A novel myogenic function residing in the 5' non-coding region of Insulin receptor substrate-1 (Irs-1) transcript.
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ABSTRACT:
SUBMITTER: Nagano H
PROVIDER: S-EPMC4373113 | biostudies-literature | 2015
REPOSITORIES: biostudies-literature
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Project description:Insulin-like growth factors (IGFs) are well known to play essential roles in enhancement of myogenic differentiation. In this report we showed that initial IGF-I signal activation but long-term IGF-1 signal termination are required for myogenic differentiation. L6 myoblast stably transfected with myc-epitope tagged insulin receptor substrate-1, myc-IRS-1 (L6-mIRS1) was unable to differentiate into myotubes, indicating that IRS-1 constitutive expression inhibited myogenesis. To elucidate the molecular mechanisms underlying myogenic inhibition, IGF-I signaling was examined. IGF-I treatment of control L6 cells for 18 h resulted in a marked suppression of IGF-I stimulated IRS-1 association with the p85 PI 3-kinase and suppression of activation of Akt that correlated with a down regulation of IRS-1 protein. L6-mIRS1 cells, in contrast, had sustained high levels of IRS-1 protein following 18 h of IGF-I treatment with persistent p85 PI 3-kinase association with IRS-1, Akt phosphorylation and phosphorylation of the downstream Akt substrate, Foxo1. Consistent with Foxo1 phosphorylation, Foxo1 protein was excluded from the nuclei in L6-mIRS1 cells, whereas Foxo1 was localized in the nuclei in control L6 cells during induction of differentiation. In addition, L6 cells stably expressing a dominant-interfering form of Foxo1, Δ256Foxo1 (L6-Δ256Foxo1) were unable to differentiate into myotubes. Together, these data demonstrate that IGF-I regulation of Foxo1 nuclear localization is essential for the myogenic program in L6 cells but that persistent activation of IGF-1 signaling pathways results in a negative feedback to prevent myogenesis.
| S-EPMC3185002 | biostudies-literature
Project description:BackgroundVascular endothelial growth factor-A (VEGF) is one of the key regulators of tumor development, hence it is considered to be an important therapeutic target for cancer treatment. However, clinical trials have suggested that anti-VEGF monotherapy was less effective than standard chemotherapy. On the basis of the evidence, we hypothesized that vegf mRNA may have unrecognized function(s) in cancer cells.Methods and findingsKnockdown of VEGF with vegf-targeting small-interfering (si) RNAs increased susceptibility of human colon cancer cell line (HCT116) to apoptosis caused with 5-fluorouracil, etoposide, or doxorubicin. Recombinant human VEGF165 did not completely inhibit this apoptosis. Conversely, overexpression of VEGF165 increased resistance to anti-cancer drug-induced apoptosis, while an anti-VEGF165-neutralizing antibody did not completely block the resistance. We prepared plasmids encoding full-length vegf mRNA with mutation of signal sequence, vegf mRNAs lacking untranslated regions (UTRs), or mutated 5'UTRs. Using these plasmids, we revealed that the 5'UTR of vegf mRNA possessed anti-apoptotic activity. The 5'UTR-mediated activity was not affected by a protein synthesis inhibitor, cycloheximide. We established HCT116 clones stably expressing either the vegf 5'UTR or the mutated 5'UTR. The clones expressing the 5'UTR, but not the mutated one, showed increased anchorage-independent growth in vitro and formed progressive tumors when implanted in athymic nude mice. Microarray and quantitative real-time PCR analyses indicated that the vegf 5'UTR-expressing tumors had up-regulated anti-apoptotic genes, multidrug-resistant genes, and growth-promoting genes, while pro-apoptotic genes were down-regulated. Notably, expression of signal transducers and activators of transcription 1 (STAT1) was markedly repressed in the 5'UTR-expressing tumors, resulting in down-regulation of a STAT1-responsive cluster of genes (43 genes). As a result, the tumors did not respond to interferon (IFN)alpha therapy at all. We showed that stable silencing of endogenous vegf mRNA in HCT116 cells enhanced both STAT1 expression and IFNalpha responses.ConclusionsThese findings suggest that cancer cells have a survival system that is regulated by vegf mRNA and imply that both vegf mRNA and its protein may synergistically promote the malignancy of tumor cells. Therefore, combination of anti-vegf transcript strategies, such as siRNA-based gene silencing, with anti-VEGF antibody treatment may improve anti-cancer therapies that target VEGF.
