The length of cargo-protein transmembrane segments drives secretory transport by facilitating cargo concentration in export domains

Abstract

The cellular destination of secretory proteins is determined by interactions of their targeting motifs with coat-protein complexes. The transmembrane domain (TMD) of secretory proteins also plays a central role in their transport and targeting. However, a comprehensive model that considers both TMD- and targeting-sequence-mediated transport has never been advanced. We focused on the secretory transport of two fluorescently tagged membrane proteins: vesicular stomatitis virus G tsO45 (VSVG), which is a cargo protein that is a thermoreversible mutant, and the Golgi-resident protein GalT-CFP. A quantitative approach was applied to analyze, in living cells, secretory transport dynamics, as well as cargo concentration of YFP-tagged VSVG mutants with one, three, five, seven, eight or nine amino acids deleted from their TMD, as well as two or four amino acids added to their TMD. Changes in TMD length affected secretory transport dynamics and the extent of cargo concentration in the ER exit sites, demonstrating that the capacity of the transport machinery to concentrate cargo depends on the length of the TMD of the cargo protein.

Getting membrane proteins on and off the shuttle bus between the endoplasmic reticulum and the Golgi complex

“…More recently, similar observations have been made for the secretory membrane protein vesicular stomatitis virus glycoprotein (VSVG) (Dukhovny et al, 2009).…”

“…Indeed, Rer1p, a cargo receptor involved in retrograde transport, recognizes the TMD of escaped membrane proteins of the ER (Sato et al, 2003), and the artificially lengthened TMD of cytochrome P450 2C1 (see Box 2) interferes with retrieval of the altered protein from the Golgi (Szczesna-Skorupa and Kemper, 2000). However, for a number of the proteins listed in Box 2, the short TMD prevents their entry into ERES, a phenomenon known as static retention; by contrast, longer TMDs are recruited to sites of vesicle budding (Ronchi et al, 2008;Dukhovny et al, 2009;Vander Heyden et al, 2011). Below, I discuss some possible mechanisms underlying this TMD-dependent sorting within the ER.…”

Role for TBC1D20 and Rab1 in Hepatitis C Virus Replication via Interaction with Lipid Droplet-Bound Nonstructural Protein 5A

“…Dominant negative N121I and constitutively active Q67L mutations were introduced using the QuikChange site-directed mutagenesis kit. VSVGtsO45-AIA, a DXE motif-deficient mutant which is retained in the ER, has been previously described (12), and cDNA encoding this mutant was subcloned into pmCherry-C1 (Clontech). HCV genotype 1b core cDNA was amplified from Bart79I and cloned into pmCherry-C1 using the XhoI and SacII restriction sites.…”

The MARVEL transmembrane motif of occludin mediates oligomerization and targeting to the basolateral surface in epithelia

“…In light of the findings on MAL, we speculated that the MARVEL domain of Ocln may exhibit comparable characteristics that could be essential for Ocln targeting or function. Based on the hypothesis that sorting and transport of membrane proteins are propelled at least in part by hydrophobic mismatch-mediated protein clustering (Dukhovny et al, 2009;Schmidt and Weiss, 2010), we tested whether the Ocln MARVEL domain is associated with targeting to the basolateral membrane pole and TJs.…”