The serine phosphorylations in the IRS-1 PIR domain abrogate IRS-1 and IR interaction.

Serine phosphorylations on insulin receptor substrate 1 (IRS-1) by diverse kinases aoccur widely during obesity-, stress-, and inflammation-induced conditions in models of insulin resistance and type 2 diabetes. In this study, we define a region within the human IRS-1, which is directly C-terminal to the PTB domain encompassing numerous serine phosphorylation sites including Ser307 (mouse Ser302) and Ser312 (mouse 307) creating a phosphorylation insulin resistance (PIR) domain. We demonstrate that the IRS-1 PTB-PIR with its unphosphorylated serine residues interacts with the insulin receptor (IR) but loses the IR-binding when they are phosphorylated. Surface plasmon resonance studies further confirm that the PTB-PIR binds stronger to IR than just the PTB domain, and that phosphorylations at Ser307, Ser312, Ser315, and Ser323 within the PIR domain result in abrogating the binding. Insulin-responsive cells containing the mutant IRS-1 with all these four serines changed into glutamates to mimic phosphorylations show decreased levels of phosphorylations in IR, IRS-1, and AKT compared to the wild-type IRS-1. Hydrogen-deuterium exchange mass spectrometry experiments indicating the PIR domain interacting with the N-terminal lobe and the hinge regions of the IR kinase domain further suggest the possibility that the IRS-1 PIR domain protects the IR from the PTP1B-mediated dephosphorylation.

Kinesin light chain 1 stabilizes insulin receptor substrate 1 to regulate the IGF-1-AKT signaling pathway during myoblast differentiation.

The IGF signaling pathway plays critical role in regulating skeletal myogenesis. We have demonstrated that KIF5B, the heavy chain of kinesin-1 motor, promotes myoblast differentiation through regulating IGF-p38MAPK activation. However, the roles of the kinesin light chain (Klc) in IGF pathway and myoblast differentiation remain elusive. In this study, we found that Klc1 was upregulated during muscle regeneration and downregulated in senescence mouse muscles and dystrophic muscles from mdx (X-linked muscular dystrophic) mice. Gain- and loss-of-function experiments further displayed that Klc1 promotes AKT-mTOR activity and positively regulates myogenic differentiation. We further identified that the expression levels of IRS1, the critical node of IGF-1 signaling, are downregulated in Klc1-depleted myoblasts. Coimmunoprecipitation study revealed that IRS1 interacted with the 88-154 amino acid sequence of Klc1 via its PTB domain. Notably, the reduced Klc1 levels were found in senescence and osteoporosis skeletal muscle samples from both mice and human. Taken together, our findings suggested a crucial role of Klc1 in the regulation of IGF-AKT pathway during myogenesis through stabilizing IRS1, which might ultimately influence the development of muscle-related disorders.

IRS1 promotes thyroid cancer metastasis through EMT and PI3K/AKT pathways.

OBJECTIVE: Insulin receptor substract 1 (IRS1) protein is an important signal transduction adapter for extracellular signal transduction from insulin-like growth factor-1 receptor and its family members to IRS1 downstream proteins. IRS1 has been reported to be involved in tumourigenesis and metastasis in some of solid tumors. Investigating the role of IRS1 in thyroid cancer can help to screen high risk patients at the initial diagnosis. DESIGN, PATIENTS AND MEASUREMENTS: Immunohistochemical assay was used to detect the expression levels of IRS1 in 131 metastatic thyroid cancer tissues. Wound healing, cell invasion and colony formation assays were used to study the functions of IRS1 in vitro. RNA sequencing (RNA-seq) and Western blot analysis analyses were performed to examine the underlying regulation mechanisms of IRS1 in thyroid cancer cells. RESULTS: IRS1 was highly expressed in thyroid cancers and its expression was positively associated with distant metastasis and advanced clinical stages. In vitro studies demonstrated that IRS1 is an important mediator of migration, invasion and colony formation of thyroid cancer cells. RNA-seq showed that IRS1 promoted the metastasis of thyroid cancer by regulating epithelial-mesenchymal transition and phosphoinositide 3-kinase (PI3K)/AKT pathway. CONCLUSIONS: IRS1 overexpression contributes to the aggressiveness of thyroid cancer and is expected to be a stratified marker and a potential therapeutic target for thyroid cancer.

