An investigation of the kinase dependent functions of CRaf

The CRaf protein kinase plays a key role in relaying proliferation, differentiation and survival signals from activated RAS or BRaf proteins to downstream effectors and is frequently hyperactivated in human cancers. Because of this essential role in signalling, CRaf kinase activity must be tightly regulated in the cell. A well-established model involves RASmediated translocation of CRaf to the plasma membrane where it is activated by a series of events including phosphorylation. Here, an important new mechanism of regulation that is distinct from the classical model has been discovered. Through the use of kinase defective crafD486A murine embryonic fibroblasts, or by the use of the Raf inhibitor sorafenib, we show that the principal function of the CRaf kinase activity is autophosphorylation of serine 621. This phosphorylation occurs in cis, does not involve MEK/ERK activation and is essential to ensure the correct folding and stability of the protein. This novel mechanism of regulation has significant implications for the control of CRaf activity in fundamental cellular processes in response to signalling from the onco-proteins RAS and BRaf. Indeed here it is demonstrated that CRaf promotes cell cycle progression, specifically the transition into S phase. Investigation of mutant craf cells demonstrated that this form of regulation is mediated in part by CRaf transcriptional repression of the CDK inhibitor p21^{CIP1}.