Prognostic value and potential regulatory relationship of miR-200c-5p in colorectal cancer.

This study aimed to investigate the relationship and potential mechanisms of miR-200c-5p in colorectal cancer (CRC) progression. Differentially expressed miRNAs were screened using the TCGA database. Subsequently, univariate analysis was performed to identify CRC survival-related miRNAs. Survival and receiver operator characteristic curves were generated. The target genes of miR-200c-5p and the relevant signaling pathways or biological processes were predicted by the miRNet database and enrichment analyses. The miR-200c-5p expression was detected using quantitative reverse-transcription polymerase chain reaction, Cell Counting Kit-8, Transwell, and cell apoptosis experiments were performed to determine miR-200c-5p's impact on CRC cell viability, invasiveness, and apoptosis. Finally, we constructed a CRC mouse model with inhibited miR-200c-5p to evaluate its impact on tumors. miR-200c-5p was upregulated in CRC, implying a favorable prognosis. Gene set enrichment analysis revealed that miR-200c-5p may participate in signaling pathways such as the TGF-ß signaling pathway, RIG-I-like receptor signaling pathway, renin-angiotensin system, and DNA replication. miR-200c-5p potentially targeted mRNAs, including KCNE4 and CYP1B1, exhibiting a negative correlation with their expression. Furthermore, these mRNAs may participate in biological processes like the regulation of intracellular transport, cAMP-dependent protein kinase regulatory activity, ubiquitin protein ligase binding, MHC class II protein complex binding, and regulation of apoptotic signaling pathway. Lastly, miR-200c-5p overexpression repressed the viability and invasiveness of CRC cells but promoted apoptosis. The tumor size, weight, and volume were significantly increased by inhibiting miR-200c-5p (p < 0.05). miR-200c-5p is upregulated in CRC, serving as a promising biomarker for predicting CRC prognosis.

Role of oncogenic long noncoding RNA KCNQ1OT1 in colon cancer.

The role of lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1) in colon cancer involves various tumorigenic processes and has been studied widely. However, the mechanism by which it promotes colon cancer remains unclear. Retroviral vector pSEB61 was retrofitted in established HCT116-siKCN and SW480-siKCN cells to silence KCNQ1OT1. Cellular proliferation was measured using CCK8 assay, and flow cytometry (FCM) detected cell cycle changes. RNA sequencing (RNA-Seq) analysis showed differentially expressed genes (DEGs). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were carried out to analyze enriched functions and signaling pathways. RT-qPCR, immunofluorescence, and western blotting were carried out to validate downstream gene expressions. The effects of tumorigenesis were evaluated in BALB/c nude mice by tumor xenografts. Our data revealed that the silencing of KCNQ1OT1 in HCT116 and SW480 cells slowed cell growth and decreased the number of cells in the G2/M phase. RNA-Seq analysis showed the data of DEGs enriched in various GO and KEGG pathways such as DNA replication and cell cycle. RT-qPCR, immunofluorescence, and western blotting confirmed downstream CCNE2 and PCNA gene expressions. HCT116-siKCN cells significantly suppressed tumorigenesis in BALB/c nude mice. Our study suggests that lncRNA KCNQ1OT1 may provide a promising therapeutic strategy for colon cancer.

The genetic landscape of inherited retinal dystrophies in Arabs.

Inherited retinal dystrophies (IRDs) are a major cause of vision loss. Altogether are highly heterogeneous genotypically and phenotypically, exhibiting substantial differences worldwide. To shed more light on these conditions, we investigated the genetic and phenotypic landscape of IRDs in the Arabs globally and per country.We analyzed 1,621 affected individuals from 16 Arabic countries reported in 198 articles. At the phenotypic level, rod-cone dystrophy (RCD) and Usher syndrome were the most prevalent conditions among non-syndromic and syndromic IRDs. At the gene level, TULP1, ABCA4, RP1, CRB1, MYO7A, RPE65, KCNV2, and IMPG2 were the most mutated genes. Interestingly, all except CRB1 were highly prevalent because they harbored founder mutations, implying that consanguinity is a major determinant in Arab countries. Of note, ~ 93% of the investigated individuals carried homozygous mutations. The country analysis for the IRDs conditions and their associated genotypes revealed that whereas Leber Congenital Amaurosis, RCD, and USHER syndrome were widely distributed, bestrophinopathies and non-syndromic hearing loss were restricted to specific countries (till now).This study could be a starting point for initiating suitable health policies towards IRDs in the Arab world. The high degree of homozygosity urges the need for genetic counsellors to provide personalized information and support the affected individuals.