HDAC inhibitors target IRS4 to enhance anti­AR therapy in AR­positive triple­negative breast cancer.

Triple­negative breast cancer (TNBC) is the most malignant subtype of breast cancer. Androgen receptor (AR) has been identified as a potential therapeutic target for AR­positive TNBC; however, clinical trials have not yet produced an effective treatment. The present study aimed to identify a novel treatment regimen to improve the prognosis of AR­positive TNBC. First, a combination of an AR inhibitor (enzalutamide, Enz) and a selective histone deacetylase inhibitor (chidamide, Chid) was used to treat AR­positive TNBC cell lines, and a synergistic effect of these drugs was observed. The combination treatment inhibited cell proliferation and migration by arresting the cell cycle at the G2/M phase. Subsequently, next­generation sequencing was performed to detect changes in gene regulation. The results showed that the PI3K/Akt signalling pathway was significantly inhibited by the combination treatment of Enz and Chid. Gene Set Enrichment Analysis revealed that the combination group was significantly enriched in KRAS signalling. Analysis of the associated genes revealed that insulin receptor substrate 4 (IRS4) may have a critical role in blocking the activation of KRAS signalling. In a mouse xenograft model, combination treatment also inhibited the PI3K/Akt signalling pathway by upregulating the expression of IRS4 and thereby suppressing tumour growth. In conclusion, the results of the present study revealed that combination treatment with Enz and Chid can upregulate IRS4, which results in the blocking of KRAS signalling and suppression of tumour growth. It may be hypothesised that the expression levels of IRS4 could be used as a biomarker for screening patients with AR­positive TNBC using Enz and Chid combination therapy.

Polysaccharides from Dendrobium officinale improve obesity-induced insulin resistance through the gut microbiota and the SOCS3-mediated insulin receptor substrate-1 signaling pathway.

BACKGROUND: Obesity induces insulin resistance and chronic inflammation, impacting human health. The relationship between obesity, gut microbiota, and regulatory mechanisms has been studied extensively. Dendrobium officinale polysaccharide (DOP), a traditional Chinese herbal medicine, potentially reduces insulin resistance. However, the mechanism through which DOP affects gut microbiota and alleviates obesity-induced insulin resistance in rats requires further investigation. RESULTS: The current study aimed to assess the impact of DOP on gut microbiota and insulin resistance in rats on a high-fat diet. The results revealed that DOP effectively reduced blood lipids, glucose disorders, oxidative stress, and inflammatory infiltration in the liver of obese Sprague Dawley rats. This was achieved by downregulating SOCS3 expression and upregulating insulin receptor substrate-1 (IRS-1) by regulating the JAK/STAT/SOCS3 signaling pathway. Notably, DOP intervention enhanced the abundance of beneficial gut microbiota and reduced harmful microbiota. Correlation analysis demonstrated significant associations among intestinal microbiota, SOCS3-mediated IRS-1 expression, and inflammatory factors. CONCLUSION: Dendrobium officinale polysaccharide regulated the gut microbiota, enhanced IRS-1 expression, and mitigated liver injury and insulin resistance due to a high-fat diet. These findings depict the potential anti-insulin resistance properties of DOP and offer further evidence for addressing obesity and its complications. © 2023 Society of Chemical Industry.

Gender Differences of Gly972Arg Polymorphism of the IRS-1 Gene Related to Cardiovascular Disease Risk Factors Among Indonesians.

