-
Current Topics in Microbiology and... 2022Macrophages display an array of activation phenotypes depending on the activation signal and the cellular microenvironment. The type and magnitude of the response depend...
Macrophages display an array of activation phenotypes depending on the activation signal and the cellular microenvironment. The type and magnitude of the response depend on signaling molecules as well as on the epigenetic and metabolic status of the cells at the time of activation. The AKT family of kinases consists of three isoforms encoded by independent genes possessing similar functions and structures. Generation of research tools such as isoform-specific gene deletion mutant mice and cells and isoform-specific antibodies has allowed us to understand the role of each kinase isoform in macrophage activation and homeostasis. This chapter discusses the current evidence on the role of AKT kinases in macrophage activation, polarization, and homeostasis, highlighting the gaps in knowledge and future challenges in the field.
Topics: Animals; Macrophage Activation; Macrophages; Mice; Protein Isoforms; Proto-Oncogene Proteins c-akt; Signal Transduction
PubMed: 36243844
DOI: 10.1007/978-3-031-06566-8_7 -
Philosophical Transactions of the Royal... Nov 2023Protein isoforms, generated through alternative splicing or promoter usage, contribute to tissue function. Here, we characterize the expression of predicted and...
Protein isoforms, generated through alternative splicing or promoter usage, contribute to tissue function. Here, we characterize the expression of predicted and isoforms in hair follicles and describe expression of , a hitherto unknown PADI2 isoform, in the oligodendrocyte lineage. transcription is initiated from a downstream intronic promoter, generating an N-terminally truncated, unstable, PADI2β. By contrast to the established role of the canonical PADI2 (PADI2α) (Falcao . 2019 . , 1090-1102.e10. (doi:10.1016/j.celrep.2019.03.108)), PADI2β inhibits oligodendrocyte differentiation, suggesting that PADI2 isoforms exert opposing effects on oligodendrocyte lineage progression. We localize and to developing hair follicles and find that both transcripts are expressed at low levels in progenitor cells, only to increase in expression concomitant with differentiation. When expressed , PADI3α and PADI3β are enriched in the cytoplasm and precipitate together. Whereas PADI3β protein stability is low and PADI3β fails to induce protein citrullination, we find that the enzymatic activity and protein stability of PADI3α is reduced in the presence of PADI3β. We propose that PADI3β modulates PADI3α activity by direct binding and heterodimer formation. Here, we establish expression and function of and isoforms, expanding on the mechanisms in place to regulate citrullination in complex tissues. This article is part of the Theo Murphy meeting issue 'The virtues and vices of protein citrullination'.
Topics: Protein-Arginine Deiminases; Cell Differentiation; Protein Isoforms
PubMed: 37778375
DOI: 10.1098/rstb.2022.0451 -
Scientific Reports Jun 2022Sepsis is caused by an uncontrolled inflammatory response, whose underlying mechanisms are not fully understood. It is well known that the majority of human genes can be...
Sepsis is caused by an uncontrolled inflammatory response, whose underlying mechanisms are not fully understood. It is well known that the majority of human genes can be expressed as alternative isoforms. While isoform switching is implicated in many diseases and is particularly prominent in cancer, it has never been reported in the context of sepsis. Patients presented to the emergency department of three tertiary care hospitals from January 2020 to December 2020 were enrolled. Clinical variables and genome-wide transcriptome of peripheral blood mononuclear cells (PBMC) were obtained. Isoform switching analysis were performed to identify significant isoform switches and relevant biological consequences. A total of 48 subjects with sepsis, involving 42 survivors and 6 non-survivors, admitted to the emergency department of three tertiary care hospitals were enrolled in this study. PBMCs were extracted for RNA sequencing (RNA-seq). Patients (n = 4) with mild stroke or acute coronary syndrome without infection were enrolled in this study as controls. The most frequent functional changes resulting from isoform switching were changes affecting the open reading frame, protein domains and intron retention. Many genes without differences in gene expression showed significant isoform switching. Many genes with significant isoform switches ([Formula: see text]> 0.1) were associated with higher mortality risk, including PIGS, CASP3, LITAF, HBB and RUVBL2. The study for the first time described the landscape of isoform switching in sepsis, including differentially expressed isoform fractions between patients with and without sepsis and survivors and nonsurvivors. The biological consequences of isoform switching, including protein domain loss, signal peptide gain, and intron retention, were identified.