The in vitro anticancer effects of FS48 from salivary glands of Xenopsylla cheopis on NCI-H460 cells via its blockage of voltage-gated K+ channels.

Voltage-gated K+ (Kv) channels play a role in the cellular processes of various cancer cells, including lung cancer cells. We previously identified and reported a salivary protein from the Xenopsylla cheopis, FS48, which exhibited inhibitory activity against Kv1.1-1.3 channels when assayed in HEK 293T cells. However, whether FS48 has an inhibitory effect on cancer cells expressing Kv channels is unclear. The present study aims to reveal the effects of FS48 on the Kv channels and the NCI-H460 human lung cancer cells through patch clamp, MTT, wound healing, transwell, gelatinase zymography, qRT-PCR and WB assays. The results demonstrated that FS48 can be effective in suppressing the Kv currents, migration, and invasion of NCI-H460 cells in a dose-dependent manner, despite the failure to inhibit the proliferation. Moreover, the expression of Kv1.1 and Kv1.3 mRNA and protein were found to be significantly reduced. Finally, FS48 decreases the mRNA level of MMP-9 while increasing TIMP-1 mRNA level. The present study highlights for the first time that blood-sucking arthropod saliva-derived protein can inhibit the physiological activities of tumour cells via the Kv channels. Furthermore, FS48 can be taken as a hit compound against the tumour cells expressing Kv channels.

Curcumin, a dietary natural supplement, prolongs the action potential duration of KCNE1-D85N-induced pluripotent stem cell-derived cardiomyocytes.

BACKGROUND: Curcumin, a polyphenolic dietary natural compound and active ingredient in turmeric, exerts antioxidant, anti-inflammatory, antidiabetic, anticancer, and antiarrhythmic properties. KCNE1-D85N, present in ∼1% of white, is a common, potentially proarrhythmic variant that predisposes individuals to drug-induced QT prolongation under certain conditions. OBJECTIVE: The purpose of this article was to test the hypothesis that curcumin might cause action potential duration (APD) prolongation in KCNE1-D85N-derived human-induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). METHODS: Gene-edited/variant-corrected isogenic control and patient-specific KCNE1-D85N-containing iPSC-CMs were generated previously. Voltage-sensing dye, multielectrode array (MEA), and whole-cell patch clamp technique were used to measure APD without and with 4-hour incubation with 10 nM curcumin. RESULTS: KCNE1-D85N-derived iPSC-CMs demonstrated significant APD prolongation with treatment of 10 nM curcumin. Using voltage-sensing dye, action potential duration at 90% repolarization (APD90) was 578 ± 7 ms (n = 39) at baseline and was prolonged to 658 ± 13 ms (n = 35) with curcumin incubation (P < .0001). Using MEA, APD90 at baseline was 237 ± 6 ms (n = 24) compared with 280 ± 6 ms (n = 12) with curcumin incubation (P = .0002). The whole-cell patch clamp technique confirmed these results, with APD90 being 544 ± 37 ms at baseline and 664 ± 40 ms with treatment of curcumin (P < .005). However, APD from isogenic control iPSC-CMs remained unchanged with curcumin treatment. CONCLUSION: This study provides pharmacological and functional evidence to suggest that curcumin, a dietary natural supplement, might cause APD prolongation in patients with common, potentially proarrhythmic functional variants such as KCNE1-D85N. Whether this supplement is potentially dangerous for the Caucasian subpopulation that has this variant warrants further investigation.

KCNE4 expression is correlated with the pathological characteristics of colorectal cancer patients and associated with the radioresistance of cancer cells.

