Drak2 Is Upstream of p70S6 Kinase: Its Implication in Cytokine-Induced Islet Apoptosis, Diabetes, and Islet Transplantation

Abstract

Drak2 is a member of the death-associated protein family and a serine threonine kinase. In this study, we investigated its role in β cell survival and diabetes. Drak2 mRNA and protein were rapidly induced in islet β cells after stimulation by inflammatory lymphokines known to be present in type 1 diabetes. Drak2 up-regulation was accompanied by increased β cell apoptosis. β cell apoptosis caused by the said stimuli was inhibited by Drak2 knockdown using small interfering RNA. Conversely, transgenic Drak2 overexpression led to aggravated β cell apoptosis triggered by the stimuli. Further in vivo experiments demonstrated that Drak2 transgenic islets were more vulnerable to streptozocin insult. We established that inducible NO synthase was upstream and caspase-9 was downstream of Drak2 in its signaling pathway. Purified Drak2 could phosphorylate ribosomal protein S6 (p70S6) kinase in an in vitro kinase assay. Drak2 overexpression in NIT-1 cells led to enhanced p70S6 kinase phosphorylation, whereas Drak2 knockdown in these cells reduced it. These mechanistic studies proved that p70S6 kinase was a bona fide Drak2 substrate.

The Role of Drak2 in T Cell Function and Autoimmunity

“…While the role of Drak2 in the EAE model was intrinsic to the T cells, and due to increased susceptibility to apoptosis of autoreactive cells, other data suggest that Drak2 may contribute to T1D by affecting survival of pancreatic β cells [14]. Treatment of islet cells with inflammatory cytokines led to the upregulation of Drak2 expression in these cells, which correlated with islet apoptosis.…”

“…Although many lines of evidence indicate that Drak2 plays a critical role within the T cells, other data suggested that Drak2 expression in β-islet cells augments T1D [14,23]. Therefore, we investigated whether Drak2 expression in T cells, β-islet cells, or both cell types regulated disease in the NOD model of T1D.…”

“…Drak2 is expressed most abundantly in lymphoid organs, although expression was also detected in various tissues during embryogenesis, in some tumor types, and in pancreatic β-islet cells after stimulation [4,5,[9][10][11][12][13][14]. Within the lymphoid compartment, Drak2 expression increases during T and B cell development, and remains abundant in mature T and B cells.…”

“…In vitro kinase assays revealed that, in addition to phosphorylating itself, Drak2 phosphorylates myosin light chain as an exogenous substrate [7,9]. Drak2 also phosphorylated p70S6 kinase, which plays important roles in protein synthesis and cell cycle progression [14]. Drak2 overexpression in NIT-1 cells increased p70S6 kinase phosphorylation, while siRNA-mediated knockdown of Drak2 resulted in reduced p70S6 kinase phosphorylation.…”

“…Drak2 augmented free fatty acid (FFA)-mediated apoptosis in vitro in primary β-islet cells and NIT-1 insulinoma cells [13]. Furthermore, transgenic overexpression of Drak2 via the β-actin promoter increased apoptosis in islet cells following FFA stimulation [14]. Drak2 overexpression in these models mediated apoptosis via reduced induction of the anti-apoptotic family members Bcl-2, Bcl-xL, and Flip.…”