Subcellular fractionation method to study endosomal trafficking of Kaposi's sarcoma-associated herpesvirus

Cell & Bioscience
Lia R WalkerShaw M Akula

Abstract

Virus entry involves multiple steps and is a highly orchestrated process on which successful infection collectively depends. Entry processes are commonly analyzed by monitoring internalized virus particles via Western blotting, polymerase chain reaction, and imaging techniques that allow scientist to track the intracellular location of the pathogen. Such studies have provided abundant direct evidence on how viruses interact with receptor molecules on the cell surface, induce cell signaling at the point of initial contact with the cell to facilitate internalization, and exploit existing endocytic mechanisms of the cell for their ultimate infectious agenda. However, there is dearth of knowledge in regards to trafficking of a virus via endosomes. Herein, we describe an optimized laboratory procedure to isolate individual organelles during different stages of endocytosis by performing subcellular fractionation. This methodology is established using Kaposi's sarcoma-associated herpesvirus (KSHV) infection of human foreskin fibroblast (HFF) cells as a model. With KSHV and other herpesviruses alike, envelope glycoproteins have been widely reported to physically engage target cell surface receptors, such as integrins, in interactions l...Continue Reading

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Citations

Jan 4, 2017·Journal of Controlled Release : Official Journal of the Controlled Release Society·Mihael Lazebnik, Daniel W Pack
Mar 30, 2018·The Journal of Biological Chemistry·Jung Mi LimRodney L Levine
Mar 30, 2019·American Journal of Physiology. Heart and Circulatory Physiology·Sumit KarParas K Mishra
Nov 17, 2016·Apoptosis : an International Journal on Programmed Cell Death·Yang LiBo Yu
Sep 22, 2018·Scientific Reports·Hosni A M HusseinShaw M Akula
Sep 9, 2017·The Journal of Biological Chemistry·Astrid RohrbeckIngo Just
Apr 14, 2021·Cellular Microbiology·Yuen-Yan ChangVirginie Stévenin

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Methods Mentioned

BETA
imaging techniques
electron
density gradient centrifugation
Assay
fluorescence imaging

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