Selenoprotein dSelK in Drosophila elevates release of Ca2+ from endoplasmic reticulum by upregulating expression of inositol 1,4,5-tris-phosphate receptor

Abstract

dSelK (G-rich), a homolog of human and mouse SelK, is one of three selenoproteins in Drosophila melanogaster. It is the only trans-membrane selenoprotein in D. melanogaster integrated into both the endoplasmic reticulum (ER) membrane and the Golgi apparatus. The gene expression profile of Drosophila Schneider 2 (S2) cells after the dsRNA interference (dsRNAi) targeting of dSelK was examined with the GeneChip Drosophila Genome 2.0 Array (Affymetrix), a high-density oligonucleotide microarray encompassing nearly the full Drosophila genome. The results showed that the transcriptional expression of eight genes whose proteins are located on (or related to) the ER or the Golgi apparatus was highly induced or repressed by the dsRNAi treatment. The mRNA levels of the inositol 1,4,5-tris-phosphate receptor (IP3 receptor), whose gene product is integrated into the ER membrane and regulates the release of Ca2+ from the ER to the cytosol, were significantly downregulated. In contrast, the expression of inositol 1,4,5-tris-phosphate kinase 1, which is a cytosolic protein with opposing functions to the IP3 receptor, was significantly upregulated. Quantitative real-time PCR verified these results. The concentration of intracellular free Ca2+ of the Drosophila S2 cells was significantly decreased after the knockdown of dSelK, whereas overexpression of dSelK significantly increased the intracellular free Ca2+ concentration. These results indicate that dSelK in D. melanogaster is involved in regulating the release of Ca2+ from the ER to the cytosol and may play important roles in the signal transduction pathways involving Ca2+ mobilization.

Selenoprotein SelK increases the secretion of insulin from MIN6 β cells

“…It was shown by ow cytometry that cytosolic free Ca 2+ level was elevated by the over-expression of mSelK and declined by the knockdown of mSelK in MIN6 b cells compared to the control group (Con) ( Fig. 2A and B), just like the results in Drosophila S2 cell line and human gastric cancer BGC 823 cell line by our group 35,36 and in T cells, neutrophils, and macrophages by Verma et al 37 And it was shown that aer the over-expression/knockdown of mSelK, the intracellular free Ca 2+ level was increased/ decreased to 149%/70% compared to the blank control group (taking blank control group as 100%), which suggested that the variation of free Ca 2+ level in MIN6 b cells by the overexpression/knockdown of mSelK was similar to that of the release of insulin.…”

“…In our previous studies, it was shown that selenoprotein SelK in Drosophila S2 cells and human gastric cancer BGC 823 cells could increase the expression of inositol trisphosphate receptor (IP3R) on the ER, which increased the level of intracellular free calcium by promoting the release of Ca 2+ from ER to cytosol. 35,36 And this effect has been veried by others. 37,38 Further studies revealed that the stable expression of IP3R needs the palmitoylation of the protein, which requires the catalysis by an enzyme complex composed of DHHC6 (letters represent the amino acids aspartic acid, histidine, and cysteine in the catalytic domain) and selenoprotein SelK.…”

“…[29][30][31][32][33] In our previous studies, it is shown that selenoprotein SelK in Drosophila S2 cells and human gastric cancer BGC 823 cells can increase the level of cytosolic free Ca 2+ by promoting the release of Ca 2+ from the ER to cytosol. 35,36 In order to illustrate the inherent mechanism of the increased insulin secretion, the inuence of the overexpression and knockdown of mSelK on the cytosolic free Ca 2+ level in MIN6 b cells was also investigated. It was shown by ow cytometry that cytosolic free Ca 2+ level was elevated by the over-expression of mSelK and declined by the knockdown of mSelK in MIN6 b cells compared to the control group (Con) ( Fig.…”

“…38 The increased expression of IP3R by the SelK up-regulation stimulates the release of Ca 2+ from the ER calcium store to the cytosol of the cells. 35,36 IP3R is a ligand-gated ion channel protein presented on ER and nuclear envelope, which was originally identied in mouse pancreas cells. IP3R is classied in ve types, among which IP3R types 1, 2, and 3 (IP3R1, IP3R2, and IP3R3) are expressed in islet b cells and IP3R3 is the most abundant in this gland.…”

Expression and purification of the membrane enzyme selenoprotein K

“…In a different study, SelK knockout mice exhibited deficiencies in calcium-related functions in the immune system [9]. A follow up study has shown that SelK undergoes proteolytic cleavage by m-calpain following Toll-like receptor activation [13] and is involved in regulation of inositol 1,4,5-tris-phosphate receptors [14]. …”