Comparative interactomic analysis of MERS-CoV & SARS-CoV-2 proteins in human, bat, and camel cells

Abstract

The Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are Betacoronaviruses capable of causing fatal human infections. Both viruses are believed to have emerged from bats via an intermediate host (camels for MERS-CoV, unknown for SARS-CoV-2) into the human population. MERSCoV and SARS-CoV-2 are enveloped positive-sense RNA viruses which encode four structural proteins (envelope (E), nucleocapsid, spike, and membrane (M)). The E and M proteins are involved in virus assembly, budding, envelope formation, and pathogenesis. Finding cellular protein interactors for these viral proteins, conserved across species, will increase our understanding of the coronavirus lifecycle and identify targets for antiviral development. Initial validation of the results for 11 cellular proteins interacting with the MERS-CoV E and/or M proteins by co-immunoprecipitation (co-IP) /Western blot analysis and immunofluorescence co-localisation, confirmed 10 of the interactions. Three cell lines (human HEK293, bat Pteropus alecto PaKiT and Camelus dromedarius Dubca) were used for transient expression of the MERS-CoV and SARS-CoV-2 E and M proteins (FLAG epitope-tagged) followed by co-IP and high-throughput mass spectrometrybased interactomic analysis. Bioinformatic analysis revealed E/M protein interactions with ER, Golgi, mitochondrial and nuclear proteins. There were 32 high-confidence cellular interaction proteins conserved amongst the different cell lines and viruses (p < 0.05, > 0 log 2 fold change compared to the controls). To determine the importance of the 11 cellular proteins interacting with the MERS-CoV E and/or M proteins and 32 cellular proteins conserved across species, in the virus lifecycle, functional validation was done by siRNA depletion in human cells, followed by infection with SARS-CoV-2. An interesting four cellular proteins were shown to be important for SARS-CoV- 2 replication. These interesting proteins include (CERS2, LPCAT1, UBA52, and TM9SF2) that when it was knocked down, succeeded in reducing the SARS-CoV-2 replication. This can be followed by working to find if these proteins can be safely targeted by a drug to be reduced in cells.