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Translational Psychiatry Nov 2023Neurexins are synaptic adhesion molecules that play diverse roles in synaptic development, function, maintenance, and plasticity. Neurexin genes have been associated...
Neurexins are synaptic adhesion molecules that play diverse roles in synaptic development, function, maintenance, and plasticity. Neurexin genes have been associated with changes in human behavior, where variants in NRXN1 are associated with autism, schizophrenia, and Tourette syndrome. While NRXN1, NRXN2, and NRXN3 all encode major α and β isoforms, NRXN1 uniquely encodes a γ isoform, for which mechanistic roles in behavior have yet to be defined. Here, we show that both α and γ isoforms of neurexin/nrx-1 are required for the C. elegans behavioral response to food deprivation, a sustained period of hyperactivity upon food loss. We find that the γ isoform regulates initiation and the α isoform regulates maintenance of the behavioral response to food deprivation, demonstrating cooperative function of multiple nrx-1 isoforms in regulating a sustained behavior. The γ isoform alters monoamine signaling via octopamine, relies on specific expression of NRX-1 isoforms throughout the relevant circuit, and is independent of neuroligin/nlg-1, the canonical trans-synaptic partner of nrx-1. The α isoform regulates the pre-synaptic structure of the octopamine producing RIC neuron and its maintenance role is conditional on neuroligin/nlg-1. Collectively, these results demonstrate that neurexin isoforms can have separate behavioral roles and act cooperatively across neuronal circuits to modify behavior, highlighting the need to directly analyze and consider all isoforms when defining the contribution of neurexins to behavior.
Topics: Animals; Humans; Caenorhabditis elegans; Octopamine; Cell Adhesion Molecules, Neuronal; Protein Isoforms; Neurons; Synapses; Caenorhabditis elegans Proteins
PubMed: 38036526
DOI: 10.1038/s41398-023-02668-z -
Cells Apr 2022Vascular endothelial growth factor A (VEGF-A) is a secreted protein that stimulates angiogenesis in response to hypoxia. Under hypoxic conditions, a non-canonical long...
Vascular endothelial growth factor A (VEGF-A) is a secreted protein that stimulates angiogenesis in response to hypoxia. Under hypoxic conditions, a non-canonical long isoform called is concomitantly expressed with . Once translated, L-VEGF is proteolytically cleaved to generate N-VEGF and VEGF-A. Interestingly, while VEGF-A is secreted and affects the surrounding cells, N-VEGF is mobilized to the nucleus. This suggests that N-VEGF participates in transcriptional response to hypoxia. In this study, we performed a series of complementary experiments to examine the functional role of N-VEGF. Strikingly, we found that the mere expression of N-VEGF followed by its hypoxia-independent mobilization to the nucleus was sufficient to induce key genes associated with angiogenesis, such as isoforms, as well as genes associated with cell survival under hypoxia. Complementarily, when was genetically depleted, key hypoxia-induced genes were downregulated and cells were significantly susceptible to hypoxia-mediated apoptosis. This is the first report of N-VEGF serving as an autoregulatory arm of VEGF-A. Further experiments will be needed to determine the role of N-VEGF in cancer and embryogenesis.
Topics: Apoptosis; Cell Hypoxia; Humans; Hypoxia; Neovascularization, Pathologic; Protein Isoforms; Vascular Endothelial Growth Factor A
PubMed: 35455969
DOI: 10.3390/cells11081289 -
Bioinformatics (Oxford, England) Mar 2020Isoforms are alternatively spliced mRNAs of genes. They can be translated into different functional proteoforms, and thus greatly increase the functional diversity of...
MOTIVATION
Isoforms are alternatively spliced mRNAs of genes. They can be translated into different functional proteoforms, and thus greatly increase the functional diversity of protein variants (or proteoforms). Differentiating the functions of isoforms (or proteoforms) helps understanding the underlying pathology of various complex diseases at a deeper granularity. Since existing functional genomic databases uniformly record the annotations at the gene-level, and rarely record the annotations at the isoform-level, differentiating isoform functions is more challenging than the traditional gene-level function prediction.
