The p53-inducible TSAP6 gene product regulates apoptosis and the cell cycle and interacts with Nix and the Myt1 kinase

doi:10.1073/pnas.0530298100

The p53-inducible TSAP6 gene product regulates apoptosis and the cell cycle and interacts with Nix and the Myt1 kinase

^*Molecular Engines Laboratories, 20 Rue Bouvier, 75011 Paris, France; ^‡Fondation Jean Dausset–Centre d'Etude du Polymorphisme Humain, 27 Rue Juliette Dodu, 75010 Paris, France; and ^§Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot 76100, Israel

Contributed by Jean Dausset

Abstract

The p53 tumor suppressor protein plays a crucial role in tumorigenesis by controlling cell-cycle progression and apoptosis. We have previously described a transcript designated tumor suppressor activated pathway-6 (TSAP6) that is up-regulated in the p53-inducible cell line, LTR6. Cloning of the murine and human full-length TSAP6 cDNA revealed that it encodes a 488-aa protein with five to six transmembrane domains. This gene is the murine and human homologue of the recently published rat pHyde. Antibodies raised against murine and human TSAP6 recognize a 50- to 55-kDa band induced by p53. Analysis of the TSAP6 promoter identified a functional p53-responsive element. Functional studies demonstrated that TSAP6 antisense cDNA diminished levels of the 50- to 55-kDa protein and decreased significantly the levels of p53-induced apoptosis. Furthermore, TSAP6 small interfering RNA inhibited apoptosis in TSAP6-overexpressing cells. Yeast two-hybrid analysis followed by GST/in vitro-transcribed/translated pull-down assays and in vivo coimmunoprecipitations revealed that TSAP6 associated with Nix, a proapoptotic Bcl-2-related protein and the Myt1 kinase, a negative regulator of the G₂/M transition. Moreover, TSAP6 enhanced the susceptibility of cells to apoptosis and cooperated with Nix to exacerbate this effect. Cell-cycle studies indicated that TSAP6 could augment Myt1 activity. Overall, these data suggest that TSAP6 may act downstream to p53 to interface apoptosis and cell-cycle progression.

Footnotes

↵ † B.J.P., V.N.-P., and N.A. contributed equally to this work.
↵ ¶ To whom correspondence should be addressed. E-mail: atelerman{at}molecular-engines.com.
Data deposition: The sequences reported in this paper have been deposited in the GenBank database [accession nos. AY214462 (murine TSAP6) and AY214461 (human TSAP6)].
Abbreviations:

TSAP,

tumor suppressor activated pathway;

HA,

hemagglutinin;

Y2H,

yeast two-hybrid;

IVT,

in vitro-transcribed/translated;

siRNA,

small interfering RNA;

DTB,

double-thymidine block;

PARP,

poly(ADP-ribose) polymerase;

AIP,

apoptosis-linked gene-2-interacting protein 1