Abstract
ABSTRACTComplex machinery is required to drive secretory cargo export from the endoplasmic reticulum (ER), which is an essential process in eukaryotic cells. In vertebrates, the MIA3 gene encodes two major forms of transport and Golgi organization protein 1 (TANGO1S and TANGO1L), which have previously been implicated in selective trafficking of procollagen. Using genome engineering of human cells, light microscopy, secretion assays, genomics and proteomics, we show that disruption of the longer form, TANGO1L, results in relatively minor defects in secretory pathway organization and function, including having limited impacts on procollagen secretion. In contrast, loss of both long and short forms results in major defects in cell organization and secretion. These include a failure to maintain the localization of ERGIC53 (also known as LMAN1) and SURF4 to the ER–Golgi intermediate compartment and dramatic changes to the ultrastructure of the ER–Golgi interface. Disruption of TANGO1 causes significant changes in early secretory pathway gene and protein expression, and impairs secretion not only of large proteins, but of all types of secretory cargo, including small soluble proteins. Our data support a general role for MIA3/TANGO1 in maintaining secretory pathway structure and function in vertebrate cells.
Highlights
The first membrane trafficking step for secretion is driven by assembly of the COPII coat complex onto the endoplasmic reticulum (ER) membrane
Validation of MIA3 disruption To investigate the relative contribution of TANGO1S and TANGO1L at the ER–Golgi interface, we generated knockout human cell lines using CRISPR-Cas9
Our data show that loss of TANGO1 expression results in dramatic morphological changes to the ER–Golgi interface coupled with significant changes in secretion
Summary
The first membrane trafficking step for secretion is driven by assembly of the COPII coat complex onto the endoplasmic reticulum (ER) membrane. In yeast and many other eukaryotes, this results in COPII vesicles that bud from the ER membrane (Bednarek et al, 1995) This process can be reconstituted in vitro using synthetic liposomes and a minimal COPII machinery comprising the small GTP-binding protein Sar1p, which, in its. GTP-bound form, recruits the inner coat of Sec23p–Sec24p, and subsequently the outer coat of Sec13p–Sec31p (Matsuoka et al, 1998) Together, these proteins are sufficient to generate 60–80 nm vesicles in an energy-dependent manner. In the most commonly accepted models, these vesicles coalesce to form an ER–Golgi intermediate compartment (ERGIC; Schweizer et al, 1988). These structures form ER exit sites (ERES; Hughes et al, 2009). While they have been detected (Martinez-Menarguez et al, 1999), they are not abundant
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
