Saudi Arabian SARS-CoV-2 genomes implicate a mutant Nucleocapsid protein in modulating host interactions and increased viral load in COVID-19 patients

Monitoring SARS-CoV-2 spread and evolution through genome sequencing is essential in handling the COVID-19 pandemic. The availability of patient hospital records is crucial for linking the genomic sequence information to virus function during the course of infections. Here, we sequenced 892 SARS-CoV-2 genomes collected from patients in Saudi Arabia from March to August 2020. From the assembled sequences, we estimate the SARS-CoV-2 effective population size and infection rate and outline the epidemiological dynamics of import and transmission events during this period in Saudi Arabia. We show that two consecutive mutations (R203K/G204R) in the SARS-CoV-2 nucleocapsid (N) protein are associated with higher viral loads in COVID-19 patients. Our comparative biochemical analysis reveals that the mutant N protein displays enhanced viral RNA binding and differential interaction with key host proteins. We found hyper-phosphorylation of the adjacent serine site (S206) in the mutant N protein by mass-spectrometry analysis. