X-linked Inhibitor of Apoptosis Protein (XIAP) Regulates PTEN Ubiquitination, Content, and Compartmentalization

Céline Van Themsche,Valérie Leblanc,Sophie Parent,Eric Asselin

doi:10.1074/jbc.c109.009522

Abstract

Apoptotic cell death plays a normal role in various physiological processes, and deregulated apoptosis is a hallmark of several diseases, including cancer. Cell fate is dictated by the balance between pro- and antiapoptotic factors. Akt is one of these antiapoptotic factors, which must be activated through phosphorylation. The phosphorylation of Akt has previously been shown to be promoted by X-linked inhibitor of apoptosis protein (XIAP), another antiapoptotic protein dictating the fate of normal and cancer cells. However, the underlying mechanisms are poorly understood. We have observed that XIAP associates with PTEN (phosphatase and tensin homolog deleted on chromosome ten), the best characterized negative regulator of Akt phosphorylation, in vitro and in vivo. XIAP knockdown reduces constitutive mono- and polyubiquitination of PTEN, increases PTEN protein levels, and prevents nuclear accumulation of PTEN. Overexpression of XIAP induces polyubiquitination of PTEN and proteasome-dependent decrease of PTEN protein levels. RNA interference experiments showed that XIAP-induced regulation of Akt phosphorylation is PTEN-dependent. Additional experiments confirmed that XIAP also regulates PTEN in vivo; primary mouse embryonic fibroblasts derived from XIAP(-/-) mice contain higher levels of PTEN protein, less mono- and polyubiquitinated PTEN, and less nuclear PTEN than primary mouse embryonic fibroblasts derived from XIAP(+/+) mice. Finally, we found that XIAP can directly ubiquitinate PTEN in vitro. We thus propose that XIAP acts as an E3 ubiquitin ligase for PTEN and promotes Akt activity by regulating PTEN content and compartmentalization.

Highlights

In normal and cancer cells, balance between survival and apoptosis is maintained by a complex network of proapoptotic and antiapoptotic factors
X-linked inhibitor of apoptosis protein (XIAP) Decreases PTEN Content to Promote Akt Activity—In accordance with a role for XIAP in promoting Akt activity, knockdown of XIAP using RNAi reduces the phosphorylation of Akt, as shown by a decrease of P-Akt levels when compared with total Akt levels (Fig. 1A)
We have found that XIAP knockdown increased PTEN protein levels (Fig. 1A)

Summary

ACCELERATED PUBLICATION

X-linked Inhibitor of Apoptosis Protein (XIAP) Regulates PTEN Ubiquitination, Content, and Compartmentalization*□S. Akt is one of these antiapoptotic factors, which must be activated through phosphorylation. The phosphorylation of Akt has previously been shown to be promoted by X-linked inhibitor of apoptosis protein (XIAP), another antiapoptotic protein dictating the fate of normal and cancer cells. We propose that XIAP acts as an E3 ubiquitin ligase for PTEN and promotes Akt activity by regulating PTEN content and compartmentalization. X-linked inhibitor of apoptosis protein (XIAP) and Akt are two antiapoptotic factors acting on distinct targets. Because XIAP has been shown to induce degradation of proapoptotic factors [4, 5], we hypothesized that XIAP promotes Akt phosphorylation by inducing PTEN degradation. Using in vitro and in vivo models, we have investigated whether XIAP regulates PTEN ubiquitination and content

EXPERIMENTAL PROCEDURES

RESULTS

XIAP Associates with PTEN and Regulates PTEN Ubiquitination in

DISCUSSION