hif-1α and the BER enzymes Ogg1 and Ref-1/Ape1 and introduction of DNA strand breaks....
MicroRNAs (miRs) are small noncoding RNAs that typically act as regulators of gene expression by ... more MicroRNAs (miRs) are small noncoding RNAs that typically act as regulators of gene expression by base pairing with the 3' UTR of messenger RNAs (mRNAs) and either repressing their translation of initiating degradation. Descriptions of the molecular origins of independent miR molecules currently support the hypothesis that miR hairpin generation is based on the adjacent insertion of two related transposable elements (TEs) at one genomic locus. Thus transcription across such TE interfaces establishes many, if not the majority of functional miRs. We have recently completed an in depth analysis of the genomic origins of all currently annotated miR loci, detailing the formations of 3,605 distinct miR loci primarily from TEs. The implications of these findings are substantial for understanding how TEs confer increased genomic fitness, describing miR transcriptional regulations, and making accurate miR target predictions. As such, hypothesizing that a miR and its mRNA target sites migh...
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