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Scientific Reports Sep 2023Sorcin (Sri), a member of penta EF-hand protein family plays a diverse role in maintaining calcium homeostasis, cell cycle and vesicular trafficking. Sri is highly...
Sorcin (Sri), a member of penta EF-hand protein family plays a diverse role in maintaining calcium homeostasis, cell cycle and vesicular trafficking. Sri is highly conserved amongst mammals and consists of N-terminal glycine rich domain and C-terminal calcium binding domain that mediates its dimerization and interacts with different compounds. In the present study, with the help of combination of computational and molecular biology techniques, we have identified a novel isoform (Sri-N) in mouse which differs only in the C-terminal domain with that of Sri reported earlier. The novel isoform contains a new last exon that is different from the one present in the reported transcript (Sri). The presence of the novel isoform was further validated in different tissues by RT-PCR and DNA sequencing. The transcript was conceptually translated and subjected to in-silico analysis using different bioinformatics tools. The novel transcript variant encodes for a longer protein isoform without any change in the sub-cellular localization as predicted by PSORT-II online tool. Molecular modelling was performed to compare the structural changes in Sri-N and Sri isoforms. The structural characterization of the novel isoform using MD simulation depicted its overall stability under the physiological conditions. The molecular docking of proteins with various chemotherapeutic drugs revealed that their binding affinity is more for Sri-N as compared to that for the previously reported transcript Sri.
Topics: Animals; Mice; Calcium; Dimerization; Molecular Docking Simulation; Protein Isoforms; Bone Density Conservation Agents; Mammals
PubMed: 37709787
DOI: 10.1038/s41598-023-40913-z -
Scientific Reports Nov 2023During osteoclast differentiation, the expression of the transcription factor nuclear factor of activated T cell 1 (Nfatc1) increases in an autoproliferative manner....
During osteoclast differentiation, the expression of the transcription factor nuclear factor of activated T cell 1 (Nfatc1) increases in an autoproliferative manner. Nfatc1 isoforms are of three sizes, and only the short isoform increases during osteoclast differentiation. Genetic ablation of the whole Nfatc1 gene demonstrated that it is essential for osteoclastogenesis; however, the specific role of the Nfatc1 short form (Nfatc1/αA) remains unknown. In this study, we engineered Nfatc1 short form-specific knockout mice and found that these mice died in utero by day 13.5. We developed a novel osteoclast culture system in which hematopoietic stem cells were cultured, proliferated, and then differentiated into osteoclasts in vitro. Using this system, we show that the Nfatc1/αA isoform is essential for osteoclastogenesis and is responsible for the expression of various osteoclast markers, the Nfatc1 short form itself, and Nfatc1 regulators.
Topics: Mice; Animals; Osteoclasts; NFATC Transcription Factors; T-Lymphocytes; Cell Differentiation; Protein Isoforms; Self-Control; RANK Ligand
PubMed: 37914750
DOI: 10.1038/s41598-023-45909-3 -
BMC Cell Biology Mar 2014Myosin IC is a single headed member of the myosin superfamily that localizes to the cytoplasm and the nucleus and is implicated in a variety of processes in both...
BACKGROUND
Myosin IC is a single headed member of the myosin superfamily that localizes to the cytoplasm and the nucleus and is implicated in a variety of processes in both compartments. We recently identified a novel isoform of myosin IC and showed that the MYOIC gene in mammalian cells encodes three isoforms (isoforms A, B, and C) that differ only in the addition of short isoform-specific N-terminal peptides. The expression pattern of the isoforms and the mechanisms of expression regulation remain unknown.
RESULTS
To determine the expression patterns of myosin IC isoforms, we performed a comprehensive expression analysis of the two myosin IC isoforms (isoform A and B) that contain isoform-specific sequences. By immunoblotting with isoform-specific antibodies and by qRT-PCR with isoform-specific primer we demonstrate that myosin IC isoforms A and B have distinct expression patterns in mouse tissues. Specifically, we show that myosin IC isoform A is expressed in a tissue specific pattern, while myosin IC isoform B is ubiquitously expressed at comparable levels in mouse tissues.
