Cyclin A-dependent phosphorylation of the ETS-related protein, MEF, restricts its activity to the G1 phase of the cell cycle.
MEF, a recently identified member of the E74 family of ETS-related transcription factors, is a strong transcriptional activator of cytokine gene expression. Using a green fluorescent protein gene reporter plasmid regulated by an MEF-responsive promoter, we determined that the transcriptional activity of MEF is largely restricted to the G1 phase of the cell cycle. MEF-dependent transcription was suppressed by the expression of cyclin A but not by cyclin D or cyclin E. This effect was due to the kinase activity generated by cyclin A expression, as co-expression of the cyclin-dependent kinase inhibitors p21 or p27, or a dominant negative form of CDK2 (DNK2), abrogated the reduction of MEF transcriptional activity by cyclin A. Cyclin A-CDK2 phosphorylated MEF protein in vitro more efficiently than cyclin D-CDK4 or cyclin E-CDK2, and phosphorylation of MEF by cyclin A-CDK2 reduced its ability to bind DNA. We determined one site of phosphorylation by cyclin A-CDK2 at the C terminus of MEF, using mass-spectrometry; mutation of three serine or threonine residues in this region significantly reduced phosphorylation of MEF by cyclin A and reduced cyclin A-mediated suppression of its transactivating activity. These amino acid substitutions also reduced the restriction of MEF activity to G1. Phosphorylation of MEF by the cyclin A-CDK2 complex controls its transcriptional activity during the cell cycle, establishing a novel link between the ETS family of proteins and the cell cycle machinery.
Miyazaki
Y
Y
Laboratory of Molecular Aspects of Hematopoiesis, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Boccuni
P
P
Mao
S
S
Zhang
J
J
Erdjument-Bromage
H
H
Tempst
P
P
Kiyokawa
H
H
Nimer
S D
SD
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D019925:Cyclin A / Q000378:metabolism*; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D016193:G1 Phase / Q000502:physiology*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D014158:Transcription, Genetic
D019925:Cyclin A; D017931:DNA Primers; D004268:DNA-Binding Proteins; D014157:Transcription Factors