Terminal exon characterization with TECtool reveals an abundance of cell-specific isoforms.
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ABSTRACT:
SUBMITTER: Gruber AJ
PROVIDER: S-EPMC7611301 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Huntington's disease (HD) is a devastating neurological disorder that is caused by an expansion of the poly-Q tract in exon 1 of the Huntingtin gene (HTT). HTT is an evolutionarily conserved and ubiquitously expressed protein that has been linked to a variety of functions including transcriptional regulation, mitochondrial function, and vesicle transport. This large protein has numerous caspase and calpain cleavage sites and can be decorated with several post-translational modifications such as phosphorylations, acetylations, sumoylations, and palmitoylations. However, the exact function of HTT and the role played by its modifications in the cell are still not well understood. Scrutiny of HTT function has been focused on a single, full length mRNA. In this study, we report the discovery of 5 novel HTT mRNA splice isoforms that are expressed in normal and HTT-expanded human embryonic stem cell (hESC) lines as well as in cortical neurons differentiated from hESCs. Interestingly, none of the novel isoforms generates a truncated protein. Instead, 4 of the 5 new isoforms specifically eliminate domains and modifications to generate smaller HTT proteins. The fifth novel isoform incorporates a previously unreported additional exon, dubbed 41b, which is hominid-specific and introduces a potential phosphorylation site in the protein. The discovery of this hominid-specific isoform may shed light on human-specific pathogenic mechanisms of HTT, which could not be investigated with current mouse models of the disease.
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Project description:Huntington’s disease (HD) is a devastating neurological disorder that is caused by an expansion of the poly-Q tract in exon 1 of the Huntingtin gene (HTT). HTT is an evolutionarily conserved and ubiquitously expressed protein that has been linked to a variety of functions including transcriptional regulation, mitochondrial function, and vesicle transport. This large protein has numerous caspase and calpain cleavage sites and can be decorated with several post-translational modifications such as phosphorylations, acetylations, sumoylations, and palmitoylations. However, the exact function of HTT and the role played by its modifications in the cell is still not well understood. Scrutiny of HTT function has been focused on a single, full length, mRNA. In this study, we report the discovery of 5 novel HTT mRNA splice isoforms that are expressed in normal and HD-hESC lines as well as cortical neurons differentiated from hESCs. Interestingly, none of the novel isoforms generates a truncated protein. Instead, 4 of the 5 new isoforms specifically eliminate domains and modifications to generate smaller HTT proteins. The fifth novel isoform incorporates a previously unreported additional exon, dubbed 41b, which is hominid-specific and introduces a potential phosphorylation site in the protein. The discovery of this hominid-specific isoform may shed light on human-specific pathogenic mechanisms of HTT, which could not be investigated with current mouse models of the disease. Furthermore, it provides a new human-specific target for drug screening in Huntington’s disease. We performed RNAseq of human embryonic stem cells in pluripotency conditions to check expression of multiple HTT isoforms.
2015-06-10 | E-GEOD-66769 | biostudies-arrayexpress
Project description:Huntington’s disease (HD) is a devastating neurological disorder that is caused by an expansion of the poly-Q tract in exon 1 of the Huntingtin gene (HTT). HTT is an evolutionarily conserved and ubiquitously expressed protein that has been linked to a variety of functions including transcriptional regulation, mitochondrial function, and vesicle transport. This large protein has numerous caspase and calpain cleavage sites and can be decorated with several post-translational modifications such as phosphorylations, acetylations, sumoylations, and palmitoylations. However, the exact function of HTT and the role played by its modifications in the cell is still not well understood. Scrutiny of HTT function has been focused on a single, full length, mRNA. In this study, we report the discovery of 5 novel HTT mRNA splice isoforms that are expressed in normal and HD-hESC lines as well as cortical neurons differentiated from hESCs. Interestingly, none of the novel isoforms generates a truncated protein. Instead, 4 of the 5 new isoforms specifically eliminate domains and modifications to generate smaller HTT proteins. The fifth novel isoform incorporates a previously unreported additional exon, dubbed 41b, which is hominid-specific and introduces a potential phosphorylation site in the protein. The discovery of this hominid-specific isoform may shed light on human-specific pathogenic mechanisms of HTT, which could not be investigated with current mouse models of the disease. Furthermore, it provides a new human-specific target for drug screening in Huntington’s disease.
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Project description:Influenza virus uses a unique cap-snatching mechanism characterized by hijacking and cleavage of host capped pre-mRNAs, resulting in short capped RNAs, which are used as primers for viral mRNA synthesis. The PA subunit of influenza polymerase carries the endonuclease activity that catalyzes the host mRNA cleavage reaction. Here, we show that PA is a sequence selective endonuclease with distinct preference to cleave at the 3' end of a guanine (G) base in RNA. The G specificity is exhibited by the native influenza polymerase complex associated with viral ribonucleoprotein particles and is conferred by an intrinsic G specificity of the isolated PA endonuclease domain PA-Nter. In addition, RNA cleavage site choice by the full polymerase is also guided by cap binding to the PB2 subunit, from which RNA cleavage preferentially occurs at the 12th nt downstream of the cap. However, if a G residue is present in the region of 10-13 nucleotides from the cap, cleavage preferentially occurs at G. This is the first biochemical evidence of influenza polymerase PA showing intrinsic sequence selective endonuclease activity.