CONCLUSIONS
The differences in the expression profile of the myosin IC isoforms indicate a tissue-specific MYOIC gene regulation and further suggest that the myosin IC isoforms, despite their high sequence homology, might have tissue-specific and isoform-specific functions.
Topics: Animals; Blotting, Western; Female; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Myosins; Protein Isoforms; RNA, Messenger; Real-Time Polymerase Chain Reaction
PubMed: 24618359
DOI: 10.1186/1471-2121-15-8 -
BMC Genomics Mar 2020Most eukaryotic genes produce different transcripts of multiple isoforms by inclusion or exclusion of particular exons. The isoforms of a gene often play diverse...
BACKGROUND
Most eukaryotic genes produce different transcripts of multiple isoforms by inclusion or exclusion of particular exons. The isoforms of a gene often play diverse functional roles, and thus it is necessary to accurately measure isoform expressions as well as gene expressions. While previous studies have demonstrated the strong agreement between mRNA sequencing (RNA-seq) and array-based gene and/or isoform quantification platforms (Microarray gene expression and Exon-array), the more recently developed NanoString platform has not been systematically evaluated and compared, especially in large-scale studies across different cancer domains.
RESULTS
In this paper, we present a large-scale comparative study among RNA-seq, NanoString, array-based, and RT-qPCR platforms using 46 cancer cell lines across different cancer types. The goal is to understand and evaluate the calibers of the platforms for measuring gene and isoform expressions in cancer studies. We first performed NanoString experiments on 59 cancer cell lines with 404 custom-designed probes for measuring the expressions of 478 isoforms in 155 genes, and additional RT-qPCR experiments for a subset of the measured isoforms in 13 cell lines. We then combined the data with the matched RNA-seq, Exon-array, and Microarray data of 46 of the 59 cell lines for the comparative analysis.
CONCLUSION
In the comparisons of the platforms for measuring the expressions at both isoform and gene levels, we found that (1) the agreement on isoform expressions is lower than the agreement on gene expressions across the four platforms; (2) NanoString and Exon-array are not consistent on isoform quantification even though both techniques are based on hybridization reactions; (3) RT-qPCR experiments are more consistent with RNA-seq and Exon-array than NanoString in isoform quantification; (4) different RNA-seq isoform quantification methods show varying estimation results, and among the methods, Net-RSTQ and eXpress are more consistent across the platforms; and (5) RNA-seq has the best overall consistency with the other platforms on gene expression quantification.
Topics: Algorithms; Exons; Gene Expression Profiling; Humans; Oligonucleotide Array Sequence Analysis; Protein Isoforms; Sequence Analysis, RNA; Software
PubMed: 32228441
DOI: 10.1186/s12864-020-6643-8 -
Human Molecular Genetics Oct 2022Aberrant splicing underlies many human diseases, including cancer, cardiovascular diseases and neurological disorders. Genome-wide mapping of splicing quantitative trait... (Review)
Review
Aberrant splicing underlies many human diseases, including cancer, cardiovascular diseases and neurological disorders. Genome-wide mapping of splicing quantitative trait loci (sQTLs) has shown that genetic regulation of alternative splicing is widespread. However, identification of the corresponding isoform or protein products associated with disease-associated sQTLs is challenging with short-read RNA-seq, which cannot precisely characterize full-length transcript isoforms. Furthermore, contemporary sQTL interpretation often relies on reference transcript annotations, which are incomplete. Solutions to these issues may be found through integration of newly emerging long-read sequencing technologies. Long-read sequencing offers the capability to sequence full-length mRNA transcripts and, in some cases, to link sQTLs to transcript isoforms containing disease-relevant protein alterations. Here, we provide an overview of sQTL mapping approaches, the use of long-read sequencing to characterize sQTL effects on isoforms, the linkage of RNA isoforms to protein-level functions and comment on future directions in the field. Based on recent progress, long-read RNA sequencing promises to be part of the human disease genetics toolkit to discover and treat protein isoforms causing rare and complex diseases.