Topics: ATPases Associated with Diverse Cellular Activities; Carrier Proteins; Cohort Studies; DNA Helicases; Gene Expression Profiling; Humans; Leukocytes, Mononuclear; Protein Isoforms; Sepsis
PubMed: 35715539
DOI: 10.1038/s41598-022-14231-9 -
The Journal of Biological Chemistry Jan 2023TMEM63B is a mechanosensitive cation channel activated by hypoosmotic stress and mechanic stimulation. We recently reported a brain-specific alternative splicing of exon...
TMEM63B is a mechanosensitive cation channel activated by hypoosmotic stress and mechanic stimulation. We recently reported a brain-specific alternative splicing of exon 4 in TMEM63B. The short variant lacking exon 4, which constitutes the major isoform in the brain, exhibits enhanced responses to hypoosmotic stimulation compared to the long isoform containing exon 4. However, the mechanisms affecting this differential response are unclear. Here, we showed that the short isoform exhibited stronger cell surface expression compared to the long variant. Using mutagenesis screening of the coding sequence of exon 4, we identified an RXR-type endoplasmic reticulum (ER) retention signal (RER). We found that this motif was responsible for binding to the COPI retrieval vesicles, such that the longer TMEM63B isoforms were more likely to be retrotranslocated to the ER than the short isoforms. In addition, we demonstrated long TMEM63Bs could form heterodimers with short isoforms and reduce their surface expression. Taken together, our findings revealed an ER retention signal in the alternative splicing domain of TMEM63B that regulates the surface expression of TMEM63B protein and channel function.
Topics: Alternative Splicing; Cations; Cell Membrane; Endoplasmic Reticulum; Protein Isoforms; Membrane Proteins; Gene Expression Regulation
PubMed: 36496074
DOI: 10.1016/j.jbc.2022.102781 -
Integrative and Comparative Biology Dec 2022Hibernation in brown bears is an annual process involving multiple physiologically distinct seasons-hibernation, active, and hyperphagia. While recent studies have...
Hibernation in brown bears is an annual process involving multiple physiologically distinct seasons-hibernation, active, and hyperphagia. While recent studies have characterized broad patterns of differential gene regulation and isoform usage between hibernation and active seasons, patterns of gene and isoform expression during hyperphagia remain relatively poorly understood. The hyperphagia stage occurs between active and hibernation seasons and involves the accumulation of large fat reserves in preparation for hibernation. Here, we use time-series analyses of gene expression and isoform usage to interrogate transcriptomic regulation associated with all three seasons. We identify a large number of genes with significant differential isoform usage (DIU) across seasons and show that these patterns of isoform usage are largely tissue-specific. We also show that DIU and differential gene-level expression responses are generally non-overlapping, with only a small subset of multi-isoform genes showing evidence of both gene-level expression changes and changes in isoform usage across seasons. Additionally, we investigate nuanced regulation of candidate genes involved in the insulin signaling pathway and find evidence of hyperphagia-specific gene expression and isoform regulation that may enhance fat accumulation during hyperphagia. Our findings highlight the value of using temporal analyses of both gene- and isoform-level gene expression when interrogating complex physiological phenotypes and provide new insight into the mechanisms underlying seasonal changes in bear physiology.
Topics: Animals; Ursidae; Hibernation; Hyperphagia; Protein Isoforms; Transcriptome; Seasons
PubMed: 35709393
DOI: 10.1093/icb/icac093 -
Stem Cell Reviews and Reports Aug 2021OCT4 plays critical roles in self-renewal and pluripotency maintenance of embryonic stem cells, and is considered as one of the main stemness markers. It also has... (Review)
Review
OCT4 plays critical roles in self-renewal and pluripotency maintenance of embryonic stem cells, and is considered as one of the main stemness markers. It also has pivotal roles in early stages of embryonic development. Most studies on OCT4 have focused on the expression and function of OCT4A, which is the biggest isoform of OCT4 known so far. Recently, many studies have shown that OCT4 has various transcript variants, protein isoforms, as well as pseudogenes. Distinguishing the expression and function of these variants and isoforms is a big challenge in expression profiling studies of OCT4. Understanding how OCT4 is functioning in different contexts, depends on knowing of where and when each of OCT4 transcripts, isoforms and pseudogenes are expressed. Here, we review OCT4 known transcripts, isoforms and pseudogenes, as well as its interactions with other proteins, and emphasize the importance of discriminating each of them in order to understand the exact function of OCT4 in stem cells, normal development and development of diseases.
