Superresolution imaging of viral protein trafficking

Abstract

The endoplasmic reticulum (ER) membrane is closely apposed to the outer mitochondrial membrane (OMM), which facilitates communication between these organelles. These contacts, known as mitochondria-associated membranes (MAM), facilitate calcium signaling, lipid transfer, as well as antiviral and stress responses. How cellular proteins traffic to the MAM, are distributed therein, and interact with ER and mitochondrial proteins are subject of great interest. The human cytomegalovirus UL37 exon 1 protein or viral mitochondria-localized inhibitor of apoptosis (vMIA) is crucial for viral growth. Upon synthesis at the ER, vMIA traffics to the MAM and OMM, where it reprograms the organization and function of these compartments. vMIA significantly changes the abundance of cellular proteins at the MAM and OMM, including proteins that regulate calcium homeostasis and cell death. Through the use of superresolution imaging, we have shown that vMIA is distributed at the OMM in nanometer scale clusters. This is similar to the clusters reported for the mitochondrial calcium channel, VDAC, as well as electron transport chain, translocase of the OMM complex, and mitochondrial inner membrane organizing system components. Thus, aside from addressing how vMIA targets the MAM and regulates survival of infected cells, biochemical studies and superresolution imaging of vMIA offer insights into the formation, organization, and functioning of MAM. Here, we discuss these insights into trafficking, function, and organization of vMIA at the MAM and OMM and discuss how the use of superresolution imaging is contributing to the study of the formation and trafficking of viruses.

Superresolution Imaging Identifies That Conventional Trafficking Pathways Are Not Essential for Endoplasmic Reticulum to Outer Mitochondrial Membrane Protein Transport

“…Using superresolution imaging by multifocal structured illumination microscopy (MSIM), gated stimulated emission depletion (gSTED) and photoactivated localization microscopy (PALM), we established that vMIA forms clusters of ~100–150 nm at the OMM of human cells 13,36 . Clustering has also been reported for inner mitochondrial membrane (IMM) proteins including mitochondrial inner membrane or cristae organizing system (MICOS or MINOS), and cytochrome C oxidase subunit 2 of complex IV 35,37 .…”

Isolation of Endoplasmic Reticulum, Mitochondria, and Mitochondria‐Associated Membrane and Detergent Resistant Membrane Fractions from Transfected Cells and from Human Cytomegalovirus‐Infected Primary Fibroblasts

“…The authors found that HCMV UL37 proteins traffic sequentially from the ER into the mitochondrial outer membrane (Bhuvanendran et al, 2014; Colberg-Poley et al, 2015; Mavinakere et al, 2006). Characterization of the subcellular localization and trafficking of HCMV UL37 proteins required establishing a protocol in which ER and mitochondria could be reliably separated from one another, despite their prevalent interconnection.…”

Metabolic signaling functions of ER–mitochondria contact sites: role in metabolic diseases

“…Concerning viral infections, several viral proteins were shown to localize to MAM (for reviews, see Williamson & Colberg-Poley 2009, Colberg-Poley et al 2015, de Armas-Rillo et al 2016. This is the case for the human cytomegalovirus exon 1 protein and viral mitochondrialocalized inhibitor of apoptosis (Bozidis et al 2010, Zhang et al 2011, for the hepatitis C virus core protein (Williamson & Colberg-Poley 2009, Horner et al 2011, 2015, for the human immunodeficiency virus protein-R (Huang et al 2012) and for the dengue virus (Chatel-Chaix et al 2016).…”