BACKGROUND: Colorectal cancer (CRC) is a common malignancy, and radioresistance limits the effectiveness of radiotherapy for rectal cancer. This study is performed to investigate the role and regulatory mechanism of Potassium Voltage-Gated Channel Subfamily E Regulatory Subunit 4 (KCNE4) in the radioresistance of CRC cells. METHODS: Immunohistochemical staining results of KCNE4 in normal tissues and CRC tissues were obtained from the Human Protein Atlas (HPA) database. The UALCAN database was used for analyzing KCNE4 mRNA expression in normal tissue samples and CRC tissue samples and its relationship with tumor stage. The relationship of KCNE4 expression with prognosis was analyzed utilizing the data of GEPIA database. LinkedOmics database was searched to analyze the co-expressed gene sets of KCNE4 in CRC, and to analyze the signaling pathways related with KCNE4 in CRC. GO and KEGG enrichment analyses were carried out on the co-expressed genes of KCNE4 with DAVID database. Ionizing radiation (IR)-resistant cell lines (HCT116/IR and HT29/IR) were established; cell viability was assessed via cell counting kit-8 (CCK-8) and EdU assays, and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay was performed for detecting cell apoptosis. Western blotting was carried out to detect the expressions of p-p85 and p-AKT. RESULTS: KCNE4 was highly expressed in CRC tissues and linked to advanced tumor stage, lymph node metastasis and poor prognosis of CRC patients. KCNE4 overexpression promoted HCT116/IR cell proliferation and inhibited the apoptosis, while KCNE4 knockdown suppressed HT29/IR cell proliferation and facilitated the apoptosis. Furthermore, high KCNE4 expression was associated with the activation of the PI3K/AKT signal pathway. CONCLUSION: KCNE4 is associated with the clinicopathological characteristics of CRC patients, and its high expression level contributes to the radioresistance of cancer cells via activating the PI3K/AKT signal pathway.

[Association study between the KCNE family gene polymorphisms of potassium channel gene and the susceptibility of atrial fibrillation].

Objective: To investigate the relationship between KCNE family gene polymorphisms of potassium channel gene and the susceptibility of atrial fibrillation (AF). Methods: In the case-control study, a total of 648 subjects were studied, of which 338 patients with atrial fibrillation were selected from the Department of Cardiovascular Medicine, Putuo Hospital Affiliated to Shanghai University of Traditional Chinese Medicine from January 2019 to December 2019, and 310 healthy people were selected from the physical examination population during the same period. DNA sequencing technology and polymerase chain reaction (PCR) were used to detect the genotype and allele frequency of rs1805127 of KCNE1, rs9984281 of KCNE2, rs9516, rs626930 of KCNE3 and rs12621643 of KCNE4. Results: The ages of subjects in atrial fibrillation group and control group were (69±13) and (73±8) years, respectively (P=0.077). Men subjects accounted for 57.70% (195 men) and 40.00% (124 men) in the two groups, respectively (P=0.092). The distribution frequencies of the allele C at rs1805127 of gene KCNE1, the allele A at rs9984281 of gene KCNE2 and the allele G at rs12621643 of gene KCNE4 were significantly different between groups (P<0.05). After adjustment for sex, smoking, hypertension, cardiac insufficiency and other factors, it was found that the increase in the frequency of the above three loci would increase the risk of atrial fibrillation (rs1805127 OR=7.064, 95%CI:1.559-31.997; rs9984281 OR=4.210, 95%CI:1.118-15.850; rs12621643 OR=2.679, 95%CI:1.025-6.998). Conclusion: The rs1805127 of KCNE1, the rs9984281 of KCNE2,the rs12621643 of KCNE4 were significantly associated with the susceptibility to atrial fibrillation.

LncRNA KCNQ1OT1 promotes the metastasis of ovarian cancer by increasing the methylation of EIF2B5 promoter.

