Induction of endoplasmic reticulum (ER)-to-Golgi blockade or ER stress induces Golgi reassembly stacking protein (GRASP)-mediated, Golgi-independent unconventional cell-surface trafficking of the folding-deficient ΔF508-cystic fibrosis transmembrane conductance regulator (CFTR). However, molecular mechanisms underlying this process remain elusive. Here, we show that phosphorylation-dependent dissociation of GRASP homotypic complexes and subsequent relocalization of GRASP to the ER play a critical role in the unconventional secretion of CFTR. Immunolocalization analyses of mammalian cells revealed that the Golgi protein GRASP55 was redistributed to the ER by stimuli that induce unconventional secretion of ΔF508-CFTR, such as induction of ER-to-Golgi blockade by the Arf1 mutant. Notably, the same stimuli also induced phosphorylation of regions near the C-terminus of GRASP55 and dissociation of GRASP homomultimer complexes. Furthermore, phosphorylation-mimicking mutations of GRASP55 induced the monomerization and ER relocalization of GRASP55, and these changes were nullified by phosphorylation-inhibiting mutations. These results provide mechanistic insights into how GRASP accesses the ER-retained ΔF508-CFTR and mediates the ER stress-induced unconventional secretion pathway.
Bibliographical noteFunding Information:
We thank Dong Soo Chang for the assistance with illustrations and the Yonsei-Carl Zeiss Advanced Imaging Center for technical assistance. This work was supported by grants 2013R1A3A2042197 and 2007?0056092 from the National Research Foundation, the Ministry of Science, ICT & Future Planning, Republic of Korea. J. K., S. H. N., H. P., D. H. K. and W. Y. C. performed molecular and immunofluorescence experiments and analyzed the data. J. S. C. and K. K. performed the surface plasmon resonance experiments. H.-S. C. and J. Y. K. contributed to data analysis. J. K., S. H. N. and M. G. L. planned the study, contributed to analysis of the data and wrote the paper. All authors discussed the results and commented on the manuscript. The authors declare no competing financial interests.
All Science Journal Classification (ASJC) codes
- Structural Biology
- Molecular Biology
- Cell Biology