Hyperphosphorylation and increased proteolytic breakdown of c-Myb induced by the inhibition of Ser/Thr protein phosphatases

Abstract

The c-myb proto-oncogene encodes a nuclear phosphoprotein that plays a crucial role in normal hematopoiesis. It is a short-lived transcription factor rapidly degraded by the 26S proteasome. Although it has been shown that instability determinants reside in its carboxyl terminus, the molecular mechanism of c-Myb degradation is unknown. Here, we report the first evidence that phosphorylation plays a role in targeting the protein to the proteasome. Inhibition of cellular serine/threonine protein phosphatase activity by okadaic acid resulted in hyperphosphorylation of c-Myb and extremely rapid turnover. The hyperphosphorylation resulted in a protein with altered properties that was indicative of conformational changes. Its mobility on gel electrophoresis was altered as well as its recognition by specific monoclonal antibody. The altered hyperphosphorylated protein still bound to DNA with an affinity similar to that of the hypophosphorylated form. Phosphorylation of three previously identified sites, serines 11, 12, and 528, does not appear to be involved in the proposed changes in conformation or stability. However, phosphoamino acid analyses of the hyperphosphorylated form of c-Myb revealed increased c-Myb phosphorylation mainly on threonine residues that correlated with other okadaic acid-induced alterations of c-Myb. These findings indicate that Ser/Thr phosphatases prevent conformational changes that may play an important role in controlled degradation of c-Myb. Oncogene (2000) 19, 2846 - 2854

Constitutive c-Myb amino-terminal phosphorylation and DNA binding activity uncoupled during entry and passage through the cell cycle

“…The protein is also subject to modi®cations that a ect protein-protein interaction (Ness, 1996) and proteosome-mediated degradation (Bies et al, 2000).…”

Covalent Attachment of the SUMO-1 Protein to the Negative Regulatory Domain of the c-Myb Transcription Factor Modifies Its Stability and Transactivation Capacity

“…In addition, our recent results suggest that proteolytic stability of c-Myb is modulated through phosphorylation of the carboxyl terminus of c-Myb. Inhibitors of Ser/Thr phosphatases rapidly induce hyperphosphorylation-dependent conformational changes in the NRD, followed by accelerated proteolytic breakdown (25,26).…”

Interferon-inducible Myc/STAT-interacting Protein Nmi Associates with IFP 35 into a High Molecular Mass Complex and Inhibits Proteasome-mediated Degradation of IFP 35

“…They also found that IFP 35 is phosphorylated and that complex formation correlates with IFP 35 dephosphorylation (21). The proteasomal degradation of many important cellular proteins including c-Myb, keratins, and Bcl-2 is modulated by their phosphorylation status (22)(23)(24). Therefore, IFP 35 levels and Nmi/IFP 35 complex formation and function may be regulated by both phosphorylation and proteasomal degradation.…”