The Novel PIAS-like Protein hZimp10 Enhances Smad Transcriptional Activity

Xiaomeng Li,Gregory Thyssen,Jason Beliakoff,Zijie Sun

doi:10.1074/jbc.m508365200

Abstract

Transforming growth factor beta (TGF-beta) plays critical roles in the control of cell proliferation, differentiation, and apoptosis. Smad proteins are substrates of the TGF-beta type I receptor and are responsible for transducing receptor signals to target genes in the nucleus. The PIAS (protein inhibitor of activated STAT) proteins were originally identified as transcriptional co-regulators of the JAK-STAT pathway. Subsequently, cross-talk between the PIAS proteins and other signaling pathways has been shown to be involved in various cellular processes. Importantly, PIAS proteins modulate TGF-beta signaling by regulating the transcriptional activity of Smad3. In this study we tested whether hZimp10, a novel PIAS-like protein, acts as other PIAS proteins to regulate Smad3-mediated transcription. We show that expression of exogenous hZimp10 enhances the transcriptional activity of Smad3, which appears to be Smad4-dependent and responsive to TGF-beta induction. Furthermore, knockdown of endogenous hZimp10 reduced the transcriptional activity of Smad3. A protein-protein interaction between Smad3 and Smad4 with hZimp10 was identified in glutathione S-transferase-pulldown and co-immunoprecipitation assays. The Miz domain of hZimp10 and the MH2 domains of Smad3 and Smad4 were mapped as the regions responsible for binding. Results from immunostaining assays further demonstrated that Smad3, Smad4, and hZimp10 co-localize within cell nuclei. Finally, we demonstrated that Smad3/4-mediated transcription is significantly impaired in response to TGF-beta induction in Zimp10 null (zimp10-/-) embryonic fibroblasts. Taken together, these results provide the first line of evidence to demonstrate a role for Zimp10 in regulating the TGF-beta/Smad signaling pathway.

Highlights

Tors activate signaling by binding and bringing together members of two subfamilies of transmembrane protein serine/ threonine kinases, the type I (T␤R-I) and type II receptors (T␤R-II)
Smad proteins are the substrates of TGF-␤ type I receptor and play a central role in transducing receptor signals to target genes in the nucleus [2]
Sequence analysis has shown that the SUMO E3 ligase RING domain shares significant homology with the Miz domain of PIAS proteins [24]

Summary

Introduction

Tors activate signaling by binding and bringing together members of two subfamilies of transmembrane protein serine/ threonine kinases, the type I (T␤R-I) and type II receptors (T␤R-II). Using several in vivo and in vitro approaches, we demonstrated that Zimp10 interacts with Smad3/4 proteins and augments Smad-mediated transcription, which provides the first line of evidence that Zimp10 plays a critical role in the regulation of the TGF-␤/Smad signaling pathway. An ϳ5-fold induction of Smad3-mediated transcriptional activity was observed when cells were transfected with Smad3 (Fig. 1A).

Results

Conclusion