Regulation of APC/C by a protein complex containing the 14-3-3 homologs Bmh1 and Bmh2.
Mark Hall, Brian Billings, Mary Jeong, Juan Martinez, Anindya Chatterjee
Biochemistry Department, Purdue University, 175 S. University St, West Lafayette, IN, 47907, United States
The anaphase-promoting complex, or cyclosome, (APC/C) is an ubiquitin ligase that regulates cell division by targeting specific substrates, such as cyclins, for degradation. During G1, APC/C is activated by Cdh1. Other groups have shown that APC/C inactivation at the G1/S transition requires inhibitory phosphorylation of Cdh1 by CDK and involves relocalization of Cdh1 to the cytoplasm. We purified a stable Cdh1 protein complex from cells arrested in early S and late M and identified the components Bmh1, Bmh2, and a previously uncharacterized protein (which we name Acm1) by mass spectrometry. Assembly of this complex is dependent on the presence of Acm1, which is absent during most of G1 and appears in late G1. acm1 deletion results in a slow growth phenotype resulting from a pronounced G1 delay. Additional deletion of cdh1 partially rescues the slow growth phenotype and alleviates the G1 delay, suggesting that a primary function of this complex is to help inactivate Cdh1-dependent APC/C activity at the G1/S transition. Further mass spec analysis revealed that Acm1 is heavily phosphorylated on several serine and threonine residues, including multiple CDK consensus sites, and we are exploring the role of Acm1 phosphorylation in the assembly and disassembly of the Cdh1 complex using site-specific mutants. We present a model for the role of this complex and its 14-3-3 subunits in deactivation of Cdh1-APC/C at the G1/S transition and reactivation at mitotic exit.