A novel N‐terminal hydrophobic motif mediates constitutive degradation of serum‐ and glucocorticoid‐induced kinase‐1 by the ubiquitin–proteasome pathway

Abstract

Serum‐ and glucocorticoid‐induced protein kinase‐1 (SGK‐1) plays a critical role in regulation of the epithelial sodium channel, ENaC. SGK‐1 also shares significant catalytic domain homology with protein kinase B (PKB/AKT‐1) and is a downstream effector of antiapoptotic phosphoinositide 3‐kinase signaling. Steady‐state levels of an active SGK‐1 are tightly regulated by rapid transcriptional activation and post‐translational modification including phosphorylation. We show here that endogenous SGK‐1 protein is polyubiquitinated and rapidly degraded by the 26S proteasome. In contrast to other rapidly degraded kinases, neither the catalytic activity of SGK‐1 nor activation site phosphorylation was required for its ubiquitin modification and degradation. Instead, SGK‐1 degradation required a lysine‐less six‐amino‐acid (amino acids 19–24) hydrophobic motif (GMVAIL) within the N‐terminal domain. Deletion of amino acids 19–24 significantly increased the half‐life of SGK1 and prevented its ubiquitin modification. Interestingly, this minimal region was also required for the association of SGK‐1 with the endoplasmic reticulum. Ubiquitin modification and degradation of SGK‐1 were increasingly inhibited by the progressive mutation of six N‐terminal lysine residues surrounding the GMVAIL motif. Mutation of all six lysines to arginine did not disrupt the subcellular localization of SGK‐1 despite a significant decrease in ubiquitination, implying that this modification per se was not required for targeting to the endoplasmic reticulum. These results suggest that constitutive ubiquitin‐mediated degradation of SGK‐1 is an important mechanism regulating its biological activity.

Multiple Translational Isoforms Give Functional Specificity to Serum- and Glucocorticoid-induced Kinase 1

“…Recently we have shown that the N-terminus of Sgk1 contains an amphipathic ␣-helix that targets the protein to the ER membrane (Arteaga et al, 2006;Bogusz et al, 2006). This finding posed the question of how ER localization is compatible with the functions hitherto proposed for Sgk1, some of which require translocation of Sgk1 to the nucleus.…”

“…Posttranscriptionally, Sgk1 is activated by sequential phosphorylation on residues S422 and T253 by a still unidentified kinase (PDK2) and phosphatidylinositol-3-phosphate dependent kinase 1 (PDK1), respectively . The abundance of Sgk1 protein is maintained at low basal levels by rapid degradation mediated by the ubiquitin proteasomal machinery associated to the endoplasmic reticulum (ER) (Arteaga et al, 2006;Bogusz et al, 2006). Lastly, under certain stimuli Sgk1 translocates from the cytoplasm to the nucleus where it phosphorylates transcriptional factors .…”

A brain-specific SGK1 splice isoform regulates expression of ASIC1 in neurons

“…The canonical SGK1 contains an amphipathic ␣-helix that targets the protein to the cytosolic surface of the endoplasmic reticulum (22,29); SGK3 contains a PX domain that binds to PtdIns(3)P, thereby targeting the protein to early endosomes (30); SGK1.1 uses a cluster of cationic and large hydrophobic residues to bind PtdIns(4,5)P 2 and tether the protein to the plasma membrane. Such motifs have been described and shown to be necessary for targeting small guanosine triphosphatases (GTPases) from the Ras, Rho, Arf, and Rab subfamilies to the plasma membrane (24).…”

Deregulated expression of pro-survival and pro-apoptotic p53-dependent genes upon Elongator deficiency in colon cancer cells

“…Of note however and in contrast to BAX, we failed to see enhanced SGK expression at the protein level upon IKAP/hELP1 depletion (data not shown). This may be due to the intrinsic instability of SGK, which is known to be quickly degraded through the proteasome pathway [44]. In any case, we clearly show here that the upregulated SGK mRNA expression requires wild type p53 as SGK is no longer induced in p53 deficient HCT116 or in HT29 cells, where p53 is not functional.…”