The C-Terminal Domain from S. cerevisiae Pat1 Displays Two Conserved Regions Involved in Decapping Factor Recruitment

Zaineb Fourati,Dominique Durand,Olga Kolesnikova,Jenny Keller,Herman Van Tilbeurgh,Noureddine Lazar,Clément Charenton,Valerio Taverniti,Marc Graille,Claudine Gaudon Plesse,Régis Back,Bertrand Séraphin

doi:10.1371/journal.pone.0096828

Abstract

Eukaryotic mRNA decay is a highly regulated process allowing cells to rapidly modulate protein production in response to internal and environmental cues. Mature translatable eukaryotic mRNAs are protected from fast and uncontrolled degradation in the cytoplasm by two cis-acting stability determinants: a methylguanosine (m7G) cap and a poly(A) tail at their 5′ and 3′ extremities, respectively. The hydrolysis of the m7G cap structure, known as decapping, is performed by the complex composed of the Dcp2 catalytic subunit and its partner Dcp1. The Dcp1-Dcp2 decapping complex has a low intrinsic activity and requires accessory factors to be fully active. Among these factors, Pat1 is considered to be a central scaffolding protein involved in Dcp2 activation but also in inhibition of translation initiation. Here, we present the structural and functional study of the C-terminal domain from S. cerevisiae Pat1 protein. We have identified two conserved and functionally important regions located at both extremities of the domain. The first region is involved in binding to Lsm1-7 complex. The second patch is specific for fungal proteins and is responsible for Pat1 interaction with Edc3. These observations support the plasticity of the protein interaction network involved in mRNA decay and show that evolution has extended the C-terminal alpha-helical domain from fungal Pat1 proteins to generate a new binding platform for protein partners.

Highlights

Protein synthesis in eukaryotic cells is finely tuned both by controlling mRNA levels and their translation
We have focused our attention on the S. cerevisiae Pat1 C domain (ScPat1C), which is required for decapping and involved in interaction with Lsm1-7 complex, Xrn1, Edc3, Dcp1 and Dcp2
Given the much lower resolution of the dataset collected from crystal form I SeMet protein crystals, this dataset was only used to obtain experimental electron density maps by the Single wavelength Anomalous Diffusion method (SAD) using the selenium anomalous signal but not for structure refinement

Summary

Introduction

Protein synthesis in eukaryotic cells is finely tuned both by controlling mRNA levels and their translation. The regulation of mRNA decay provides a rapid mechanism to adapt to changing conditions. In the 39R59 mRNA decay, the mRNA is degraded by the exosome, a multiprotein complex with both exoand endo-nucleolytic activities embedded within a single protein Dis3 [2,3,4,5,6]. This generates a 59 capped oligonucleotide byproduct (less than 10 base long) that is further degraded by the scavenger protein DcpS (Dcs in budding yeast) to release m7GMP [7]. In the 59R39 pathway, the 59 cap is eliminated by a critical and highly regulated step known as decapping, which is followed by the rapid digestion of the mRNA body from its 59 end, a process mediated by the exonuclease Xrn1 [8,9]

Methods

Results

Conclusion