Topics: Humans; Human Genetics; Protein Isoforms; RNA Isoforms; RNA, Messenger; Sequence Analysis, RNA
PubMed: 35960994
DOI: 10.1093/hmg/ddac196 -
Cell Death and Differentiation Dec 2011The TP53 tumour-suppressor gene is expressed as several protein isoforms generated by different mechanisms, including use of alternative promoters, splicing sites and... (Review)
Review
The TP53 tumour-suppressor gene is expressed as several protein isoforms generated by different mechanisms, including use of alternative promoters, splicing sites and translational initiation sites, that are conserved through evolution and within the TP53 homologues, TP63 and TP73. Although first described in the eighties, the importance of p53 isoforms in regulating the suppressive functions of p53 has only become evident in the last 10 years, by analogy with observations that p63 and p73 isoforms appeared indispensable to fully understand the biological functions of TP63 and TP73. This review summarizes recent advances in the field of 'p53 isoforms', including new data on p63 and p73 isoforms. Details of the alternative mechanisms that produce p53 isoforms and cis- and trans-regulators identified are provided. The main focus is on their biological functions (apoptosis, cell cycle, aging and so on) in cellular and animal models, including mouse, zebrafish and Drosophila. Finally, the deregulation of p53 isoform expression in human cancers is reviewed. Based on these latest results, several developments are expected in the future: the identification of drugs modulating p53 isoform expression; the generation of animal models and the evaluation of the use of p53 isoform as biomarkers in human cancers.
Topics: Amino Acid Sequence; Animals; Disease Models, Animal; Evolution, Molecular; Humans; Molecular Sequence Data; Mutation; Neoplasms; Protein Isoforms; Tumor Suppressor Protein p53; Tumor Suppressor Proteins
PubMed: 21941372
DOI: 10.1038/cdd.2011.120 -
Nature Communications Apr 2022Alternative splicing (AS) is a highly-regulated post-transcriptional mechanism known to modulate isoform expression within genes and contribute to cell-type identity....
Alternative splicing (AS) is a highly-regulated post-transcriptional mechanism known to modulate isoform expression within genes and contribute to cell-type identity. However, the extent to which alternative isoforms establish co-expression networks that may be relevant in cellular function has not been explored yet. Here, we present acorde, a pipeline that successfully leverages bulk long reads and single-cell data to confidently detect alternative isoform co-expression relationships. To achieve this, we develop and validate percentile correlations, an innovative approach that overcomes data sparsity and yields accurate co-expression estimates from single-cell data. Next, acorde uses correlations to cluster co-expressed isoforms into a network, unraveling cell type-specific alternative isoform usage patterns. By selecting same-gene isoforms between these clusters, we subsequently detect and characterize genes with co-differential isoform usage (coDIU) across cell types. Finally, we predict functional elements from long read-defined isoforms and provide insight into biological processes, motifs, and domains potentially controlled by the coordination of post-transcriptional regulation. The code for acorde is available at https://github.com/ConesaLab/acorde .
Topics: Alternative Splicing; Protein Isoforms; Sequence Analysis, RNA
PubMed: 35383181
DOI: 10.1038/s41467-022-29497-w -
Biological Chemistry Oct 2007Selenoproteins have been recognized as modulators of brain function and signaling. Phospholipid hydroperoxide glutathione peroxidase (GPx4/PHGPx) is a unique member of... (Review)
Review
Selenoproteins have been recognized as modulators of brain function and signaling. Phospholipid hydroperoxide glutathione peroxidase (GPx4/PHGPx) is a unique member of the selenium-dependent glutathione peroxidases in mammals with a pivotal role in brain development and function. GPx4 exists as a cytosolic, mitochondrial, and nuclear isoform derived from a single gene. In mice, the GPx4 gene is located on chromosome 10 in close proximity to a functional retrotransposome that is expressed under the control of captured regulatory elements. Elucidation of crystallographic data uncovered structural peculiarities of GPx4 that provide the molecular basis for its unique enzymatic properties and substrate specificity. Monomeric GPx4 is multifunctional: it acts as a reducing enzyme of peroxidized phospholipids and thiols and as a structural protein. Transcriptional regulation of the different GPx4 isoforms requires several isoform-specific cis-regulatory sequences and trans-activating factors. Cytosolic and mitochondrial GPx4 are the major isoforms exclusively expressed by neurons in the developing brain. In stark contrast, following brain trauma, GPx4 is specifically upregulated in non-neuronal cells, i.e., reactive astrocytes. Molecular approaches to genetic modification in mice have revealed an essential and isoform-specific function for GPx4 in development and disease. Here we review recent findings on GPx4 with emphasis on its molecular structure and function and consider potential mechanisms that underlie neural development and neuropathological conditions.