BACKGROUND: Cardiovascular disease is driven by traditional risk factors, sex, and genetic differences. The Asian population, specifically Indonesians, has been known at high risk of insulin resistance and endothelial dysfunction. A possible genetic risk factor related to cardiovascular diseases is Gly972Arg polymorphism of insulin receptor substrate 1 (IRS-1) gene, as this impairs endothelial function. To date, whether there is a gender difference in Gly972Arg polymorphism of the IRS-1 gene in Indonesians is unknown. This study aimed to to define whether there is a gender difference in Gly972Arg polymorphism of the IRS-1 gene in Indonesians. METHODS: We studied adults living in two areas (rural and urban) in Indonesia. We collected demographic and clinical data from the study subjects. Gly972Arg polymorphism of the IRS-1 gene (rs1801278) was detected using TaqMan real-time polymerase chain reaction. RESULTS: A total of 378 subjects were recruited. The wild-type allele (CC) was found in 86 (22.8%) subjects, heterozygous mutant allele (CT) in 245 (64.8%), and homozygous mutant allele in 47 (12.4%). The proportion of subjects with T alleles was significantly higher among women than men (54.6% vs. 45.4%, odds ratio: 1.89; p = 0.01). Subjects with T allele more often have hypertension (odds ratio: 1.69, p = 0.058). CONCLUSION: There were a higher proportion of women than men carrying the T allele of Gly972Arg polymorphism among Indonesians. Individuals with the T allele appeared to show a greater prevalence of hypertension. These results may explain a possible mechanism of the high prevalence of metabolic syndrome in Indonesia, especially in women.

Phenolic Constituents from Black Quinoa Alleviate Insulin Resistance in HepG2 Cells via Regulating IRS1/PI3K/Akt/GLUTs Signaling Pathways.

Quinoa is a nutrient-rich pseudocereal with a lower glycemic index and glycemic load. However, its therapeutic potency and underlying mechanism against insulin resistance (IR) have not been fully elucidated. In this work, network pharmacology was applied to screen IR targets and their related pathways. The efficacy and mechanism of black quinoa polyphenols (BQP) on IR improvement were evaluated and uncovered based on the IR model in vitro combined with molecular docking. Ten phenolic constituents of BQP were detected, and the network pharmacology results show that PI3K/Akt pathways are the main pathways in BQP against IR. The in vitro assay proved that BQP increases the glucose consumption and glycogen synthesis via upregulating insulin receptor substrate 1 (IRS1)/PI3K/Akt/glucose transporters (GLUTs) signaling pathways to alleviate IR. Rutin, resveratrol, and catechin show lower binding energy docking with IRS1, PI3K, Akt, and GLUT4 proteins, indicating better interactions. It might be an effective constituent against IR. Hence, BQP could become a potential functional food source for blood glucose management among insulin-resistant people.

The IGF-Independent Role of IRS-2 in the Secretion of MMP-9 Enhances the Growth of Prostate Carcinoma Cell Line PC3.

Insulin receptor substrate-2 (IRS-2), a substrate of the insulin-like growth factor (IGF)-I receptor, is highly expressed in the prostate cancer cell line, PC3. We recently demonstrated that extracellular signal-regulated kinase (Erk1/2), a kinase downstream of IGF signaling, is activated in PC3 cells under serum starvation, and this activation can be inhibited by IRS-2 knockdown. Here, we observed that adding an IGF-I-neutralizing antibody to the culture medium inhibited the activation of Erk1/2. Suppression of Erk1/2 in IRS-2 knockdown cells was restored by the addition of a PC3 serum-free conditioned medium. In contrast, the IRS-2-silenced PC3 conditioned medium could not restore Erk1/2 activation, suggesting that IRS-2 promotes the secretion of proteins that activate the IGF signaling pathway. Furthermore, gelatin zymography analysis of the conditioned medium showed that matrix metalloproteinase-9 (MMP-9) was secreted extracellularly in an IRS-2 dependent manner when PC3 was cultured under serum starvation conditions. Moreover, MMP-9 knockdown suppressed Erk1/2 activation, DNA synthesis, and migratory activity. The IRS-2 levels were positively correlated with Gleason grade in human prostate cancer tissues. These data suggest that highly expressed IRS-2 activates IGF signaling by enabling the secretion of MMP-9, which is associated with hyperproliferation and malignancy of prostate cancer cell line, PC3.

