Regulation of the erythroid transcription factor NF-E2 by cyclic adenosine monophosphate-dependent protein kinase.

Abstract

Activation of cyclic adenosine monophosphate (cAMP)-dependent protein kinase (A-kinase) promotes hemoglobin synthesis in several erythropoietin-dependent cell lines, whereas A-kinase-deficient murine erythroleukemia (MEL) cells show impaired hemoglobin production; A-kinase may regulate the erythroid transcription factor NF-E2 by directly phosphorylating its p45 subunit or by changing p45 interactions with other proteins. We have mapped the major A-kinase phosphorylation site of p45 to Ser(169); Ala substitution for Ser(169) resulted in a protein that was no longer phosphorylated by A-kinase in vitro or in vivo. The mutant protein formed NF-E2 complexes that bound to DNA with the same affinity as wild-type p45 and functioned normally to restore beta-globin gene expression in a p45-deficient MEL cell line. Transactivation properties of the (Ser (169)--> Ala) mutant p45 were also indistinguishable from wild-type p45 when Gal4-p45 fusion constructs were tested with a Gal4-dependent reporter gene. Transactivation of the reporter by both mutant and wild-type p45 was significantly enhanced when A-kinase was activated by membrane-permeable cAMP analogs or when cells were cotransfected with the catalytic subunit of A-kinase. Stimulation of p45 transactivation by A-kinase required only the N-terminal transactivation domain of p45, suggesting that A-kinase regulates the interaction of p45 with downstream effectors.

• Authors: D Casteel, M Suhasini, T Gudi, R Naima, R B Pilz
• Journal: Blood (Blood) Vol. 91 Issue 9 Pg. 3193-201 (May 01 1998) ISSN: 0006-4971 [Print] United States
• PMID: 9558374 (Publication Type: Journal Article, Research Support, U.S. Gov't, Non-P.H.S.)

Chemical References

• DNA-Binding Proteins
• Erythroid-Specific DNA-Binding Factors
• NF-E2 Transcription Factor
• NF-E2 Transcription Factor, p45 Subunit
• Nfe2 protein, mouse
• Trans-Activators
• Transcription Factors
• Erythropoietin
• Phosphoserine
• Cyclic AMP
• Histone Acetyltransferases
• Ncoa3 protein, mouse
• Nuclear Receptor Coactivator 3
• Cyclic AMP-Dependent Protein Kinases

Topics

• Animals
• Cyclic AMP (physiology)
• Cyclic AMP-Dependent Protein Kinases (physiology)
• DNA-Binding Proteins (metabolism)
• Erythroid-Specific DNA-Binding Factors
• Erythropoiesis
• Erythropoietin (physiology)
• Histone Acetyltransferases
• Mice
• Mutagenesis, Site-Directed
• NF-E2 Transcription Factor
• NF-E2 Transcription Factor, p45 Subunit
• Nuclear Receptor Coactivator 3
• Phosphoserine (metabolism)
• Structure-Activity Relationship
• Trans-Activators (physiology)
• Transcription Factors (metabolism)
• Transcriptional Activation