Clathrin Adaptor Complex-interacting Protein Irc6 Functions through the Conserved C-Terminal Domain

Huajun Zhou,Gregory S Payne,Giancarlo Costaguta

doi:10.1038/s41598-019-40852-8

Huajun Zhou, Gregory S Payne + Show 1 more

Open Access

https://doi.org/10.1038/s41598-019-40852-8

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Abstract

Clathrin coats drive transport vesicle formation from the plasma membrane and in pathways between the trans-Golgi network (TGN) and endosomes. Clathrin adaptors play central roles orchestrating assembly of clathrin coats. The yeast clathrin adaptor-interacting protein Irc6 is an orthologue of human p34, which is mutated in the inherited skin disorder punctate palmoplantar keratoderma type I. Irc6 and p34 bind to clathrin adaptor complexes AP-1 and AP-2 and are members of a conserved family characterized by a two-domain architecture. Irc6 is required for AP-1-dependent transport between the TGN and endosomes in yeast. Here we present evidence that the C-terminal two amino acids of Irc6 are required for AP-1 binding and transport function. Additionally, like the C-terminal domain, the N-terminal domain when overexpressed partially restores AP-1-mediated transport in cells lacking full-length Irc6. These findings support a functional role for Irc6 binding to AP-1. Negative genetic interactions with irc6∆ are enriched for genes related to membrane traffic and nuclear processes, consistent with diverse cellular roles for Irc6.

Highlights

Clathrin-coated vesicles mediate transport from the plasma membrane and between the trans-Golgi network (TGN) and endosomes
Dependent TGN-endosome transport, we used an assay for sensitivity to the chitin-binding dye, calcofluor white[1] (CCFW; Fig. 1a)
Growth inhibition of chs6∆ cells by calcofluor white1 (CCFW) can serve as a sensitive measure of defects in AP-1/Irc6-dependent localization of Chs[3]

Summary

Introduction

Clathrin-coated vesicles (ccv) mediate transport from the plasma membrane and between the trans-Golgi network (TGN) and endosomes. The adaptor complex-interacting protein Irc[6] is a member of a conserved protein family defined by an N-terminal G protein-like domain and a C-terminal adaptin-binding domain[1,2]. Yeast cells lacking Irc[6] (irc6∆) are defective in AP-1-dependent transport between the TGN and endosomes, but AP-2-mediated endocytosis appears normal[1,2]. TGN-endosome traffic can be restored in irc6∆ cells by overexpressing p34, providing evidence for evolutionarily conserved function in this pathway[1]. Another interaction partner of Irc[6], the Rab GTPase Ypt[31], acts in TGN-endosome transport. A novel conserved BC-YY motif (beta-strand connecting YY motif), located in an accessible loop connecting beta strands in the N-terminal domain, contributes to Irc[6] and p34 function in yeast

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