Influence of polymorphisms in IRS1, IRS2, MC3R, and MC4R on metabolic and inflammatory status and food intake in Brazilian adults: An exploratory pilot study.

Polymorphisms in genes of leptin-melanocortin and insulin pathways have been associated with obesity and type 2 diabetes. We hypothesized that polymorphisms in IRS1, IRS2, MC3R, and MC4R influence metabolic and inflammatory markers and food intake composition in Brazilian subjects. This exploratory pilot study included 358 adult subjects. Clinical, anthropometric, and laboratory data were obtained through interview and access to medical records. The variants IRS1 rs2943634 A˃C, IRS2 rs1865434 C>T, MC3R rs3746619 C>A, and MC4R rs17782313 T>C were analyzed by real-time polymerase chain reaction. Food intake composition was assessed in a group of subjects with obesity (n = 84) before and after a short-term nutritional counseling program (9 weeks). MC4R rs17782313 was associated with increased risk of obesity (P = .034). Multivariate linear regression analysis adjusted by covariates indicated associations of IRS2 rs1865434 with reduced low-density lipoprotein cholesterol and resistin, MC3R rs3746619 with high glycated hemoglobin, and IRS1 rs2943634 and MC4R rs17782313 with increased high-sensitivity C-reactive protein (P < .05). Energy intake and carbohydrate and total fat intakes were reduced after the diet-oriented program (P < .05). Multivariate linear regression analysis showed associations of IRS2 rs1865434 with high basal fiber intake, IRS1 rs2943634 with low postprogram carbohydrate intake, and MC4R rs17782313 with low postprogram total fat and saturated fatty acid intakes (P < .05). Although significant associations did not survive correction for multiple comparisons using the Benjamini-Hochberg method in this exploratory study, polymorphisms in IRS1, IRS2, MC3R, and MC4R influence metabolic and inflammatory status in Brazilian adults. IRS1 and MC4R variants may influence carbohydrate, total fat, and saturated fatty acid intakes in response to a diet-oriented program in subjects with obesity.

Genetic Reduction of Insulin Signaling Mitigates Amyloid-ß Deposition by Promoting Expression of Extracellular Matrix Proteins in the Brain.

The insulin/IGF-1 signaling (IIS) regulates a wide range of biological processes, including aging and lifespan, and has also been implicated in the pathogenesis of Alzheimer's disease (AD). We and others have reported that reduced signaling by genetic ablation of the molecules involved in IIS (e.g., insulin receptor substrate 2 [IRS-2]) markedly mitigates amyloid plaque formation in the brains of mouse models of AD, although the molecular underpinnings of the amelioration remain unsolved. Here, we revealed, by a transcriptomic analysis of the male murine cerebral cortices, that the expression of genes encoding extracellular matrix (ECM) was significantly upregulated by the loss of IRS-2. Insulin signaling activity negatively regulated the phosphorylation of Smad2 and Smad3 in the brain, and suppressed TGF-ß/Smad-dependent expression of a subset of ECM genes in brain-derived cells. The ECM proteins inhibited Aß fibril formation in vitro, and IRS-2 deficiency suppressed the aggregation process of Aß in the brains of male APP transgenic mice as revealed by injection of aggregation seeds in vivo Our results propose a novel mechanism in AD pathophysiology whereby IIS modifies Aß aggregation and amyloid pathology by altering the expression of ECM genes in the brain.SIGNIFICANCE STATEMENT The insulin/IGF-1 signaling (IIS) has been recognized as a regulator of aging, a leading risk factor for the onset of Alzheimer's disease (AD). In AD mouse models, genetic deletion of key IIS molecules markedly reduces the amyloid plaque formation in the brain, although the molecular underpinnings of this amelioration remain elusive. We found that the deficiency of insulin receptor substrate 2 leads to an increase in the expression of various extracellular matrices (ECMs) in the brain, potentially through TGF-ß/Smad signaling. Furthermore, some of those ECMs exhibited the potential to inhibit amyloid plaque accumulation by disrupting the formation of Aß fibrils. This study presents a novel mechanism by which IIS regulates Aß accumulation, which may involve altered brain ECM expression.