BACKGROUND: Long non-coding RNAs (lncRNAs) have emerged as regulators of human malignancies, including ovarian cancer (OC). LncRNA KCNQ1OT1 could promote OC progression, and EIF2B5 was associated with development of several tumors. This project was aimed to explore the role of lncRNA KCNQ1OT1 in OC development, as well as the involving action mechanism. METHODS: Reverse transcription quantitative polymerase chain reaction (RT-qPCR) or Western blotting was employed to determine the expression levels of KCNQ1OT1 and EIF2B5. OC cell proliferation was evaluated by MTT and colony formation assays, and wound healing and Transwell assays were implemented to monitor cell migration and invasion, respectively. The methylation status of EIF2B5 promoter was examined by MS-PCR, to clarify whether the expression of EIF2B5 was decreased. The binding activity of KCNQ1OT1 to methyltransferases DNMT1, DNMT3A and DNMT3B was determined by dual luciferase reporter assay or RIP assay, to explore the potential of KCNQ1OT1 alters the expression of its downstream gene. ChIP assay was carried out to verify the combination between EIF2B5 promoter and above three methyltransferases. RESULTS: Expression of lncRNA KCNQ1OT1 was increased in OC tissues and cells. EIF2B5 expression was downregulated in OC, which was inversely correlated with KCNQ1OT1. Knockdown of KCNQ1OT1 inhibited OC cell proliferation and metastasis. KCNQ1OT1 could downregulate EIF2B5 expression by recruiting DNA methyltransferases into EIF2B5 promoter. Furthermore, interference of EIF2B5 expression rescued KCNQ1OT1 depletion-induced inhibitory impact on OC cell proliferation and metastasis. CONCLUSION: Our findings evidenced that lncRNA KCNQ1OT1 aggravated ovarian cancer metastasis by decreasing EIF2B5 expression level, and provided a novel therapeutic strategy for OC.

Clinically Relevant KCNQ1 Variants Causing KCNQ1-KCNE2 Gain-of-Function Affect the Ca2+ Sensitivity of the Channel.

Dominant KCNQ1 variants are well-known for underlying cardiac arrhythmia syndromes. The two heterozygous KCNQ1 missense variants, R116L and P369L, cause an allelic disorder characterized by pituitary hormone deficiency and maternally inherited gingival fibromatosis. Increased K+ conductance upon co-expression of KCNQ1 mutant channels with the beta subunit KCNE2 is suggested to underlie the phenotype; however, the reason for KCNQ1-KCNE2 (Q1E2) channel gain-of-function is unknown. We aimed to discover the genetic defect in a single individual and three family members with gingival overgrowth and identified the KCNQ1 variants P369L and V185M, respectively. Patch-clamp experiments demonstrated increased constitutive K+ conductance of V185M-Q1E2 channels, confirming the pathogenicity of the novel variant. To gain insight into the pathomechanism, we examined all three disease-causing KCNQ1 mutants. Manipulation of the intracellular Ca2+ concentration prior to and during whole-cell recordings identified an impaired Ca2+ sensitivity of the mutant KCNQ1 channels. With low Ca2+, wild-type KCNQ1 currents were efficiently reduced and exhibited a pre-pulse-dependent cross-over of current traces and a high-voltage-activated component. These features were absent in mutant KCNQ1 channels and in wild-type channels co-expressed with calmodulin and exposed to high intracellular Ca2+. Moreover, co-expression of calmodulin with wild-type Q1E2 channels and loading the cells with high Ca2+ drastically increased Q1E2 current amplitudes, suggesting that KCNE2 normally limits the resting Q1E2 conductance by an increased demand for calcified calmodulin to achieve effective channel opening. Our data link impaired Ca2+ sensitivity of the KCNQ1 mutants R116L, V185M and P369L to Q1E2 gain-of-function that is associated with a particular KCNQ1 channelopathy.

Artificial pore blocker acts specifically on voltage-gated potassium channel isoform KV1.6.

Among voltage-gated potassium channel (KV) isoforms, KV1.6 is one of the most widespread in the nervous system. However, there are little data concerning its physiological significance, in part due to the scarcity of specific ligands. The known high-affinity ligands of KV1.6 lack selectivity, and conversely, its selective ligands show low affinity. Here, we present a designer peptide with both high affinity and selectivity to KV1.6. Previously, we have demonstrated that KV isoform-selective peptides can be constructed based on the simplistic α-hairpinin scaffold, and we obtained a number of artificial Tk-hefu peptides showing selective blockage of KV1.3 in the submicromolar range. We have now proposed amino acid substitutions to enhance their activity. As a result, we have been able to produce Tk-hefu-11 that shows an EC50 of ≈70 nM against KV1.3. Quite surprisingly, Tk-hefu-11 turns out to block KV1.6 with even higher potency, presenting an EC50 of ≈10 nM. Furthermore, we have solved the peptide structure and used molecular dynamics to investigate the determinants of selective interactions between artificial α-hairpinins and KV channels to explain the dramatic increase in KV1.6 affinity. Since KV1.3 is not highly expressed in the nervous system, we hope that Tk-hefu-11 will be useful in studies of KV1.6 and its functions.

