Requirement of N-linked glycosylation in the luminal domain for optimal proteolytic activation of liver-enriched transcription factor CREB-H

Abstract

CREB-H is a liver-enriched bZIP transcription factor closely related to CREB3. CREB-H is activated by intramembrane proteolysis that releases an N-terminal fragment. Aberrant expression of CREB-H is implicated in hepatocellular carcinoma. In this study we characterized N-linked glycosylation of CREB-H which occurs at three sites in the C-terminal luminal domain. When all three sites were disrupted by sitedirected mutagenesis, N-linked glycosylation of CREB-H was completely abrogated. The CREB-H mutants defective for N-linked glycosylation poorly activated transcription driven by unfolded protein response element or C-reactive protein promoter. Upon stimulation with an activator of intramembrane proteolysis such as brefeldin A, unglycosylated CREB-H remained largely uncleaved, anchored to the endoplasmic reticulum membrane, and unable to activate transcription. Taken together, our findings suggest that N-linked glycosylation is necessary for full activation of CREB-H through intramembrane proteolysis. Our work also reveals a novel mechanism for post-translational regulation of the CREB3 subfamily of membrane-associated transcription factors.