BRD7 improves glucose homeostasis independent of IRS proteins.

Bromodomain-containing protein 7 (BRD7) has emerged as a player in the regulation of glucose homeostasis. Hepatic BRD7 levels are decreased in obese mice, and the reinstatement of hepatic BRD7 in obese mice has been shown to establish euglycemia and improve glucose homeostasis. Of note, the upregulation of hepatic BRD7 levels activates the AKT cascade in response to insulin without enhancing the sensitivity of the insulin receptor (InsR)-insulin receptor substrate (IRS) axis. In this report, we provide evidence for the existence of an alternative insulin signaling pathway that operates independently of IRS proteins and demonstrate the involvement of BRD7 in this pathway. To investigate the involvement of BRD7 as a downstream component of InsR, we utilized liver-specific InsR knockout mice. Additionally, we employed liver-specific IRS1/2 knockout mice to examine the requirement of IRS1/2 for the action of BRD7. Our investigation of glucose metabolism parameters and insulin signaling unveiled the significance of InsR activation in mediating BRD7's effect on glucose homeostasis in the liver. Moreover, we identified an interaction between BRD7 and InsR. Notably, our findings indicate that IRS1/2 is not necessary for BRD7's regulation of glucose metabolism, particularly in the context of obesity. The upregulation of hepatic BRD7 significantly reduces blood glucose levels and restores glucose homeostasis in high-fat diet-challenged liver-specific IRS1/2 knockout mice. These findings highlight the presence of an alternative insulin signaling pathway that operates independently of IRS1/2 and offer novel insights into the mechanisms of a previously unknown insulin signaling in obesity.

MiR-222-3p suppresses C2C12 myoblast proliferation and differentiation via the inhibition of IRS-1/PI3K/Akt pathway.

Numerous studies have revealed the profound impact of microRNAs on regulating skeletal muscle development and regeneration. However, the biological function and regulation mechanism of miR-222-3p in skeletal muscle remains largely unknown. In this study, miR-222-3p was found to be abundantly expressed in the impaired skeletal muscles, indicating that it might have function in the development and regeneration process of the skeletal muscle. MiR-222-3p overexpression impeded C2C12 myoblast proliferation and myogenic differentiation, whereas inhibition of miR-222-3p got the opposite results. The dual-luciferase reporter assay showed that insulin receptor substrate-1 (IRS-1) was the target gene of miR-222-3p. We next found that knockdown of IRS-1 could obviously suppress C2C12 myoblast proliferation and differentiation. Additionally, miR-222-3p-induced repression of myoblast proliferation and differentiation was verified to be associated with a decrease in phosphoinositide 3-kinase (PI3K)-Akt signaling. Overall, we demonstrated that miR-222-3p inhibited C2C12 cells myogenesis via IRS-1/PI3K/Akt pathway. Therefore, miR-222-3p may be used as a therapeutic target for alleviating muscle loss caused by inherited and nonhereditary diseases.

Association of insulin receptor substrate 1 (IRS-1) gene polymorphism with growth and litter-related traits in NMER rabbits.

This study investigated the associations between the c.189G > T polymorphism of the insulin receptor substrate-1 (IRS-1) gene and the growth and litter size-related traits in the Native rabbit in Middle Egypt (NMER). One hundred sixty-two NMER rabbits were genotyped by RFLP-PCR using Sau3AI restriction enzyme and the associations of the reported genotypes with body weights at 5th, 6th, 8th, 10th, and 12th week old, body gain, and daily gain plus, the litter size-related traits were determined. Additionally, the genotypic and allelic frequencies, the effective (Ne) and observed (NA) numbers of alleles, observed (Ho) and expected (He) heterozygosity, Hardy-Weinberg equilibrium (HWE), and the decrease in heterozygosity because of inbreeding (FIS) were calculated. Three genotypes; GG, GT, and TT with 0.65, 0.33, and 0.02 frequencies, respectively which fit HWE were reported. These genotypes displayed a marked low FIS value. Significant associations of the genotypes with the body weights, and gains, except at the 5th week old determined with superiority of the GT genotype compared with the other genotypes. All reported litter size-related traits significantly varied among different genotypes. In summary, the c.189G > T SNP of the IRS-1 gene is an effective genetic marker to improve growth performance and litter size traits of the NMER rabbits.