Comprehensive analysis of KCTD family genes associated with hypoxic microenvironment and immune infiltration in lung adenocarcinoma.

To obtain novel insights into the tumor biology and therapeutic targets of LUAD, we performed a comprehensive analysis of the KCTD family genes. The expression patterns and clinical significance of the KCTD family were identified through multiple bioinformatics mining. Moreover, the molecular functions and potential mechanisms of differentially expressed KCTDs were evaluated using TIMER 2.0, cBioPortal, GeneMANIA, LinkedOmics and GSEA. The results indicated that the mRNA and protein expression levels of KCTD9, KCTD10, KCTD12, KCTD15 and KCTD16 were significantly decreased in LUAD, while those of KCTD5 were significantly increased. High KCTD5 expression was significantly associated with advanced tumor stage, lymph node metastasis, TP53 mutation and poor prognosis. In addition, KCTD5 was positively correlated with CD8 + T cell, neutrophil, macrophage and dendritic cell infiltration. Additionally, KCTDs demonstrate promising prospects in the diagnosis of LUAD. Importantly, high KCTD5 expression was enriched in signaling pathways associated with the malignant progression of tumors, including the inflammatory response, the IL6/JAK/STAT3 signaling pathway, EMT and hypoxia. Further association analysis showed that KCTD5 was positively correlated with hypoxia-related genes such as HIF1. Overall, KCTDs can be used as molecular targets for the treatment of LUAD, as well as effective molecular biomarkers for diagnosis and prognosis prediction.

Progesterone Changes the Pregnancy-Induced Adaptation of Cardiomyocyte Kv2.1 Channels via MicroRNA-29b.

The cardiovascular system adaptation occurs during pregnancy to ensure adequate maternal circulation. Progesterone (P4) is widely used in hormone therapy to support pregnancy, but little is known about its effects on maternal cardiac function. In this study, we investigated the cardiac repolarization and ion channel expression in pregnant subjects and mice models and studied the effects of P4 administrations on these pregnancy-mediated adaptations. P4 administrations shortened the prolongation of QTC intervals and action potential duration (APD) that occurred during pregnancy, which was mainly attributable to the reduction in the voltage-gated potassium (Kv) current under basal conditions. In vitro studies indicated that P4 regulated the Kv2.1 channel in a bidirectional manner. At a low dose (1 µM), P4 induced upregulation of Kv2.1 through P4 receptor, while at a higher dose (5 µM), P4 downregulated Kv2.1 by targeting microRNA-29b (miR-29b). Our data showed that P4 modulated maternal cardiac repolarization by regulating Kv2.1 channel activity during pregnancy. Kv2.1, as well as miR-29b, might be used as potential therapeutic targets for adaptations of the maternal cardiovascular system or evaluation of progesterone medication during pregnancy.

Downregulation of Kcnq1ot1 attenuates ß-cell proliferation and insulin secretion via the miR-15b-5p/Ccnd1 and Ccnd2 axis.

