A role for Cdc37 in EGFRvIII biogenesis

Abstract

The mutant epidermal growth factor receptor, EGFRvIII, is associated with tumour aggressiveness and drug resistance in glioblastoma. Our lab has shown that the molecular chaperone Hsp90 interacts with nascent EGFRvIII, and that EGFRvIII expression and function is dependent upon Hsp90 activity. In tumours, Hsp90 is found exclusively in an active form, where it is bound to cochaperones. One of these cochaperones, Cdc37, was found in the Hsp90/nascent EGFRvIII complex. Cdc37's role is to recruit certain kinase proteins to Hsp90 for folding. Its ability to bind these kinases is thought to be dependent upon phosphorylation of serine 13 in its client-binding domain by the kinase CK2. Here I have shown that Cdc37 is a limiting factor in EGFRvIII maturation, and its absence or inactivity may limit recruitment to Hsp90 and result in degradation. Using mutant constructs designed to mimic non-phosphorylated and phosphorylated Cdc37, I evaluated the role of Cdc37 phosphorylation in the stability of nascent EGFRvIII. Expression of the phospho-mimetic Cdc37 construct raised protein levels of nascent and mature EGFRvIII, while the non-phosphorylatable Cdc37 S13A mutant could not. Approximately 100 times more of the phospho-mimetic construct bound to nascent EGFRvIII than did the non-phosphorylatable form, and this increased Cdc37 binding was associated with more Hsp90 binding as well. Treatment of cells with the selective CK2 inhibitor, TBB, lowered the amount of Cdc37 and Hsp90 bound to nascent EGFRvIII, and reduced EGFRvIII protein levels. Taken together, these data indicate a positive role for Cdc37 and CK2 in EGFRvIII biogenesis, and suggest that Cdc37 phosphorylation may be a key factor in the recruitment of client to the Hsp90 complex.