[Effect of miR-96-5p targeting IRS1 on apoptosis of PC12 cells induced by aluminum maltol].

Objective: To investigate the effect and mechanism of miR-96-5p on apoptosis of PC12 cells induced by maltol aluminum. Methods: In January 2021, PC12 cells at logarithmic growth phase were divided into blank control group and low, medium and high dose group. Cells in each group were treated with 0, 100, 200 and 400 µmol/L maltol aluminum for 24 hours respectively. Cells were collected and cell apoptosis rates were detected by flow cytometry, miR-96-5p and insulin receptor substrate 1 (IRS1) mRNA expressions were detected by qRT-PCR, and the protein expression levels of cysteine protease 3 (Caspase3) 、activated cysteine protease 3 (Cleaved-caspase3) 、IRS1、phosphorylated protein kinase B (p-AKT) and phosphorylated glucose synthesis kinase 3ß (p-GSK3ß) were detected by western blotting. The target binding relationship between miR-96-5p and IRS1 was detected by double luciferase reporter gene experiment. The miR-96-5p inhibitor cells and negative control cells were constructed after transfecting PC12 cells with miR-96-5p inhibitor for 24 hours. The cells were divided into blank control group, negative control group, aluminum exposure group, aluminum exposure+negative control group, aluminum exposure+miR-96-5p inhibition group, and miR-96-5p inhibition group. After transfecting PC12 cells with miR-96-5p inhibition and IRS1 siRNA for 24 h, the cells were divided into aluminum exposure+miR-96-5p inhibition+negative control group and aluminum exposure+miR-96-5p inhibition+IRS1 inhibition group. The control group was cultured in complete culture medium, and cells in the aluminum exposure group were treated with 200 µmol/L maltol aluminum for 24 hours. Cells in each group were collected and the apoptosis rate, miR-96-5p and IRS1 mRNA expression levels, as well as protein expression levels of Caspase3, Cleaved-caspase3, IRS1, p-AKT, and p-GSK3ß were measured. Results: After 24 hours of exposure, compared with blank control group and low-dose group, the apoptosis rates, relative expressions of Caspase3 and Cleaved-caspase3 proteins, and relative expressions of miR-96-5p in the medium and high-dose groups of PC12 cells were significantly increased, while the relative expression levels of IRS1 mRNA, IRS1, p-AKT and p-GSK3ß proteins were significantly decreased (P<0.05). Targetscan prediction and double luciferase report experiment both proved that IRS1 was a direct target gene of miR-96-5p. In the transfection experiment, compared with the aluminum exposure group, the apoptosis rate, the relative expressions of Caspase3 and Cleaved-caspase3 proteins, the relative expression of miR-96-5p in the aluminum exposure+miR-96-5p inhibition group were significantly decreased, while the relative expression levels of IRS1 mRNA and IRS1, p-AKT and p-GSK3ß proteins were significantly increased (P<0.05). In the IRS1 low expression experiment, compared with the aluminum exposure+miR-96-5p inhibition+negative control group, the apoptosis rate, the relative expressions of Caspase3 and Cleaved-caspase3 proteins in the aluminum exposure+miR-96-5p inhibition+IRS1 inhibition group were significantly increased, while the relative expression levels of IRS1 mRNA and IRS1, p-AKT and p-GSK3ß proteins were significantly decreased (P<0.05) . Conclusion: The increased expression of miR-96-5p and the targeted inhibition of IRS1 may be one of the mechanisms of apoptosis of PC12 cells induced by maltol aluminum exposure.

High risk genetic variants of human insulin receptor substrate 1(IRS1) infer structural instability and functional interference.

