Phosphorylation-dependent targeting of cAMP response element binding protein to the ubiquitin/proteasome pathway in hypoxia

Hypoxia activates a number of gene products through degradation of the tran scriptional coactivator cAMP response element binding protein (CREB). Other transcriptional regulators (e.g., beta -catenin and NF-kappaB) are control led through phosphorylation-targeted proteasomal degradation, and thus, we hypothesized a similar degradative pathway for CREB. Differential display a nalysis of mRNA derived from hypoxic epithelia revealed a specific and time -dependent repression of protein phosphatase 1 (PP1), a serine phosphatase important in CREB dephosphorylation. Subsequent studies identified a previo usly unappreciated proteasomal-targeting motif within the primary structure of CREB (DSVTDS), which functions as a substrate for PP1. Ambient hypoxia resulted in temporally sequential CREB serine phosphorylation, ubiquitinati on, and degradation tin vitro and in vivo). HIV-tat peptide-facilitated loa ding of intact epithelia with phosphopeptides corresponding to this proteas ome targeting motif resulted in inhibition of CREB ubiquitination. Further studies revealed that PP1 inhibitors mimicked hypoxia-induced gene expressi on, whereas proteasome inhibitors reversed the hypoxic phenotype. Thus, hyp oxia establishes conditions that target CREB to proteasomal degradation. Th ese studies may provide unique insight into a general mechanism of transcri ptional regulation by hypoxia.