RESULTS
Several approaches have been proposed to differentiate the functions of isoforms. They generally follow the multi-instance learning paradigm by viewing each gene as a bag and the spliced isoforms as its instances, and push functions of bags onto instances. These approaches implicitly assume the collected annotations of genes are complete and only integrate multiple RNA-seq datasets. As such, they have compromised performance. We propose a data integrative solution (called DisoFun) to Differentiate isoform Functions with collaborative matrix factorization. DisoFun assumes the functional annotations of genes are aggregated from those of key isoforms. It collaboratively factorizes the isoform data matrix and gene-term data matrix (storing Gene Ontology annotations of genes) into low-rank matrices to simultaneously explore the latent key isoforms, and achieve function prediction by aggregating predictions to their originating genes. In addition, it leverages the PPI network and Gene Ontology structure to further coordinate the matrix factorization. Extensive experimental results show that DisoFun improves the area under the receiver operating characteristic curve and area under the precision-recall curve of existing solutions by at least 7.7 and 28.9%, respectively. We further investigate DisoFun on four exemplar genes (LMNA, ADAM15, BCL2L1 and CFLAR) with known functions at the isoform-level, and observed that DisoFun can differentiate functions of their isoforms with 90.5% accuracy.
AVAILABILITY AND IMPLEMENTATION
The code of DisoFun is available at mlda.swu.edu.cn/codes.php?name=DisoFun.
SUPPLEMENTARY INFORMATION
Supplementary data are available at Bioinformatics online.
Topics: Computational Biology; Gene Ontology; Molecular Sequence Annotation; Protein Isoforms; ROC Curve
PubMed: 32176770
DOI: 10.1093/bioinformatics/btz847 -
Bioinformatics (Oxford, England) Sep 2022High-resolution annotation of gene functions is a central task in functional genomics. Multiple proteoforms translated from alternatively spliced isoforms from a single...
MOTIVATION
High-resolution annotation of gene functions is a central task in functional genomics. Multiple proteoforms translated from alternatively spliced isoforms from a single gene are actual function performers and greatly increase the functional diversity. The specific functions of different isoforms can decipher the molecular basis of various complex diseases at a finer granularity. Multi-instance learning (MIL)-based solutions have been developed to distribute gene(bag)-level Gene Ontology (GO) annotations to isoforms(instances), but they simply presume that a particular annotation of the gene is responsible by only one isoform, neglect the hierarchical structures and semantics of massive GO terms (labels), or can only handle dozens of terms.
RESULTS
We propose an efficacy approach IsofunGO to differentiate massive functions of isoforms by GO embedding. Particularly, IsofunGO first introduces an attributed hierarchical network to model massive GO terms, and a GO network embedding strategy to learn compact representations of GO terms and project GO annotations of genes into compressed ones, this strategy not only explores and preserves hierarchy between GO terms but also greatly reduces the prediction load. Next, it develops an attention-based MIL network to fuse genomics and transcriptomics data of isoforms and predict isoform functions by referring to compressed annotations. Extensive experiments on benchmark datasets demonstrate the efficacy of IsofunGO. Both the GO embedding and attention mechanism can boost the performance and interpretability.
AVAILABILITYAND IMPLEMENTATION
The code of IsofunGO is available at http://www.sdu-idea.cn/codes.php?name=IsofunGO.
SUPPLEMENTARY INFORMATION
Supplementary data are available at Bioinformatics online.
Topics: Gene Ontology; Computational Biology; Molecular Sequence Annotation; Protein Isoforms; Semantics
PubMed: 35997558
DOI: 10.1093/bioinformatics/btac576 -
Cell Death & Disease Mar 2024The advancement of RNAseq and isoform-specific expression platforms has led to the understanding that isoform changes can alter molecular signaling to promote... (Review)
Review
The advancement of RNAseq and isoform-specific expression platforms has led to the understanding that isoform changes can alter molecular signaling to promote tumorigenesis. An active area in cancer research is uncovering the roles of ubiquitination on spliceosome assembly contributing to transcript diversity and expression of alternative isoforms. However, the effects of isoform changes on functionality of ubiquitination machineries (E1, E2, E3, E4, and deubiquitinating (DUB) enzymes) influencing onco- and tumor suppressor protein stabilities is currently understudied. Characterizing these changes could be instrumental in improving cancer outcomes via the identification of novel biomarkers and targetable signaling pathways. In this review, we focus on highlighting reported examples of direct, protein-coded isoform variation of ubiquitination enzymes influencing cancer development and progression in gastrointestinal (GI) malignancies. We have used a semi-automated system for identifying relevant literature and applied established systems for isoform categorization and functional classification to help structure literature findings. The results are a comprehensive snapshot of known isoform changes that are significant to GI cancers, and a framework for readers to use to address isoform variation in their own research. One of the key findings is the potential influence that isoforms of the ubiquitination machinery have on oncoprotein stability.