| S-EPMC2386836 | biostudies-literature
Project description:Insulin/IGF-1 action is driven by a complex and highly integrated signalling network. Loss-of-function studies indicate that the major insulin/IGF-1 receptor substrate (IRS) proteins, IRS-1 and IRS-2, mediate different biological functions in vitro and in vivo, suggesting specific signalling properties despite their high degree of homology. To identify mechanisms contributing to the differential signalling properties of IRS-1 and IRS-2 in the mediation of insulin/IGF-1 action, we performed comprehensive mass spectrometry (MS)-based phosphoproteomic profiling of brown preadipocytes from wild type, IRS-1-/- and IRS-2-/- mice in the basal and IGF-1-stimulated states. We applied stable isotope labeling by amino acids in cell culture (SILAC) for the accurate quantitation of changes in protein phosphorylation. We found ~10% of the 6262 unique phosphorylation sites detected to be regulated by IGF-1. These regulated sites included previously reported substrates of the insulin/IGF-1 signalling pathway, as well as novel substrates including Nuclear Factor I X and Semaphorin-4B. In silico prediction suggests the protein kinase B (PKB), protein kinase C (PKC), and cyclin-dependent kinase (CDK) as the main mediators of these phosphorylation events. Importantly, we found preferential phosphorylation patterns depending on the presence of either IRS-1 or IRS-2, which was associated with specific sets of kinases involved in signal transduction downstream of these substrates such as PDHK1, MAPK3, and PKD1 for IRS-1, and PIN1 and PKC beta for IRS-2. Overall, by generating a comprehensive phosphoproteomic profile from brown preadipocyte cells in response to IGF-1 stimulation, we reveal both common and distinct insulin/IGF-1 signalling events mediated by specific IRS proteins.
| S-EPMC5951756 | biostudies-literature
Project description:The insulin-like growth factor-1 (IGF-1) signaling pathway has been implicated in non-small cell lung cancer (NSCLC) outcomes and resistance to targeted therapies. However, little is known regarding the molecular mechanisms by which this pathway contributes to the biology of NSCLC. The insulin receptor substrate (IRS) proteins are cytoplasmic adaptor proteins that signal downstream of the IGF-1R and determine the functional outcomes of this signaling pathway. In this study, we assessed the expression patterns of IRS-1 and IRS-2 in NSCLC to identify associations between IRS-1 and IRS-2 expression levels and survival outcomes in the two major histological subtypes of NSCLC, adenocarcinoma (ADC) and squamous cell carcinoma (SCC). High IRS-2 expression was significantly associated with decreased overall survival in adenocarcinoma (ADC) patients, whereas low IRS-1 cytoplasmic expression showed a trend toward association with decreased overall survival in squamous cell carcinoma (SCC) patients. Tumors with low IRS-1 and high IRS-2 expression were found to be associated with poor outcomes in ADC and SCC, indicating a potential role for IRS-2 in the aggressive behavior of NSCLC. Our results suggest distinct contributions of IRS-1 and IRS-2 to the biology of ADC and SCC that impact disease progression.
| S-EPMC6687170 | biostudies-literature
Project description:BackgroundInsulin receptor substrate (Irs) proteins are essential for insulin signaling as they allow downstream effectors to dock with, and be activated by, the insulin receptor. A family of four Irs proteins have been identified in mice, however the gene for one of these, IRS3, has been pseudogenized in humans. While it is known that the Irs gene family originated in vertebrates, it is not known when it originated and which members are most closely related to each other. A better understanding of the evolution of Irs genes and proteins should provide insight into the regulation of metabolism by insulin.ResultsMultiple genes for Irs proteins were identified in a wide variety of vertebrate species. Phylogenetic and genomic neighborhood analyses indicate that this gene family originated very early in vertebrae evolution. Most Irs genes were duplicated and retained in fish after the fish-specific genome duplication. Irs genes have been lost of various lineages, including Irs3 in primates and birds and Irs1 in most fish. Irs3 and Irs4 experienced an episode of more rapid protein sequence evolution on the ancestral mammalian lineage. Comparisons of the conservation of the proteins sequences among Irs paralogs show that domains involved in binding to the plasma membrane and insulin receptors are most strongly conserved, while divergence has occurred in sequences involved in interacting with downstream effector proteins.ConclusionsThe Irs gene family originated very early in vertebrate evolution, likely through genome duplications, and in parallel with duplications of other components of the insulin signaling pathway, including insulin and the insulin receptor. While the N-terminal sequences of these proteins are conserved among the paralogs, changes in the C-terminal sequences likely allowed changes in biological function.