Insulin receptor substrate 1(IRS1) is a signaling adapter protein encoded by the IRS1 gene. This protein delivers signals from insulin and insulin-like growth factor-1(IGF-1) receptors to the phosphatidylinositol 3-kinases (P13K)/protein kinase B (Akt) and Extracellular signal-regulated kinases (Erk) - Mitogen-activated protein (MAP) kinase pathways, which regulate particular cellular processes. Mutations in this gene have been linked to type 2 diabetes mellitus, a heightened risk of insulin resistance, and an increased likelihood of developing a number of different malignancies. The structure and function of IRS1 could be severely compromised as a result of single nucleotide polymorphism (SNP) type genetic variants. In this study, we focused on identification of the most harmful non-synonymous SNPs (nsSNPs) of the IRS1 gene as well as prediction of their structural and functional consequences. Six different algorithms made the initial prediction that 59 of the 1142 IRS1 nsSNPs would have a negative impact on the protein structure. In-depth evaluations detected 26 nsSNPs located inside the functional domains of IRS1. Following that, 16 nsSNPs were identified as more harmful based on conservation profile, hydrophobic interaction, surface accessibility, homology modelling, and inter-atomic interactions. Following an in-depth analysis of protein stability, M249T (rs373826433), I223T (rs1939785175) and V204G (rs1574667052) were identified as three most deleterious SNPs and were subjected to molecular dynamics simulation for further insights. These findings will help us understand the implications for disease susceptibility, cancer progression, and the efficacy of therapeutic development against IRS1 gene mutants.Communicated by Ramaswamy H. Sarma.

Enhancer RNA IRS2e is essential for IRS2 expression and the oncogenic properties in oral squamous cell carcinoma.

The crucial roles of enhancer RNAs (eRNAs) in the regulation of gene expression in human diseases have drawn wider and wider attention in recent years. However, the specific expression profile and function of eRNAs are still rarely discussed in oral squamous cell carcinoma (OSCC), the most common subtype of head and neck squamous cell carcinoma (HNSC). In this study, we aimed to investigate the expression and function of an uncharacterized eRNA, insulin receptor substrate 2 enhancer RNA (IRS2e), in OSCC. We found that IRS2e was overexpressed in HNSC and its overexpression was positively correlated with a poor prognosis. The downregulation of IRS2e by short hairpin RNA significantly inhibited cell growth and induced cellular apoptosis and cell-cycle arrest in OSCC cells. Furthermore, the ablation of IRS2e inhibited tumor growth in vivo. Mechanically, IRS2e is essential for the expression of insulin receptor substrate 2 (IRS2), an oncogene nearby IRS2e in chromosome 13. Altogether, our study demonstrated that IRS2e is a novel oncogenic eRNA required for oncogene IRS2 expression in OSCC.

Insulin receptor substrate-1 gene polymorphism and lipid panel data in type 2 diabetic patients with comorbid obesity and/or essential hypertension.

Objective. The hallmarks of type 2 diabetes mellitus (T2DM) are insulin resistance (IR) and insulin receptor substrate (IRS) proteins essential for the insulin signaling. IRS-1 gene has not only been shown to be associated with T2DM, but also has indicated that it may significantly correlate with diabetic complications, such as coronary heart disease and obesity. The aim of this study was to evaluate changes of the lipid panel data in T2DM patients with comorbid obesity and/or essential hypertension in connection with the IRS-1 (rs2943640) polymorphism. Methods. The study involved 33 T2DM patients and 10 healthy individuals. The IRS-1 (rs2943640) polymorphism was genotyped using a TaqMan real-time polymerase chain reaction method. Blood serum lipid panel data were determined with commercially available kits using a Cobas 6000 analyzer. Results. Analysis of the serum lipid panel data depending on the presence of the C/A alleles of IRS-1 (rs2943640) polymorphism in T2DM patients, regardless of the presence/absence of comorbidities, showed significantly lower level of high-density lipoprotein cholesterol (HDL-C) and significantly higher level of non-HDL-C in the carriers of C allele vs. carriers of A allele. In T2DM patients with comorbid obesity and essential hypertension, proatherogenic lipid changes were found in both C and A alleles carriers. Analysis of the effect of IRS-1 (rs2943640) genotypes on serum lipid panel data in T2DM patients, regardless of the presence/absence of comorbidities, showed that the CC genotype carriers had more pronounced pro-atherogenic changes vs. carriers of СА and АА genotypes. In the comorbid course of T2DM (both in combination with obesity and obesity and essential hypertension), pro-atherogenic changes were found in the carriers of the CA genotype of IRS-1 (rs2943640) polymorphism. Conclusions. The presence of the C allele of IRS-1 (rs2943640) polymorphism in both homo-zygous and heterozygous states indicates increased risk of pro-atherogenic changes in T2DM patients with comorbid obesity and/or essential hypertension.