Topics: Humans; Ubiquitination; Protein Isoforms; Gastrointestinal Neoplasms; Carcinogenesis; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases
PubMed: 38453895
DOI: 10.1038/s41419-024-06575-z -
PloS One 2022Planar cell polarity (PCP) signaling regulates several polarization events during development of ommatidia in the Drosophila eye, including directing chirality by...
Planar cell polarity (PCP) signaling regulates several polarization events during development of ommatidia in the Drosophila eye, including directing chirality by polarizing a cell fate choice and determining the direction and extent of ommatidial rotation. The pksple isoform of the PCP protein Prickle is known to participate in the R3/R4 cell fate decision, but the control of other polarization events and the potential contributions of the three Pk isoforms have not been clarified. Here, by characterizing expression and subcellular localization of individual isoforms together with re-analyzing isoform specific phenotypes, we show that the R3/R4 fate decision, its coordination with rotation direction, and completion of rotation to a final ±90° rotation angle are separable polarization decisions with distinct Pk isoform requirements and contributions. Both pksple and pkpk can enforce robust R3/R4 fate decisions, but only pksple can correctly orient them along the dorsal-ventral axis. In contrast, pksple and pkpk can fully and interchangeably sustain coordination of rotation direction and rotation to completion. We propose that expression dynamics and competitive interactions determine isoform participation in these processes. We propose that the selective requirement for pksple to orient the R3/R4 decision and their interchangeability for coordination and completion of rotation reflects their previously described differential interaction with the Fat/Dachsous system which is known to be required for orientation of R3/R4 decisions but not for coordination or completion of rotation.
Topics: Animals; Brain; CRISPR-Cas Systems; DNA-Binding Proteins; Drosophila; Drosophila Proteins; Eye; Gene Editing; Genotype; LIM Domain Proteins; Membrane Proteins; Phenotype; Protein Isoforms; RNA Interference; RNA, Small Interfering; Signal Transduction
PubMed: 35148314
DOI: 10.1371/journal.pone.0262328 -
Nature Communications Sep 2023Shotgun proteomics is essential for protein identification and quantification in biomedical research, but protein isoform characterization is challenging due to the...
Shotgun proteomics is essential for protein identification and quantification in biomedical research, but protein isoform characterization is challenging due to the extensive number of peptides shared across proteins, hindering our understanding of protein isoform regulation and their roles in normal and disease biology. We systematically assess the challenge and opportunities of shotgun proteomics-based protein isoform characterization using in silico and experimental data, and then present SEPepQuant, a graph theory-based approach to maximize isoform characterization. Using published data from one induced pluripotent stem cell study and two human hepatocellular carcinoma studies, we demonstrate the ability of SEPepQuant in addressing the key limitations of existing methods, providing more comprehensive isoform-level characterization, identifying hundreds of isoform-level regulation events, and facilitating streamlined cross-study comparisons. Our analysis provides solid evidence to support a widespread role of protein isoform regulation in normal and disease processes, and SEPepQuant has broad applications to biological and translational research.
Topics: Humans; Proteomics; Protein Isoforms; Biomedical Research; Carcinoma, Hepatocellular; Liver Neoplasms
PubMed: 37726316
DOI: 10.1038/s41467-023-41558-2 -
European Journal of Medicinal Chemistry Oct 2023Hsp90 isoform-selective inhibitors represent a new paradigm for novel anti-cancer drugs as each of the four isoforms have specific cellular localization, function, and...