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Project description:Despite intensive efforts on annotation of eukaryotic transcriptoms, little is known about the regulation of low-abundance transcripts. To address this question, we analysed the regulation of novel low-abundance transcript variants of human acyl-CoA binding protein (ACBP), an important multifunctional housekeeping protein, which we have identified by screening of human expressed sequence tags in combination with ab initio gene prediction. By using RT-, real-time RT- and rapid amplification of cDNA ends-PCR in five human tissues, we find these transcripts, which are generated by a consequent use of alternative promoters and alternate first or first two exons, to be authentic ones. They show a tissue-specific distribution and intrinsic responsiveness to glucose and insulin. Promoter analyses of the corresponding transcripts revealed a differential regulation mediated by sterol regulatory element-binding protein-2, hepatocyte nuclear factor-4α and nuclear factor κB (NF-κB), central transcription factors of fat and glucose metabolism and inflammation. Subcellular localization studies of deduced isoforms in liver HepG2 cells showed that they are distributed in different compartments. By demonstrating that ACBP is a target of NF-κB, our findings link fatty acid metabolism with inflammation. Furthermore, our findings show that low-abundance transcripts are regulated in a similar mode than their high-abundance counterparts.
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Project description:BackgroundBrassica napus is an important agricultural species, improving stress resistance was one of the main breeding goals at present. Non-specific lipid transfer proteins (nsLTPs) are small, basic proteins which are involved in some biotic or abiotic stress responses. B. napus is susceptible to a variety of fungal diseases, so identify the BnLTPs and their expression in disease responses is very important. The common reference genome of B. napus does not contain all B. napus genes because of gene presence/absence variations between individuals. Therefore, it was necessary to search for candidate BnLTP genes in the B. napus pangenome.ResultsIn the present study, the BnLTP genes were identified throughout the pangenome, and different BnLTP genes were presented among varieties. Totally, 246 BnLTP genes were identified and could be divided into five types (1, 2, C, D, and G). The classification, phylogenetic reconstruction, chromosome distribution, functional annotation, and gene expression were analyzed. We also identified potential cis-elements that respond to biotic and abiotic stresses in the 2 kb upstream regions of all BnLTP genes. RNA sequencing analysis showed that the BnLTP genes were involved in the response to Sclerotinia sclerotiorum infection. We identified 32 BnLTPs linked to blackleg resistance quantitative trait locus (QTL).ConclusionThe identification and analysis of LTP genes in the B. napus pangenome could help to elucidate the function of BnLTP family members and provide new information for future molecular breeding in B. napus.
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Project description:The SALL2 transcription factor, an evolutionarily conserved gene through vertebrates, is involved in normal development and neuronal differentiation. In disease, SALL2 is associated with eye, kidney, and brain disorders, but mainly is related to cancer. Some studies support a tumor suppressor role and others an oncogenic role for SALL2, which seems to depend on the cancer type. An additional consideration is tissue-dependent expression of different SALL2 isoforms. Human and mouse SALL2 gene loci contain two promoters, each controlling the expression of a different protein isoform (E1 and E1A). Also, several improvements on the human genome assembly and gene annotation through next-generation sequencing technologies reveal correction and annotation of additional isoforms, obscuring dissection of SALL2 isoform-specific transcriptional targets and functions. We here integrated current data of normal/tumor gene expression databases along with ChIP-seq binding profiles to analyze SALL2 isoforms expression distribution and infer isoform-specific SALL2 targets. We found that the canonical SALL2 E1 isoform is one of the lowest expressed, while the E1A isoform is highly predominant across cell types. To dissect SALL2 isoform-specific targets, we analyzed publicly available ChIP-seq data from Glioblastoma tumor-propagating cells and in-house ChIP-seq datasets performed in SALL2 wild-type and E1A isoform knockout HEK293 cells. Another available ChIP-seq data in HEK293 cells (ENCODE Consortium Phase III) overexpressing a non-canonical SALL2 isoform (short_E1A) was also analyzed. Regardless of cell type, our analysis indicates that the SALL2 long E1 and E1A isoforms, but not short_E1A, are mostly contributing to transcriptional control, and reveals a highly conserved network of brain-specific transcription factors (i.e., SALL3, POU3F2, and NPAS3). Our data integration identified a conserved molecular network in which SALL2 regulates genes associated with neural function, cell differentiation, development, and cell adhesion between others. Also, we identified PODXL as a gene that is likely regulated by SALL2 across tissues. Our study encourages the validation of publicly available ChIP-seq datasets to assess a specific gene/isoform's transcriptional targets. The knowledge of SALL2 isoforms expression and function in different tissue contexts is relevant to understanding its role in disease.
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