AIM: To examine the effect of lncRNA Kcnq1ot1 on pancreatic ß cells in the development of diabetes. METHODS: The expression levels of Kcnq1ot1 were detected in the islets of diabetes mouse models and the serum of patients with type 2 diabetes by qRT-PCR. CCK8, Ki67 staining, immunohistochemical analyses, glucose-stimulated insulin secretion and intraperitoneal glucose tolerance test were performed to detect the effect of Kcnq1ot1 on ß-cell proliferation and insulin secretion in vitro and in vivo. The relationship between Kcnq1ot1 and miR-15b-5p was predicted by bioinformatics prediction, which was confirmed by luciferase reporter assay. RESULTS: Kcnq1ot1 was more abundant in the pancreas. The expression of Kcnq1ot1 was decreased in the islets of db/db mice and diet-induced obese mice and in the serum of patients with type 2 diabetes. Silencing Kcnq1ot1 inhibited the ß-cell proliferation concomitant with a reduction in the levels of Ccnd1 and Ccnd2. Insulin synthesis and secretion were impaired, along with the decreased expression of Ins1, Ins2, and insulin-related transcription factors. Moreover, Kcnq1ot1 knockdown in vivo reduced glucose tolerance and decreased insulin secretion, consistent with the reduction in the relative islet area and Ki67-positive ß-cells detected by immunochemistry and immunofluorescence staining, respectively. Mechanistically, Kcnq1ot1 directly targeted miR-15b-5p which regulated ß-cell proliferation and insulin secretion through Ccnd1 and Ccnd2. Notably, the suppression of miR-15b-5p attenuated the inhibition of Min6 proliferation and insulin production induced by Kcnq1ot1 knockdown. CONCLUSION: Kcnq1ot1 regulated ß-cell proliferation and insulin secretion via the miR-15b-5p/Ccnd1 and Ccnd2 axis, which is worthy of further investigation considering its potential in diabetes treatment.

Long Non-Coding RNA KCNQ1OT1 Regulates Protein Kinase CK2 Via miR-760 in Senescence and Calorie Restriction.

Long non-coding RNAs (lncRNAs) play important biological roles. Here, the roles of the lncRNA KCNQ1OT1 in cellular senescence and calorie restriction were determined. KCNQ1OT1 knockdown mediated various senescence markers (increased senescence-associated ß-galactosidase staining, the p53-p21Cip1/WAF1 pathway, H3K9 trimethylation, and expression of the senescence-associated secretory phenotype) and reactive oxygen species generation via CK2α downregulation in human cancer HCT116 and MCF-7 cells. Additionally, KCNQ1OT1 was downregulated during replicative senescence, and its silencing induced senescence in human lung fibroblast IMR-90 cells. Additionally, an miR-760 mimic suppressed KCNQ1OT1-mediated CK2α upregulation, indicating that KCNQ1OT1 upregulated CK2α by sponging miR-760. Finally, the KCNQ1OT1-miR-760 axis was involved in both lipopolysaccharide-mediated CK2α reduction and calorie restriction (CR)-mediated CK2α induction in these cells. Therefore, for the first time, this study demonstrates that the KCNQ1OT1-miR-760-CK2α pathway plays essential roles in senescence and CR, thereby suggesting that KCNQ1OT1 is a novel therapeutic target for an alternative treatment that mimics the effects of anti-aging and CR.

Curcumin Suppresses TGF-ß2-Induced Proliferation, Migration, and Invasion in Lens Epithelial Cells by Targeting KCNQ1OT1/miR-377-3p/COL1A2 Axis in Posterior Capsule Opacification.

Purpose: Posterior capsule opacification (PCO) is a common complication after cataract surgery, which can lead to secondary loss of vision. Curcumin has been reported to play a suppressive role in PCO progression, and the potential molecular mechanism was explored in this study. Methods: Cell viability and proliferation were analyzed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and 5-Ethynyl-2'-deoxyuridine (EdU) assay. Transwell assay and wound healing assay were performed to assess cell invasion and migration abilities. Western blot assay and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) were conducted to measure the expression of proteins and RNAs. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were conducted to confirm the interaction between microRNA-377-3p (miR-377-3p) and KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1) or collagen type I alpha 2 chain (COL1A2). Results: Curcumin dose-dependently alleviated transforming growth factor-ß2 (TGF-ß2)-induced proliferation, migration, and invasion in SRA01/04 cells. KCNQ1OT1 was up-regulated in PCO patients and TGF-ß2-induced SRA01/04 cells. Curcumin-induced protective effects in TGF-ß2-induced SRA01/04 cells were largely overturned by KCNQ1OT1 overexpression. KCNQ1OT1 directly interacted with miR-377-3p and negatively regulated its expression. miR-377-3p silencing overturned Curcumin-mediated protective effects in SRA01/04 cells upon TGF-ß2 treatment. miR-377-3p directly interacted with the 3' untranslated region (3'UTR) of COL1A2. COL1A2 overexpression largely counteracted KCNQ1OT1 silencing-induced effects in TGF-ß2-stimulated SRA01/04 cells. KCNQ1OT1 could up-regulate COL1A2 expression by sponging miR-377-3p in SRA01/04 cells. Conclusion: In conclusion, Curcumin suppressed TGF-ß2-induced malignant changes in lens epithelial cells by targeting KCNQ1OT1/miR-377-3p/COL1A2 axis.

