Molecular insight into the potential functional role of pseudoenzyme GFOD1 via interaction with NKIRAS2.

The glucose-fructose oxidoreductase/inositol dehydrogenase/rhizopine catabolism protein (Gfo/Idh/MocA) family includes a variety of oxidoreductases with a wide range of substrates that utilize NAD or NADP as redox cofactor. Human contains two members of this family, namely glucose-fructose oxidoreductase domain-containing protein 1 and 2 (GFOD1 and GFOD2). While GFOD1 exhibits low tissue specificity, it is notably expressed in the brain, potentially linked to psychiatric disorders and severe diseases. Nevertheless, the specific function, cofactor preference, and enzymatic activity of GFOD1 remain largely unknown. In this work, we find that GFOD1 does not bind to either NAD or NADP. Crystal structure analysis unveils that GFOD1 exists as a typical homodimer resembling other family members, but lacks essential residues required for cofactor binding, suggesting that it may function as a pseudoenzyme. Exploration of GFOD1-interacting partners in proteomic database identifies NK-κB inhibitor-interacting Ras-like 2 (NKIRAS2) as one potential candidate. Co-immunoprecipitation (co-IP) analysis indicates that GFOD1 interacts with both GTP- and GDP-bound forms of NKIRAS2. The predicted structural model of the GFOD1-NKIRAS2 complex is validated in cells using point mutants and shows that GFOD1 selectively recognizes the interswitch region of NKIRAS2. These findings reveal the distinct structural properties of GFOD1 and shed light on its potential functional role in cellular processes.

Circ_0001955 promotes non-small cell lung cancer cell proliferation and invasion by regulating MiR-29a-3p/NKIRAS2 axis to activate the nuclear factor-κB pathway.

Non-small cell lung cancer (NSCLC) is one of the most common malignant tumors worldwide. Circular RNAs (circRNAs) have been widely reported to play a role in the pathogenesis of various tumors. Nevertheless, the function of circ_0001955 in NSCLC progression has not been explored yet. This study aims to explore the functions of circ_0001955 in NSCLC and investigate its regulatory molecular mechanism. First, we determined that circ_0001955 was upregulated in NSCLC cells. Subsequently, we demonstrated that knockdown of circ_0001955 restrained cell proliferation and invasion. In vivo experiments further proved the suppressive effect of circ_0001955 silence on tumor growth. Mechanism assays revealed that circ_0001955 enhanced nuclear factor-κB (NF-κB) inhibitor interacting Ras-like protein 2 (NKIRAS2) expression by sponging microRNA-29a-3p (miR-29a-3p). Upregulation of NKIRAS2 led to the deceased level of IκBß but increased levels of nuclear p65, thus activating the NF-κB signaling pathway. In conclusion, Circ_0001955 activates the NF-κB pathway to promote NSCLC cell proliferation and invasion by regulating miR-29a-3p/NKIRAS2 axis.

MiR-1180 promotes cardiomyocyte cell cycle re-entry after injury through the NKIRAS2-NFκB pathway.

Heart failure (HF) is associated with a considerable number of symptoms and significantly impaired health for humans, including reduced quality of life and physical functioning. Previous studies have indicated that miRNAs have important roles in regulating the development of HF. MiR-1180 is involved in the proliferation, migration, invasiveness, and chemoresistance of cancer cells; however, the underlying mechanisms and role of miR-1180 in the functioning of cardiomyocytes remains unclear. In this study, we found that miR-1180 promotes cell activity and cell cycle processes by driving energy generation through NKIRAS2, which declines over time during development. The expression of miR-1180 is down-regulated in cells subjected to hypoxia-reoxygenation, and use of an miR-1180 mimic significantly reduced myocardial injury and cell apoptosis. In addition, miR-1180 regulates the NFκB pathway through NKIRAS2 in cardiomyocytes. These findings suggest that miR-1180 maybe a novel therapeutic target for use in getting cardiomyocytes to re-enter the cell cycle as well as for cardiac repair following myocardial injury.

miRNA-125b Regulates Osteogenic Differentiation of Periodontal Ligament Cells Through NKIRAS2/NF-κB Pathway.

BACKGROUND/AIMS: Osteogenesis of periodontal ligament cells (PDLCS) is essential for alveolar bone repair. Varieties of factors have been found involved in the regulation of PDLCs osteoblast differentiation. Aim of this study was to identify microRNA as a regulator of the os-teogenic differentiation of PDLCs. METHODS: The CD markers were analyzed by flow cytometry analysis. Osteoblast differentiation of PDLCs was induced by treatment with dexamethasone, ß-glycerol phosphate and α-ascorbic acid. The expression of osteoblastic phenotype was evaluated after the induction by simultaneous monitoring of alkaline phosphatase activity, the expression of genes involved in osteoblastic differentiation by RT-qPCR and Western Blot, and mineralization at the same time. MicroRNA and NKIRAS2 expression was determined by RT-qPCR. Luciferase reporter assays were performed to test whether miR-125b is capable of interacting with the 3'UTR sequence of NKIRAS2. The possible signaling pathway was determined by Western Blot. RESULTS: In this study, we found that the expression of miR-125b was down regulated during the process of ostoblast differentiation of PDLCs. When the expression of miR-125b was up regulated, the osteogenic differentiation of PDLCs was inhibited. During this process, the over-expressed miR-125b led to the activation of NF-κB. NF-κB inhibitor interacting RAS-like 2 (NKIRAS2) is one of target gene of miR-125b, and it is a regulator of NF-κB signaling that plays various roles in osteoblastic differentiation. We demonstrate thatmiR-125b is involved in osteogenic differentiation of PDLCs. CONCLUSION: Our data support the hypothesis that that miR-125b attenuates PDLCs osteoblastic differentiation by targeting NKIRAS2 and enhancing NF-κB signaling.