| S-EPMC5482937 | biostudies-literature
Project description:Insulin receptor substrate-1 (IRS-1) plays a pivotal role in insulin signaling, therefore its degradation is exquisitely regulated. Here, we show that insulin-stimulated degradation of IRS-1 requires the presence of a highly conserved Ser/Thr-rich domain that we named domain involved in degradation of IRS-1 (DIDI). DIDI (amino acids 386-430 of IRS-1) was identified by comparing the intracellular degradation rate of several truncated forms of IRS-1 transfected into CHO cells. The isolated DIDI domain underwent insulin-stimulated Ser/Thr phosphorylation, suggesting that it serves as a target for IRS-1 kinases. The effects of deletion of DIDI were studied in Fao rat hepatoma and in CHO cells expressing Myc-IRS-1(WT) or Myc-IRS-1(Δ386-430). Deletion of DIDI maintained the ability of IRS-1(Δ386-434) to undergo ubiquitination while rendering it insensitive to insulin-induced proteasomal degradation, which affected IRS-1(WT) (80% at 8 h). Consequently, IRS-1(Δ386-434) mediated insulin signaling (activation of Akt and glycogen synthesis) better than IRS-1(WT). IRS-1(Δ386-434) exhibited a significant greater preference for nuclear localization, compared with IRS-1(WT). Higher nuclear localization was also observed when cells expressing IRS-1(WT) were incubated with the proteasome inhibitor MG-132. The sequence of DIDI is conserved more than 93% across species, from fish to mammals, as opposed to approximately 40% homology of the entire IRS-1. These findings implicate DIDI as a novel, highly conserved domain of IRS-1, which mediates its cellular localization, rate of degradation, and biological activity, with a direct impact on insulin signal transduction.
| S-EPMC5417385 | biostudies-literature
Project description:Muscle atrophy occurs under various catabolic conditions, including insulin deficiency, insulin resistance, or increased levels of glucocorticoids. This results from reduced levels of insulin receptor substrate 1 (IRS-1), leading to decreased phosphatidylinositol 3-kinase activity and thereby activation of FoxO transcription factors. However, the precise mechanism of reduced IRS-1 under a catabolic condition is unknown. Here, we report that C1-Ten is a novel protein tyrosine phosphatase (PTPase) of IRS-1 that acts as a mediator to reduce IRS-1 under a catabolic condition, resulting in muscle atrophy. C1-Ten preferentially dephosphorylated Y612 of IRS-1, which accelerated IRS-1 degradation. These findings suggest a novel type of IRS-1 degradation mechanism which is dependent on C1-Ten and extends our understanding of the molecular mechanism of muscle atrophy under catabolic conditions. C1-Ten expression is increased by catabolic glucocorticoid and decreased by anabolic insulin. Reflecting these hormonal regulations, the muscle C1-Ten is upregulated in atrophy but downregulated in hypertrophy. This reveals a previously unidentified role of C1-Ten as a relevant PTPase contributing to skeletal muscle atrophy.