Repressing IRS1/2 by NT157 inhibits the malignant behaviors of ovarian cancer through inactivating PI3K/AKT/mTOR pathway and inducing autophagy.

Insulin receptor substrate 1 and 2 (IRS1/2) have been found involved in many cancers development and their inhibitors exert significant tumor-suppressive effects. Here, we tried to explore the function of NT157, an IGF1R-IRS1/2 inhibitor, in ovarian cancer. We treated ovarian cancer cells with varying doses of NT157. The MTT assay was employed to evaluate cell proliferation and colony formation assay was used for detecting colony-forming ability. TUNEL assay was adopted to test cell apoptosis. Cell invasion was checked by the Transwell assay. The expression of apoptosis-related proteins, autophagy markers, IRS1/2, and PI3K/AKT/mTOR pathway was compared by Western blot, immunofluorescence, or qRT-PCR. As indicated by the data, NT157 abated the viability, proliferation, and induced autophagy of ovarian cancer cells. Overexpressing IRS1/2 attenuated the tumor-suppressive effect of NT157 and heightened the PI3K/AKT/mTOR pathway activation. Inhibition of the PI3K/AKT/mTOR pathway enhanced the tumor-suppressive effect of NT157 and facilitated NT157-mediated autophagy. However, the autophagy inhibitor 3-MA partly reversed NT-157-mediated antitumor effects. In conclusion, this study disclosed that NT157 suppressed the malignant phenotypes of ovarian cancer cells by inducing autophagy and hampering the expression of IRS1/2 and PI3K/AKT/mTOR pathway.

Overexpression of Insulin Receptor Substrate 1 (IRS1) Relates to Poor Prognosis and Promotes Proliferation, Stemness, Migration, and Oxidative Stress Resistance in Cholangiocarcinoma.

Cholangiocarcinoma (CCA) is one of the oxidative stress-driven carcinogenesis through chronic inflammation. Insulin receptor substrate 1 (IRS1), an adaptor protein of insulin signaling pathways, is associated with the progression of many inflammation-related cancers. This study hypothesized that oxidative stress regulates IRS1 expression and that up-regulation of IRS1 induces CCA progression. The localizations of IRS1 and an oxidative stress marker (8-oxodG) were detected in CCA tissues using immunohistochemistry (IHC). The presence of IRS1 in CCA tissues was confirmed using immortal cholangiocyte cells (MMNK1), a long-term oxidative-stress-induced cell line (ox-MMNK1-L), and five CCA cell lines as cell culture models. IRS1 was overexpressed in tumor cells and this was associated with a shorter patient survival time and an increase in 8-oxodG. IRS1 expression was higher in ox-MMNK1-L cells than in MMNK1 cells. Knockdown of IRS1 by siRNA in two CCA cell lines led to inhibition of proliferation, cell cycle progression, migration, invasion, stemness, and oxidative stress resistance properties. Moreover, a transcriptomics study demonstrated that suppressing IRS1 in the KKU-213B CCA cell line reduced the expression levels of several genes and pathways involved in the cellular functions. The findings indicate that IRS1 is a key molecule in the connection between oxidative stress and CCA progression. Therefore, IRS1 and its related genes can be used as prognostic markers and therapeutic targets for CCA therapy.