LncRNA KCNQ1OT1 depletion inhibits the malignant development of atherosclerosis by miR-145-5p.

BACKGROUND: Atherosclerosis (AS) is a lipid-driven inflammatory disease of the arterial intima. Evidence is growing that dysregulation of lncRNAs is implicated in the pathogenesis of AS. In this research, the role of lncRNA KCNQ1OT1 in AS was investigated. METHODS: ApoE-/- mice were fed on a high fat diet to establish mouse models of AS. Macrophages (THP-1) were treated with oxidized low-density lipoprotein (ox-LDL) to establish cell models of AS. Atherosclerotic lesions of AS mice were determined by performing Oil red O staining. Lipid metabolic disorders and inflammatory were detected using specific assay kits. KCNQ1OT1 and miR-145-5p expression was measured using RT-qPCR. Levels of PPARα and CPT1 were measured using western blot. RESULTS: KCNQ1OT1 expression was upregulated and miR-145-5p was downregulated in atherosclerotic plaques of AS mice and ox-LDL-treated THP-1 cells. Lipid metabolic disorders and inflammation in vivo and in vitro were attenuated by either KCNQ1OT1 knockdown or miR-145-5p overexpression. Additionally, KCNQ1OT1 acted as a molecular sponge of miR-145-5p and downregulated miR-145-5p expression. Furthermore, silencing miR-145-5p abolished the effect of KCNQ1OT1 knockdown. CONCLUSION: Silencing KCNQ1OT1 attenuates AS progression by sponging miR-145-5p.

Membrane pools of phosphatidylinositol-4-phosphate regulate KCNQ1/KCNE1 membrane expression.

Plasma membrane phosphatidylinositol 4-phosphate (PI4P) is a precursor of PI(4,5)P2, an important regulator of a large number of ion channels. Although the role of the phospholipid PI(4,5)P2 in stabilizing ion channel function is well established, little is known about the role of phospholipids in channel membrane localization and specifically the role of PI4P in channel function and localization. The phosphatidylinositol 4-kinases (PI4Ks) synthesize PI4P. Our data show that inhibition of PI4K and prolonged decrease of levels of plasma membrane PI4P lead to a decrease in the KCNQ1/KCNE1 channel membrane localization and function. In addition, we show that mutations linked to Long QT syndrome that affect channel interactions with phospholipids lead to a decrease in membrane expression. We show that expression of a LQT1-associated C-terminal deletion mutant abolishes PI4Kinase-mediated decrease in membrane expression and rescues membrane expression for phospholipid-targeting mutations. Our results indicate a novel role for PI4P on ion channel regulation. Our data suggest that decreased membrane PI4P availability to the channel, either due to inhibition of PI4K or as consequence of mutations, dramatically inhibits KCNQ1/KCNE1 channel membrane localization and current. Our results may have implications to regulation of other PI4P binding channels.

KCNG1-Related Syndromic Form of Congenital Neuromuscular Channelopathy in a Crossbred Calf.

