Patient-derived mutations within the N-terminal domains of p85α impact PTEN or Rab5 binding and regulation

The p85 alpha protein regulates flux through the PI3K/PTEN signaling pathway, and also controls receptor trafficking via regulation of Rab-family GTPases. In this report, we determined the impact of several cancer patient-derived p85 alpha mutations located within the N-terminal domains of p85 alpha previously shown to bind PTEN and Rab5, and regulate their respective functions. One p85 alpha mutation, L30F, significantly reduced the steady state binding to PTEN, yet enhanced the stimulation of PTEN lipid phosphatase activity. Three other p85 alpha mutations (E137K, K288Q, E297K) also altered the regulation of PTEN catalytic activity. In contrast, many p85 alpha mutations reduced the binding to Rab5 (L30F, I69L, I82F, I177N, E217K), and several impacted the GAP activity of p85 alpha towards Rab5 (E137K, I177N, E217K, E297K). We determined the crystal structure of several of these p85 alpha BH domain mutants (E137K, E217K, R262T E297K) for bovine p85 alpha BH and found that the mutations did not alter the overall domain structure. Thus, several p85 alpha mutations found in human cancers may deregulate PTEN and/or Rab5 regulated pathways to contribute to oncogenesis. We also engineered several experimental mutations within the p85 alpha BH domain and identified L191 and V263 as important for both binding and regulation of Rab5 activity.