Topics: DNA; Embryonic Stem Cells; Humans; Octamer Transcription Factor-3; Protein Isoforms; RNA
PubMed: 33389631
DOI: 10.1007/s12015-020-10098-3 -
American Journal of Physiology. Heart... Oct 2022Protein kinase C-α (PKCα) plays a major role in a diverse range of cellular processes. Studies to date have defined the regulatory controls and function of PKCα...
Protein kinase C-α (PKCα) plays a major role in a diverse range of cellular processes. Studies to date have defined the regulatory controls and function of PKCα entirely based upon the previously annotated ubiquitously expressed prototypical isoform. From RNA-seq-based transcriptome analysis in murine heart, we identified a previously unannotated PKCα variant produced by alternative RNA splicing. This PKCα transcript variant, which we named PKCα-novel exon (PKCα-NE), contains an extra exon between exon 16 and exon 17, and is specifically detected in adult mouse cardiac and skeletal muscle, but not other tissues; it is also detected in human hearts. This transcript variant yields a PKCα isoform with additional 16 amino acids inserted in its COOH-terminal variable region. Although the canonical PKCα enzyme is a lipid-dependent kinase, in vitro kinase assays show that PKCα-NE displays a high level of basal lipid-independent catalytic activity. Our unbiased proteomic analysis identified a specific interaction between PKCα-NE and eukaryotic elongation factor-1α (eEF1A1). Studies in cardiomyocytes link PKCα-NE expression to an increase in eEF1A1 phosphorylation and elevated protein synthesis. In summary, we have identified a previously uncharacterized muscle-specific PKCα splicing variant, PKCα-NE, with distinct biochemical properties that plays a unique role in the control of the protein synthesis machinery in cardiomyocytes. PKCα is an important signaling molecule extensively studied in many cellular processes. However, no isoforms have been reported for PKCα except one prototypic isoform. Alternative mRNA splicing of gene was detected for the first time in rodent and human cardiac tissue, which can produce a previously unknown PKCα-novel exon (NE) isoform. The biochemistry and molecular effects of PKCα-NE are markedly different from PKCα wild type, suggesting potential functional diversity of PKCα signaling in muscle.
Topics: Adult; Alternative Splicing; Amino Acids; Animals; Humans; Lipids; Mice; Muscle, Skeletal; Myocytes, Cardiac; Protein Isoforms; Protein Kinase C-alpha; Proteomics; RNA, Messenger
PubMed: 36112502
DOI: 10.1152/ajpheart.00304.2022 -
Molecules (Basel, Switzerland) Dec 2021After being rather neglected as a research field in the past, carbonic anhydrase activators (CAAs) were undoubtedly demonstrated to be useful in diverse pharmaceutical... (Review)
Review
After being rather neglected as a research field in the past, carbonic anhydrase activators (CAAs) were undoubtedly demonstrated to be useful in diverse pharmaceutical and industrial applications. They also improved the knowledge of the requirements to selectively interact with a CA isoform over the others and confirmed the catalytic mechanism of this class of compounds. Amino acid and amine derivatives were the most explored in in vitro, in vivo and crystallographic studies as CAAs. Most of them were able to activate human or non-human CA isoforms in the nanomolar range, being proposed as therapeutic and industrial tools. Some isoforms are better activated by amino acids than amines derivatives and the stereochemistry may exert a role. Finally, non-human CAs have been very recently tested for activation studies, paving the way to innovative industrial and environmental applications.
