SARS-CoV-2 viral budding and entry can be modeled using BSL-2 level virus-like particles

J Biol Chem. 2021 Jan-Jun;296:100103. doi: 10.1074/jbc.RA120.016148. Epub 2020 Nov 27.

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first discovered in December 2019 in Wuhan, China, and expeditiously spread across the globe causing a global pandemic. Research on SARS-CoV-2, as well as the closely related SARS-CoV-1 and MERS coronaviruses, is restricted to BSL-3 facilities. Such BSL-3 classification makes SARS-CoV-2 research inaccessible to the majority of functioning research laboratories in the United States; this becomes problematic when the collective scientific effort needs to be focused on such in the face of a pandemic. However, a minimal system capable of recapitulating different steps of the viral life cycle without using the virus' genetic material could increase accessibility. In this work, we assessed the four structural proteins from SARS-CoV-2 for their ability to form virus-like particles (VLPs) from human cells to form a competent system for BSL-2 studies of SARS-CoV-2. Herein, we provide methods and resources of producing, purifying, fluorescently and APEX2-labeling of SARS-CoV-2 VLPs for the evaluation of mechanisms of viral budding and entry as well as assessment of drug inhibitors under BSL-2 conditions. These systems should be useful to those looking to circumvent BSL-3 work with SARS-CoV-2 yet study the mechanisms by which SARS-CoV-2 enters and exits human cells.

Keywords: Golgi; SARS-CoV-2; coronavirus; electron microscopy (EM); membrane bilayer; viral protein; virology; virus; virus assembly; virus budding; virus entry; virus-like particle (VLP).

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Biomimetic Materials / chemistry
  • Biomimetic Materials / metabolism
  • Containment of Biohazards / classification
  • Coronavirus Envelope Proteins / genetics*
  • Coronavirus Envelope Proteins / metabolism
  • Gene Expression
  • Genes, Reporter
  • Government Regulation
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HEK293 Cells
  • Humans
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Microscopy, Electron
  • Nucleocapsid Proteins / genetics*
  • Nucleocapsid Proteins / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • SARS-CoV-2 / genetics
  • SARS-CoV-2 / growth & development*
  • SARS-CoV-2 / metabolism
  • SARS-CoV-2 / ultrastructure
  • Spike Glycoprotein, Coronavirus / genetics*
  • Spike Glycoprotein, Coronavirus / metabolism
  • Viral Matrix Proteins / genetics*
  • Viral Matrix Proteins / metabolism
  • Virion / genetics
  • Virion / growth & development*
  • Virion / metabolism
  • Virion / ultrastructure
  • Virus Assembly / physiology
  • Virus Internalization
  • Virus Release / physiology

Substances

  • Coronavirus Envelope Proteins
  • Luminescent Proteins
  • Nucleocapsid Proteins
  • Recombinant Proteins
  • Spike Glycoprotein, Coronavirus
  • Viral Matrix Proteins
  • envelope protein, SARS-CoV-2
  • membrane protein, SARS-CoV-2
  • red fluorescent protein
  • spike protein, SARS-CoV-2
  • Green Fluorescent Proteins