Inherited channelopathies are a clinically and heritably heterogeneous group of disorders that result from ion channel dysfunction. The aim of this study was to characterize the clinicopathologic features of a Belgian Blue x Holstein crossbred calf with paradoxical myotonia congenita, craniofacial dysmorphism, and myelodysplasia, and to identify the most likely genetic etiology. The calf displayed episodes of exercise-induced generalized myotonic muscle stiffness accompanied by increase in serum potassium. It also showed slight flattening of the splanchnocranium with deviation to the right side. On gross pathology, myelodysplasia (hydrosyringomielia and segmental hypoplasia) in the lumbosacral intumescence region was noticed. Histopathology of the muscle profile revealed loss of the main shape in 5.3% of muscle fibers. Whole-genome sequencing revealed a heterozygous missense variant in KCNG1 affecting an evolutionary conserved residue (p.Trp416Cys). The mutation was predicted to be deleterious and to alter the pore helix of the ion transport domain of the transmembrane protein. The identified variant was present only in the affected calf and not seen in more than 5200 other sequenced bovine genomes. We speculate that the mutation occurred either as a parental germline mutation or post-zygotically in the developing embryo. This study implicates an important role for KCNG1 as a member of the potassium voltage-gated channel group in neurodegeneration. Providing the first possible KCNG1-related disease model, we have, therefore, identified a new potential candidate for related conditions both in animals and in humans. This study illustrates the enormous potential of phenotypically well-studied spontaneous mutants in domestic animals to provide new insights into the function of individual genes.

Detection of Circulating Serum microRNA/Protein Complexes in ASD Using Functionalized Chips for an Atomic Force Microscope.

MicroRNAs, which circulate in blood, are characterized by high diagnostic value; in biomedical research, they can be considered as candidate markers of various diseases. Mature microRNAs of glial cells and neurons can cross the blood-brain barrier and can be detected in the serum of patients with autism spectrum disorders (ASD) as components of macrovesicles, macromolecular protein and low-density lipoprotein particles. In our present study, we have proposed an approach, in which microRNAs in protein complexes can be concentrated on the surface of AFM chips with oligonucleotide molecular probes, specific against the target microRNAs. MicroRNAs, associated with the development of ASD in children, were selected as targets. The chips with immobilized molecular probes were incubated in serum samples of ASD patients and healthy volunteers. By atomic force microscopy (AFM), objects on the AFM chip surface have been revealed after incubation in the serum samples. The height of these objects amounted to 10 nm and 6 nm in the case of samples of ASD patients and healthy volunteers, respectively. MALDI-TOF-MS analysis of protein components on the chip surface allowed us to identify several cell proteins. These proteins are involved in the binding of nucleic acids (GBG10, RT24, RALYL), in the organization of proteasomes and nucleosomes (PSA4, NP1L4), and participate in the functioning of the channel of active potassium transport (KCNE5, KCNV2).

Long non-coding RNA KCNQ1OT1 facilitates the progression of cervical cancer and tumor growth through modulating miR-296-5p/HYOU1 axis.

Literature reports that lncRNA KCNQ1OT1 is markedly up-regulated in cervical cancer (CC) tissues and cell lines, and KCNQ1OT1 can promote the proliferation and metastasis of CC cells. This current work was designed to investigate the molecular mechanism underlying the participation of KCNQ1OT1 in CC progression. Herein, RT-qPCR was utilized for determining the levels of KCNQ1OT1, miR-296-5p and HYOU1 in clinical tumor tissue specimens and CC cell lines. Then, starBase predicted the complementary binding sites of KCNQ1OT1 and miR-296-5p or miR-296-5p and HYOU1. Dual-luciferase reporter assay/RIP assay validated the interplays among KCNQ1OT1/miR-296-5p/HYOU1. In addition, CCK-8, wound healing and transwell assays were employed to assess the proliferative, migrative and invasive properties of CC cells. Moreover, nude mice xenograft model was established by subcutaneously injection with SiHa cells in order to validate the precise functions of KCNQ1OT1/miR-296-5p/HYOU1 axis in CC in vivo. Besides, Immunohistochemical staining examined Ki-67 expression in xenograft tumors and western blotting analysis detected expressions of MMP2/9 and Wnt/ß-catenin signaling pathway in CC cells and xenograft tumors. Elevated KCNQ1OT1 and HYOU1 as well as reduced miR-296-5p were observed in clinical tumor tissue specimens and CC cell lines. Results revealed that upregulation of miR-296-5p counteracted the enhancing effects of overexpressed KCNQ1OT1 on the proliferative, migrative and invasive abilities of CC cells. Additionally, HYOU1 overexpression abolished the suppressing effects of silenced KCNQ1OT1 on the malignant behaviors of CC cells and tumor growth. To conclude, KCNQ1OT1 could aggravate the malignant behaviors of CC and facilitate tumor growth through modulating miR-296-5p/HYOU1 axis.