Topics: Animals; Brain; Crystallography, X-Ray; Gene Expression Regulation, Enzymologic; Genome; Glutathione Peroxidase; Humans; Models, Molecular; Neurons; Phospholipid Hydroperoxide Glutathione Peroxidase; Protein Isoforms; Signal Transduction; Structure-Activity Relationship; Transcription, Genetic
PubMed: 17937614
DOI: 10.1515/BC.2007.126 -
Bioinformatics (Oxford, England) Jun 2022RNA isoforms contribute to the diverse functionality of the proteins they encode within the cell. Visualizing how isoform expression differs across cell types and brain...
SUMMARY
RNA isoforms contribute to the diverse functionality of the proteins they encode within the cell. Visualizing how isoform expression differs across cell types and brain regions can inform our understanding of disease and gain or loss of functionality caused by alternative splicing with potential negative impacts. However, the extent to which this occurs in specific cell types and brain regions is largely unknown. This is the kind of information that ScisorWiz plots can provide in an informative and easily communicable manner. ScisorWiz affords its user the opportunity to visualize specific genes across any number of cell types, and provides various sorting options for the user to gain different ways to understand their data. ScisorWiz provides a clear picture of differential isoform expression through various clustering methods and highlights features such as alternative exons and single-nucleotide variants. Tools like ScisorWiz are key for interpreting single-cell isoform sequencing data. This tool applies to any single-cell long-read RNA sequencing data in any cell type, tissue or species.
AVAILABILITY AND IMPLEMENTATION
Source code is available at http://github.com/ans4013/ScisorWiz. No new data were generated for this publication. Data used to generate figures was sourced from GEO accession token GSE158450 and available on GitHub as example data.
Topics: Protein Isoforms; Alternative Splicing; Software; RNA Isoforms; Exons; Sequence Analysis, RNA
PubMed: 35604081
DOI: 10.1093/bioinformatics/btac340 -
The Journal of Nutrition May 2015Precursor mRNA (pre-mRNA) splicing is a critical step in gene expression that results in the removal of intronic sequences from immature mRNA, leading to the production... (Review)
Review
Precursor mRNA (pre-mRNA) splicing is a critical step in gene expression that results in the removal of intronic sequences from immature mRNA, leading to the production of mature mRNA that can be translated into protein. Alternative pre-mRNA splicing is the process whereby alternative exons and/or introns are selectively included or excluded, generating mature mRNAs that encode proteins that may differ in function. The resulting alterations in the pattern of protein isoform expression can result in changes in protein-protein interaction, subcellular localization, and flux through metabolic pathways. Although basic mechanisms of pre-mRNA splicing of introns and exons are reasonably well characterized, how these mechanisms are regulated remains poorly understood. The goal of this review is to highlight selected recent advances in our understanding of the regulation of pre-mRNA splicing by nutrients and modulation of nutrient metabolism that result from changes in pre-mRNA splicing.
Topics: Alternative Splicing; Animals; Diet; Energy Intake; Energy Metabolism; Gene Expression Regulation; Humans; Models, Biological; Protein Isoforms; Protein Transport; RNA Precursors; RNA Splicing; RNA, Messenger
PubMed: 25761502
DOI: 10.3945/jn.114.203216