| S-EPMC3624259 | biostudies-literature
Project description:Pregnant women with gestational diabetes mellitus (GDM) and type 2 diabetes mellitus (T2DM) share a common pathophysiology associated with similar risk factors. Genetic variants used to determine the risk of developing T2DM might also be associated with the prevalence of GDM. The aim of the present study was to scrutinize the relationship between the G972R polymorphism of the insulin receptor substrate-1 (IRS-1) gene with GDM in the Saudi female population. This is a case-control study that monitored 500 Saudi women. Subjects with GDM (n = 200) were compared with non-GDM (n = 300) controls. We opted to evaluate rs1801278 polymorphism in the IRS1 gene, which plays a critical role in the insulin-signaling pathway. Genotyping was performed with the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) method. The frequency of the rs1801278 polymorphism was significantly higher in women with GDM than in women with non-GDM (for TT + CT versus CC: P = 0.02). Additionally, there was a significant increase in the frequency of the Arg-encoding mutant allele from GDM to non-GDM (for T versus C: P = 0.01). Our results suggest that the rs1801278 polymorphism in the IRS-1 gene is involved in the occurrence of GDM in the Saudi population.
| S-EPMC3948357 | biostudies-literature
Project description:We have reported previously that renal hemodynamic abnormalities exist in the prediabetic stage of type II diabetic rats. At this prediabetic stage these rats have hyperinsulinemia, insulin resistance and metabolic syndrome. It is well known that insulin resistance is frequently associated with renal abnormalities, but the mechanism underlying this association has remained speculative. Although insulin is known to modify renal hemodynamics, little is known about the roles of insulin receptor substrates (IRS1, IRS2) in the renal actions of insulin. To address this issue, the effects of insulin on renal function and renal hemodynamics were investigated in C57BL/6 (WT: wild type), insulin receptor substrate 1- knockout (IRS1-/-), and IRS2-knockout (IRS2-/-) mice. IRS2-/-mice had elevated glucose level as expected. 24-h urine collections and serum creatinine revealed that creatinine clearance did not significantly differ between these groups. Albuminuria was found in IRS1-/-and IRS2-/-groups. We examined the effects on the IRS during the administration of Losartan, which is widely used for diabetic nephropathy. After the administration of Losartan the IRS displayed improved renal hemodynamics. Moreover, the subjects were also given Pioglitazone, which improves insulin resistance. Losartan significantly reduced albuminuria in both groups. Pioglitazone also showed similar results. We assessed the autoregulatory responses of the total renal blood flow (RBF), the superficial (SBF) and the deep renal cortical blood flow (DBF) with stepwise reductions of renal perfusion pressure (RPP), which was induced by a manual clamp on the abdominal aorta. During the clamp induced reductions of the RPP by 10 to 20mm HG, RBF, SBF and the DBF fell significantly more in the IRS1 and IRS2 than in the WT mice. Furthermore micropuncture studies showded that compared to the WT tubuloglomerular feedback (TGF) responses of the stop flow pressure (Psf) were reduced in both the IRS1 -/- and IRS2 -/-. The results of the IRS1 and IRS2 mice displayed the pressence of hemodynamic abnormalities. Losartan and Pioglitazone have shown the potential to improve these abnormalities. In conclusion the results indicate that IRS plays a major role in the stimulation of renal functions and renal hemodynamics in type type II diabetes.
| S-EPMC7714100 | biostudies-literature
Project description:AbstractLong non-coding RNAs (lncRNAs) are a family of non-protein-coding RNAs that span a length of over 200 nucleotides. Research reports have illustrated that lncRNAs are involved in various cellular processes and that their abnormal expression leads to the occurrence and development of various tumors. Colon cancer-associated transcript 2 (CCAT2) was first reported as an oncogene in colon cancer. LncRNA CCAT2 is abnormally expressed in hepatocellular carcinoma, cholangiocarcinoma, lung cancer, breast cancer, ovarian cancer, glioma, and other tumors. In tumor tissues, abnormally overexpressed CCAT2 can affect cell proliferation, migration, epithelial-mesenchymal transition, apoptosis, and other biological behaviors through endogenous RNAs mechanisms, various signaling pathways, transcriptional regulation, and other complex mechanisms. Additionally, the overexpression of CCAT2 is also closely related to the tumor size, tumor node metastasis (TNM) stage, survival time, and other prognostic factors, suggesting that it is a potential prognostic indicator. This article reviews the biological functions of CCAT2 and its mechanisms of action in tumors from previous studies. In this review, we attempt to provide a molecular basis for future clinical applications of lncRNA CCAT2.
| S-EPMC9945556 | biostudies-literature