Characterization of LncRNA SNHG22 as a protector of NKIRAS2 through miR-4492 binding in osteosarcoma.

Many studies have revealed the function of long noncoding RNA (LncRNA) in regulating tumorigenesis of osteosarcoma (OS). As a subgroup of LncRNA, small nucleolar RNA host genes (SNHGs) have emerged as potentially important in OS. According to our recent findings, small nucleolar RNA host gene 22 (SNHG22) plays an important role in inhibiting the growth and metastasis of OS. However, the underlying mechanism of SNHG22 in regulating OS progression remains unknown. In this study, we confirmed that SNHG22 was downregulated in OS, and the overexpression of SNHG22 significantly inhibited OS progression in vivo and in vitro. Meanwhile, overexpression of SNHG22 also inhibited the migration and proliferation of human umbilical vein endothelial cells (HUVECs) and prevented the epithelial-to-mesenchymal transition (EMT) in OS. Furthermore, the interaction between miR-4492 and SNHG22 we previously predicted was validated by RNA pull-down assays and RNA immunoprecipitation assays. Dual-luciferase reporter assays showed that SNHG22 could directly interact with miR-4492 and upregulate the expression of NK-κB inhibitor-interacting Ras-like 2 (NKIRAS2) by its competing endogenous RNA (ceRNA) activity on miR-4492. In conclusion, our study has clarified the function of SNHG22 in OS progression and suggests a novel therapeutic target for OS.

Epstein-Barr virus-coded miR-BART13 promotes nasopharyngeal carcinoma cell growth and metastasis via targeting of the NKIRAS2/NF-κB pathway.

Based on analysis of Epstein-Barr virus (EBV) BART microRNA expression profiles, we previously reported that EBV-encoded miR-BART13 is upregulated in nasopharyngeal carcinoma (NPC) plasma specimens. However, the effects and molecular mechanisms of miR-BART13 in NPC remain largely unknown. We found that miR-BART13 was significantly upregulated in NPC tissue specimens. Ectopic expression of miR-BART13 promoted NPC cell proliferation, epithelial mesenchymal transition, and metastasis in vitro, and facilitated xenograft tumor growth and lung metastasis in vivo. Molecularly, NF-κB inhibitor interacting Ras-like 2 (NKIRAS2), a negative regulator of the NF-κB signaling, was identified to be a direct target of miR-BART13 in NPC cells, and NKIRAS2 mRNA and protein expression was inversely correlated with miR-BART13 in NPC tissues, respecitvely. Furthermore, the NF-κB signaling pathway was activated by miR-BART13. By rescued experiments, reconstitution of NKIRAS2 expression abrogated all the phenotypes upregulated by miR-BART13, and attenuated activity of NF-κB signaling pathway activated by miR-BART13 in NPC cells. Our findings indicated the newly identified miR-BART13/NKIRAS2/NF-κB signaling axis may provide further insights into better understanding of NPC initiation and development, and targeting of this pathway could be further studied as a therapeutic strategy for NPC patients.

MicroRNA-1908-5p contributes to the oncogenic function of the splicing factor SRSF3.

Serine/arginine (SR)-rich proteins that contain RS domains and SR repeats have diverse cellular functions including transcription, polyadenylation, translation, and RNA export. The splicing factor SRSF3, also termed SRp20, is the smallest member of the SR protein family and is a known proto-oncogene. Although it is implicated in the malignant phenotypes of various cancer cells, the molecular mechanism underlying SRSF3-mediated cancer progression is still obscure. We investigated here the oncogenic functions of SRSF3 in osteosarcoma U2OS cells. Knockdown of SRSF3 inhibited proliferation, clonogenicity, and metastatic potential including migration and invasion. It also decreased the level of miR-1908 independent of its host gene FADS1. Although FADS1 was not associated with SRSF3-mediated malignant properties, overexpression of miR-1908-5p increased cell proliferation, migration, and invasion, suggesting that miR-1908-5p is responsible for the oncogenic functions of SRSF3. Knockdown of SRSF3 decreased the expression of miR-1908-5p by inhibiting transactivation of NF-κB. We observed that miR-1908-5p downregulated NF-κB inhibitor interacting Ras-like 2 (NKIRAS2), a negative regulator of the NF-κB pathway by directly binding to the 3'UTR of NKIRAS2 mRNA. Consistent with overexpression of miR-1908-5p, knockdown of NKIRAS2 diminished the expression level of IκB-ß and provoked translocation of NF-κB into the nucleus where it transcriptionally activates its target genes including miR-1908-5p expression, thus elevating the proliferation and metastatic potential. Taken together, our results demonstrate that SRSF3 confers the malignant characteristics on cancer cells via the SRSF3/miR-1908-5p/NKIRAS2 axis.