Structure of the Unphosphorylated STAT5a Dimer*

Edith Pfitzner,Sophie Verrier,Klaus Klumpp,Kenneth Straub,Ana Mirela Neculai,Dante Neculai,Stefan Becker

doi:10.1074/jbc.m507682200

Abstract

STAT proteins have the function of signaling from the cell membrane into the nucleus, where they regulate gene transcription. Latent mammalian STAT proteins can form dimers in the cytoplasm even before receptor-mediated activation by specific tyrosine phosphorylation. Here we describe the 3.21-A crystal structure of an unphosphorylated STAT5a homodimer lacking the N-terminal domain as well as the C-terminal transactivation domain. The overall structure of this fragment is very similar to phosphorylated STATs. However, important differences exist in the dimerization mode. Although the interface between phosphorylated STATs is mediated by their Src-homology 2 domains, the unphosphorylated STAT5a fragment dimerizes in a completely different manner via interactions between their beta-barrel and four-helix bundle domains. The STAT4 N-terminal domain dimer can be docked onto this STAT5a core fragment dimer based on shape and charge complementarities. The separation of the dimeric arrangement, taking place upon activation and nuclear translocation of STAT5a, is demonstrated by fluorescence resonance energy transfer experiments in living cells.

Highlights

Overall Structure—The core fragment of unphosphorylated STAT5a used for crystallization was chosen from an alignment with the STAT3 fragment that had been crystallized in complex with DNA [4]
Each monomer bears the general architecture already found in the phosphorylated STAT core fragments (Fig. 1A): an N-terminal four-helix bundle, an eight-stranded ␤-barrel, an ␣-helical linker domain, and a SH2 domain
The residues 128 –137, 423– 432, and 691–712 are not ordered, and they are not included in our model

Summary

Introduction

In this structure the N-terminal domains seem to stabilize the interaction between the core fragments, well in agreement with these biochemical and cell biological data. The connecting region between the N-terminal domain and the core fragment is not visible in the STAT1 structure, resulting in two possible dimerization modes of the two proteins because of their tetrameric arrangement in the crystal.

Results

Conclusion