NFATC3–PLA2G15 Fusion Transcript Identified by RNA Sequencing Promotes Tumor Invasion and Proliferation in Colorectal Cancer Cell Lines / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: This study was designed to identify novel fusion transcripts (FTs) and their functional significance in colorectal cancer (CRC) lines. MATERIALS AND METHODS: We performed paired-end RNA sequencing of 28 CRC cell lines. FT candidates were identified using TopHat-fusion, ChimeraScan, and FusionMap tools and further experimental validation was conducted through reverse transcription-polymerase chain reaction and Sanger sequencing. FT was depleted in human CRC line and the effects on cell proliferation, cell migration, and cell invasion were analyzed. RESULTS: One thousand three hundred eighty FT candidates were detected through bioinformatics filtering. We selected six candidate FTs, including four inter-chromosomal and two intrachromosomal FTs and each FT was found in at least one of the 28 cell lines. Moreover, when we tested 19 pairs of CRC tumor and adjacent normal tissue samples, NFATC3–PLA2G15 FT was found in two. Knockdown of NFATC3–PLA2G15 using siRNA reduced mRNA expression of epithelial–mesenchymal transition (EMT) markers such as vimentin, twist, and fibronectin and increased mesenchymal–epithelial transition markers of E-cadherin, claudin-1, and FOXC2 in colo-320 cell line harboring NFATC3–PLA2G15 FT. The NFATC3–PLA2G15 knockdown also inhibited invasion, colony formation capacity, and cell proliferation. CONCLUSION: These results suggest that that NFATC3–PLA2G15 FTs may contribute to tumor progression by enhancing invasion by EMT and proliferation.

Effect of PARP1 through regulating NFATs on cardiac hypertrophy / 中国药理学通报

Aim To investigate the interaction and mechanism of PARP1 and NFATc3, NFATc4 in ISO-induced pathological cardiac hypertrophy. Methods To establish the model of cardiac hypertrophy in vitro and in vivo, primary neonatal rat cardiomyocytes were treated with ISO (10 jimol • L-1) for 24 h; SD rats were subcutaneously injected with 1. 2 mg • kg-1 • d-1 ISO for 7 d. The nuclear and cytoplasmic proteins were separated by Cellytic Nuclear Extraction Kit. The subcellular localization of NFATc3 and NAFTc4 were detected by Western blot and immunofluorescence. The recombinant adenovirus (Ad-PARPl) infection was used to overexpress PARP1 and knockdown PARP1 by transfecting with siRNA of PARP1 in cardiomyocytes. Results The models of cardiac hypertrophy were successfully built both in vivo and vitro by ISO. It was determined that NFATc3 and NFATc4 were transfered into the nuclear from the cytoplasm in primary neonatal rat cardiomyocytes (NRCMs) after being treated with ISO. And the enzymatic activity of PARP1 was boosted in TSO-trpatpH prmin. OvprpYnrp.ssinn of PARP1 nromo-ted the nuclear translocation of NFATc3 and NFATc4 in cardiomyocytes, while knockdown of PARP1 could reverse the nuclear translocation induced by ISO. PARP1 inhibitor 3AB retarded ISO-induced nuclear transportation of NFATc3 and NFATc4 to some extent. Conclusions ISO leads to the up-regulation of enzymatic activity of PARP1 and promotes nuclear translocation of NFATc3 and NFATc4, thus aggravating car-diac hypertrophy.

L-Carnitine inhibits H2 O2-mediated NFATc3 nuclear translocation / 中国病理生理杂志

AIM: To explore the effect of L-carnitine on nuclear factor of activated T-cells, cytoplasmic 3 (NFATc3) in cardiomyocytes under H2O2 stimulation.METHODS: Primary cultured neonatal rat myocardial cells were stimulated by H2 O2 at concentration of 200μmol/L for 12 h to induce oxidative stress injury.In treatment group, L-carni-tine and cyclosporin A ( CsA) , a specific inhibitor of calcineurin ( CaN) , were administered 30 min prior to H2 O2 stimula-tion.After treatment, total, cytoplasmic and nuclear NFATc3 protein levels were determined by Western blotting.The method of immunofluoresence was used to evaluate the distribution of NFATc3.RESULTS: H2 O2 treatment produced no effect on the expression of total NFATc3, but caused its translocation from the cytosolic to nuclear compartment, which was greatly blunted by L-carnitine pretreatment.CONCLUSION:L-carnitine antagonized oxidative stress injury via alleviating NFATc3 nuclear translocation.

NFATc1 and NFATc3 is Involved in the Expression of Receptor Activator of NF-kappaB Ligand in Activated T Lymphocytes

Receptor activator of NF-kappaB ligand (RANKL) is an essential cytokine for osteoclast differentiation, activation and survival. T lymphocytes such as T17 cells, a subset of T helper cells that produce IL-17, play an important role in rheumatoid arthritic bone resorption by producing inflammatory cytokines and RANKL. It has not yet been clearly elucidated how T cell activation induces RANKL expression. T cell receptor activation induces the activation of nuclear factor of activated T cell (NFAT) and expression of its target genes. In this study, we examined the role of NFAT in T cell activation-induced RANKL expression. EL-4, a murine T lymphocytic cell line, was used. When T cell activation was induced by phorbol 12-myristate 13-acetate (PMA) and ionomycin, RANKL expression increased in a time-dependent manner. In the presence of cyclosporin, an inhibitor of NFAT activation, this PMA/ionomycin-induced RANKL expression was blocked. Overexpression of either NFATc1 or NFATc3 induced RANKL expression. Chromatin immunoprecipitation results demonstrated that PMA/ionomycin treatment induced the binding of NFATc1 and NFATc3 to the mouse RANKL gene promoter. These results suggest that NFATc1 and NFATc3 mediates T cell receptor activation-induced RANKL expression in T lymphocytes.