Structural Insight into Golgi Membrane Stacking by GRASP65 and GRASP55 Proteins

Yanbin Feng,Wenying Yu,Xinxin Li,Shaoyu Lin,Ying Zhou,Junjie Hu,Xinqi Liu

doi:10.1074/jbc.m113.478024

Yanbin Feng, Wenying Yu + Show 5 more

Open Access

https://doi.org/10.1074/jbc.m113.478024

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Abstract

The stacking of Golgi cisternae involves GRASP65 and GRASP55. The oligomerization of the N-terminal GRASP domain of these proteins, which consists of two tandem PDZ domains, is required to tether the Golgi membranes. However, the molecular basis for GRASP assembly is unclear. Here, we determined the crystal structures of the GRASP domain of GRASP65 and GRASP55. The structures reveal similar homotypic interactions: the GRASP domain forms a dimer in which the peptide-binding pockets of the two neighboring PDZ2 domains face each other, and the dimers are further connected by the C-terminal tail of one GRASP domain inserting into the binding pocket of the PDZ1 domain in another dimer. Biochemical analysis suggests that both types of contacts are relatively weak but are needed in combination for GRASP-mediated Golgi stacking. Our results unveil a novel mode of membrane tethering by GRASP proteins and provide insight into the mechanism of Golgi stacking.

Highlights

The oligomerization of GRASP65 and GRASP55 is required to tether Golgi membranes
A 2.2-Å dataset was collected, and the structure was determined by molecular replacement (Table 1), but only residues 12–210 were visible in the electron density
The dimers are linked through interactions between the two C-terminal tails (CTs) of one dimer and two peptide-binding pockets of the PDZ1 domains in the dimer (Fig. 1C). These results suggest that GRASP65 utilizes two distinct interfaces in the GRASP domain, both involving the peptide-binding pocket of the PDZ domain, to achieve oligomerization

Summary

Introduction

The oligomerization of GRASP65 and GRASP55 is required to tether Golgi membranes. Results: The crystal structures reveal two types of intermolecular interactions, and biochemical and cellular assays confirm these observations. Conclusion: Two relatively weak interactions in combination are needed for GRASP-mediated Golgi stacking. Significance: These data suggest a novel mode of Golgi membrane stacking by the GRASP proteins. The oligomerization of the N-terminal GRASP domain of these proteins, which consists of two tandem PDZ domains, is required to tether the Golgi membranes. Biochemical analysis suggests that both types of contacts are relatively weak but are needed in combination for GRASP-mediated Golgi stacking. Our results unveil a novel mode of membrane tethering by GRASP proteins and provide insight into the mechanism of Golgi stacking

Methods

Results

Conclusion