Topics: Amines; Amino Acids; Animals; Carbonic Anhydrases; Enzyme Activation; Enzyme Activators; Humans; Models, Molecular; Protein Isoforms
PubMed: 34885917
DOI: 10.3390/molecules26237331 -
Clinical & Experimental Metastasis Jun 2022Cellular plasticity lies at the core of cancer progression, metastasis, and resistance to treatment. Stemness and epithelial-mesenchymal plasticity in cancer are... (Review)
Review
Cellular plasticity lies at the core of cancer progression, metastasis, and resistance to treatment. Stemness and epithelial-mesenchymal plasticity in cancer are concepts that represent a cancer cell's ability to coopt and adapt normal developmental programs to promote survival and expansion. The cancer stem cell model states that a small subset of cancer cells with stem cell-like properties are responsible for driving tumorigenesis and metastasis while remaining especially resistant to common chemotherapeutic drugs. Epithelial-mesenchymal plasticity describes a cancer cell's ability to transition between epithelial and mesenchymal phenotypes which drives invasion and metastasis. Recent research supports the existence of stable epithelial/mesenchymal hybrid phenotypes which represent highly plastic states with cancer stem cell characteristics. The cell adhesion molecule CD44 is a widely accepted marker for cancer stem cells, and it lies at a functional intersection between signaling networks regulating both stemness and epithelial-mesenchymal plasticity. CD44 expression is complex, with alternative splicing producing many isoforms. Interestingly, not only does the pattern of isoform expression change during transitions between epithelial and mesenchymal phenotypes in cancer, but these isoforms have distinct effects on cell behavior including the promotion of metastasis and stemness. The role of CD44 both downstream and upstream of signaling pathways regulating epithelial-mesenchymal plasticity and stemness make this protein a valuable target for further research and therapeutic intervention.
Topics: Alternative Splicing; Epithelial-Mesenchymal Transition; Humans; Hyaluronan Receptors; Neoplasm Metastasis; Neoplasms; Neoplastic Stem Cells; Protein Isoforms
PubMed: 35023031
DOI: 10.1007/s10585-022-10146-x -
Cancer Biomarkers : Section a of... 2022Secreted phosphoprotein 1 (SPP1), also known as osteopontin (OPN), is a multifunctional protein expressed in diverse normal tissues, and functionally is involved in...
BACKGROUND
Secreted phosphoprotein 1 (SPP1), also known as osteopontin (OPN), is a multifunctional protein expressed in diverse normal tissues, and functionally is involved in cellular matrix and signaling processes. Many studies have linked SPP1 to pathophysiological conditions including cancer.
OBJECTIVE
The aim of this study is to evaluate the 3'UTR length of SPP1 gene in glioblastoma cell line.
METHODS
3' Rapid Amplification of cDNA End (3'-RACE) was used to determine the 3' end of SPP1 gene. APAatlas data base, GEPIA web server, and miRcode were also used to extract related information and bioinformatic analysis part.
RESULTS
In this study we show that SPP1 gene undergoes Alternative cleavage and Polyadenylation (APA) mechanism, by which it generates two 3' termini, longer isoform and shorter isoform, in glioblastoma derived cell line, U87-MG. Further bioinformatic analysis reveals that SPP1 alternative 3'UTR (aUTR), which is absent in shorter isoform, is targeted by two families of microRNAs-miR-181abcd/4262 and miR-154/872. These miRNAs also target and perhaps negatively regulate NAP1L1 and ENAH genes that are involved in cell proliferation and cell polarity, respectively. Relative expression difference (RED), obtained from RNA-seq data of diverse normal tissues, representing APA usage appears to be negatively correlated with expression of NAP1L1 and ENAH, emphasizing co-expression of SPP1 longer isoform with these two genes, indicating miRNA sponge function of aUTR (longer 3'UTR). Bioinformatic analysis also shows that in normal brain tissue longer APA isoform of SPP1 is expressed; however shorter isoform appears to be expressed in cancer condition.
CONCLUSION
Together, this study reveals that SPP1 APA isoforms have different pattern in normal and cancerous conditions, which can be considered as a diagnostic and prognostic marker in cancers.
Topics: 3' Untranslated Regions; Glioblastoma; Humans; MicroRNAs; Microfilament Proteins; Nucleosome Assembly Protein 1; Osteopontin; Polyadenylation; Protein Isoforms
PubMed: 35275520
DOI: 10.3233/CBM-210135