Shank-interacting protein–like 1 promotes tumorigenesis via PTEN inhibition in human tumor cells
Lizhi He, Alistair Ingram, Adrian P. Rybak, Damu Tang
Lizhi He, Alistair Ingram, Adrian P. Rybak, Damu Tang
View: Text | PDF
Research Article Oncology

Shank-interacting protein–like 1 promotes tumorigenesis via PTEN inhibition in human tumor cells

Abstract

Inactivation of phosphatase and tensin homolog (PTEN) is a critical step during tumorigenesis, and PTEN inactivation by genetic and epigenetic means has been well studied. There is also evidence suggesting that PTEN negative regulators (PTEN-NRs) have a role in PTEN inactivation during tumorigenesis, but their identity has remained elusive. Here we have identified shank-interacting protein–like 1 (SIPL1) as a PTEN-NR in human tumor cell lines and human primary cervical cancer cells. Ectopic SIPL1 expression protected human U87 glioma cells from PTEN-mediated growth inhibition and promoted the formation of HeLa cell–derived xenograft tumors in immunocompromised mice. Conversely, siRNA-mediated knockdown of SIPL1 expression inhibited the growth of both HeLa cells and DU145 human prostate carcinoma cells in vitro and in vivo in a xenograft tumor model. These inhibitions were reversed by concomitant knockdown of PTEN, demonstrating that SIPL1 affects tumorigenesis via inhibition of PTEN function. Mechanistically, SIPL1 was found to interact with PTEN through its ubiquitin-like domain (UBL), inhibiting the phosphatidylinositol 3,4,5-trisphosphate (PIP3) phosphatase activity of PTEN. Furthermore, SIPL1 expression correlated with loss of PTEN function in PTEN-positive human primary cervical cancer tissue. Taken together, these observations indicate that SIPL1 is a PTEN-NR and that it facilitates tumorigenesis, at least in part, through its PTEN inhibitory function.

Authors

Lizhi He, Alistair Ingram, Adrian P. Rybak, Damu Tang

×

Figure 7

SIPL1 reduces PTEN’s PIP3 phosphatase activity in the cell.

Options: View larger image (or click on image) Download as PowerPoint
SIPL1 reduces PTEN’s PIP3 phosphatase activity in the cell. (A) 0, 5...
(A) 0, 5, 10, 20 μg SIPL1 and 5 μg PTEN (S0P5, S5P5, S10P5 and S20P5, where S represents SIPL1) were cotransfected into 293T cells. 5 μg PTEN C124S mutant (C124S) served as a negative control. IPs were carried out using an anti-PTEN antibody, followed by assaying for PTEN’s PIP3 phosphatase activity. S20P0, 20 μg SIPL1 (FLAG-tagged) was used (negative control). Other negative controls (a reaction without the addition of PTEN and a reaction without the addition of PIP3 substrate) showed no detectable activities (data not shown). Western blot shows the immunoprecipitated PTEN and SIPL1. (B) DU145 cells were treated with control siRNA or SIPL1 siRNA retrovirus as indicated. Knockdown of SIPL1 was demonstrated (right panel). PTEN was immunoprecipitated from both cells, followed by assaying for PTEN’s PIP3 phosphatase activity in vitro. Comparable levels of PTEN were immunoprecipitated from both cell lines (data not shown). Experiments were repeated 3 times. (C) PTEN (5 μg) was transiently coexpressed in 293T cells with the indicated SIPL1 mutants (20 μg), followed by IP with anti-PTEN or M2 anti-FLAG (for SIPL1 mutants), and then assayed for PTEN’s PIP3 phosphatase activity. PTEN C124S (C124S, 5 μg) was used as a negative control. Under this assay system, 20 μg of the SIPL1 construct completely inhibited the PIP3 phosphatase activity produced by cotransfection of cells with 5 μg of the PTEN construct (data not shown; also see A).