Phosphorylation-Dependent Targeting of cAMP Response Element Binding Protein to the Ubiquitin/Proteasome Pathway in Hypoxia

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 97; no. 22; pp. 12091 - 12096
• Main Authors: Taylor, Cormac T., Furuta, Glenn T., Synnestvedt, Kristin, Colgan, Sean P.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 24-10-2000; National Acad Sciences; National Academy of Sciences; The National Academy of Sciences
• Subjects: Amino Acid Sequence; Anaerobic conditions; Animals; Base Sequence; Biological Sciences; Cell Hypoxia; Cyclic AMP Response Element-Binding Protein - metabolism; Cysteine Endopeptidases - metabolism; DNA Primers; Epithelial cells; Genes; HeLa Cells; Humans; Hypoxia; Messenger RNA; Mice; Models, Animal; Multienzyme Complexes - metabolism; Oxygen; Phenotypes; Phosphatases; Phosphorylation; Proteasome Endopeptidase Complex; Proteins; Response elements; Tissues; Transcription factors; Transcription, Genetic; Ubiquitins - metabolism
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.220211797

Hypoxia activates a number of gene products through degradation of the transcriptional coactivator cAMP response element binding protein (CREB). Other transcriptional regulators (e.g., β -catenin and NF-κ B) are controlled through phosphorylation-targeted proteasomal degradation, and thus, we hypothesized a similar degradative pathway for CREB. Differential display analysis 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 previously 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, ubiquitination, and degradation (in vitro and in vivo). HIV-tat peptide-facilitated loading of intact epithelia with phosphopeptides corresponding to this proteasome targeting motif resulted in inhibition of CREB ubiquitination. Further studies revealed that PP1 inhibitors mimicked hypoxia-induced gene expression, whereas proteasome inhibitors reversed the hypoxic phenotype. Thus, hypoxia establishes conditions that target CREB to proteasomal degradation. These studies may provide unique insight into a general mechanism of transcriptional regulation by hypoxia.