The maxi-anion channels (MACs) are expressed in cells from mammals to amphibians with ~60% exhibiting a phenotype called Maxi-Cl. Maxi-Cl serves as the most efficient pathway for regulated fluxes of inorganic and organic anions including ATP However, its molecular entity has long been elusive. By subjecting proteins isolated from bleb membranes rich in Maxi-Cl activity to LC-MS/MS combined with targeted siRNA screening, CRISPR/Cas9-mediated knockout, and heterologous overexpression, we identified the organic anion transporter SLCO2A1, known as a prostaglandin transporter (PGT), as a key component of Maxi-Cl. Recombinant SLCO2A1 exhibited Maxi-Cl activity in reconstituted proteoliposomes. When SLCO2A1, but not its two disease-causing mutants, was heterologously expressed in cells which lack endogenous SLCO2A1 expression and Maxi-Cl activity, Maxi-Cl currents became activated. The charge-neutralized mutant became weakly cation-selective with exhibiting a smaller single-channel conductance. silencing and respectively, suppressed the release of ATP from swollen C127 cells and from Langendorff-perfused mouse hearts subjected to ischemia-reperfusion. These findings indicate that SLCO2A1 is an essential core component of the ATP-conductive Maxi-Cl channel.

Ovarian cancer is a highly lethal cancer in females. Therefore, it is necessary to explore effective biomarkers for the diagnosis and prognosis of the disease. Stratifin (SFN) is a cell cycle checkpoint protein that has been reported to be involved in oncogenesis. Our studies detected the expression of SFN in ovarian cancer by Oncomine, Human Protein Atlas database and ULCAN database. Meanwhile, we found its coexpression gene by cBioPortal online tool and validated their expression in different ovarian cancer cells by western blot and reverse transcription quantitative PCR. Then, we also investigated their prognostic values via the Kaplan–Meier plotter database in different subtypes of ovarian cancer patients. The results demonstrated that SFN was found to be increased in ten various ovarian cancer datasets, compared with healthy tissues. Additionally, up-regulation of SFN expression is associated with age and cancer grades. The higher expression of SFN in all patients with ovarian cancers is significantly correlated with worse postprogression survival. In addition, high SFN expression is associated with significantly worse overall survival in patients who received chemotherapy contains gemcitabine, taxol, taxol+platin, paclitaxel and avastin. In human ovarian carcinoma SKOV3 and A2780 cells, the expression of SFN and its coexpression gene MICB were also increased at protein and mRNA levels compared with the normal ovarian epithelial cells. Based on above results, overexpression of SFN was correlated with the prognosis in ovarian cancer. The present study might be useful for better understanding the clinical significance of SFN mRNA.

The slow component of the delayed rectifier K(+) current (I Ks) plays an important role in the repolarization of action potentials in cardiac pacemaker cells and ventricular myocytes, and is regulated by various signaling pathways. Recent evidence has shown that calmodulin (CaM) is involved in modulation of diverse ion channels in cardiac myocytes under physiological and pathophysiological conditions. In the present study, we examined regulation of I Ks by Ca(2+)/CaM in guinea pig sinoatrial (SA) node cells using the whole-cell patch-clamp method. The density of I Ks was larger during intracellular dialysis with a higher Ca(2+) concentration (pCa 7, Ca (+)) compared to that with a low Ca(2+) concentration (pCa 10, Ca (-)). Intracellular application of CaM (400 nM) markedly potentiated I Ks with a Ca (+) pipette solution but not with a Ca (-) solution, thus showing that CaM potentiates I Ks in an intracellular Ca(2+)-dependent manner. Intracellular application of a specific Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) inhibitor, autocamtide-2 inhibitory peptide (AIP, 500 nM), markedly reduced I Ks activity in the presence of higher intracellular Ca(2+). Similarly, bath application of another inhibitor, KN-93 (1 μM) also significantly suppressed I Ks. Finally, the stimulatory action on I Ks of Ca(2+)/CaM was abolished by pretreatment with KN-93. Taken together, these observations suggest that Ca(2+)/CaM stimulates I Ks in guinea pig SA node cells through activation of CaMKII. This enhancement of I Ks by CaMKII may be involved in modulation of SA node automaticity under physiological or pathophysiological condition.

We herein investigated the effect of low temperature exposure on the expression, degradation, localization and activity of human Kv1.5 (hKv1.5). In hKv1.5-expressing CHO cells, the currents were significantly increased when cultured at a reduced temperature (28°C) compared to those observed at 37°C. Western blot analysis indicated that the protein levels (both immature and mature proteins) of hKv1.5 were significantly elevated under the hypothermic condition. Treatment with a proteasome inhibitor, MG132, significantly increased the immature, but not the mature, hKv1.5 protein at 37°C, however, there were no changes in either the immature or mature hKv1.5 proteins at low temperature following MG132 exposure. These observations suggest that the enhancement of the mature hKv1.5 protein at reduced temperature may not result from the inhibition of proteolysis. Moreover, the hKv1.5 fluorescence signal in the cells increased significantly on the cell surface at 28°C versus those cultured at 37°C. Importantly, the low temperature treatment markedly shifted the subcellular distribution of the mature hKv1.5, which showed considerable overlap with the trans-Golgi component. Experiments using tunicamycin, an inhibitor of N-glycosylation, indicated that the N-glycosylation of hKv1.5 is more effective at 28°C than at 37°C. Finally, the hypothermic treatment also rescued the protein expression and currents of trafficking-defective hKv1.5 mutants. These results indicate that low temperature exposure stabilizes the protein in the cellular organelles or on the plasma membrane, and modulates its maturation and trafficking, thus enhancing the currents of hKv1.5 and its trafficking defect mutants.