Hsp90 isoform-selective inhibitors represent a new paradigm for novel anti-cancer drugs as each of the four isoforms have specific cellular localization, function, and client proteins. The mitochondrial isoform, TRAP1, is the least understood member of the Hsp90 family due to the lack of small molecule tools to study its biological function. Herein, we report novel TRAP1-selective inhibitors used to interrogate TRAP1's biological function along with co-crystal structures of such compounds bound to the N-terminus of TRAP1. Solution of the co-crystal structure allowed for a structure-based approach that resulted in compound 36, which is a 40 nM inhibitor with >250-fold TRAP1 selectivity over Grp94, the isoform with the highest structural similarity to TRAP1 within the N-terminal ATP binding site. Lead compounds 35 and 36 were found to selectively induce TRAP1 client protein degradation without inducing the heat shock response or disrupting Hsp90-cytosolic clients. They were also shown to inhibit OXPHOS, alter cellular metabolism towards glycolysis, disrupt TRAP1 tetramer stability, and disrupt the mitochondrial membrane potential.
Topics: Humans; HSP90 Heat-Shock Proteins; Protein Binding; Protein Isoforms
PubMed: 37307624
DOI: 10.1016/j.ejmech.2023.115531 -
Nature Protocols Feb 2021Although mammalian embryo development depends on critical protein isoforms that arise from embryo-specific nucleic acid modifications, the role of these isoforms is not...
Although mammalian embryo development depends on critical protein isoforms that arise from embryo-specific nucleic acid modifications, the role of these isoforms is not yet clear. Challenges arise in measuring protein isoforms and nucleic acids from the same single embryos and blastomeres. Here we present a multimodal technique for performing same-embryo nucleic acid and protein isoform profiling (single-embryo nucleic acid and protein profiling immunoblot, or snapBlot). The method integrates protein isoform measurement by fractionation polyacrylamide gel electrophoresis (fPAGE) with off-chip analysis of nucleic acids from the nuclei. Once embryos are harvested and cultured to the desired stage, they are sampled into the snapBlot device and subjected to fPAGE. After fPAGE, 'gel pallets' containing nuclei are excised from the snapBlot device for off-chip nucleic acid analyses. fPAGE and nuclei analyses are indexed to each starting sample, yielding same-embryo multimodal measurements. The entire protocol, including processing of samples and data analysis, takes 2-3 d. snapBlot is designed to help reveal the mechanisms by which embryo-specific nucleic acid modifications to both genomic DNA and messenger RNA orchestrate the growth and development of mammalian embryos.
Topics: Animals; Blastocyst; Blastomeres; DNA; Electrophoresis, Polyacrylamide Gel; Embryo, Mammalian; Embryonic Development; Female; Immunoblotting; Mice; Nucleic Acids; Protein Isoforms; RNA, Messenger
PubMed: 33452502
DOI: 10.1038/s41596-020-00449-2 -
Scientific Reports Jan 2020The advent of RNA-seq technologies has switched the paradigm of genetic analysis from a genome to a transcriptome-based perspective. Alternative splicing generates...
The advent of RNA-seq technologies has switched the paradigm of genetic analysis from a genome to a transcriptome-based perspective. Alternative splicing generates functional diversity in genes, but the precise functions of many individual isoforms are yet to be elucidated. Gene Ontology was developed to annotate gene products according to their biological processes, molecular functions and cellular components. Despite a single gene may have several gene products, most annotations are not isoform-specific and do not distinguish the functions of the different proteins originated from a single gene. Several approaches have tried to automatically annotate ontologies at the isoform level, but this has shown to be a daunting task. We have developed ISOGO (ISOform + GO function imputation), a novel algorithm to predict the function of coding isoforms based on their protein domains and their correlation of expression along 11,373 cancer patients. Combining these two sources of information outperforms previous approaches: it provides an area under precision-recall curve (AUPRC) five times larger than previous attempts and the median AUROC of assigned functions to genes is 0.82. We tested ISOGO predictions on some genes with isoform-specific functions (BRCA1, MADD,VAMP7 and ITSN1) and they were coherent with the literature. Besides, we examined whether the main isoform of each gene -as predicted by APPRIS- was the most likely to have the annotated gene functions and it occurs in 99.4% of the genes. We also evaluated the predictions for isoform-specific functions provided by the CAFA3 challenge and results were also convincing. To make these results available to the scientific community, we have deployed a web application to consult ISOGO predictions (https://biotecnun.unav.es/app/isogo). Initial data, website link, isoform-specific GO function predictions and R code is available at https://gitlab.com/icassol/isogo.
Topics: Algorithms; Alternative Splicing; Gene Ontology; Humans; Molecular Sequence Annotation; Open Reading Frames; Protein Isoforms
PubMed: 31974522
DOI: 10.1038/s41598-020-57974-z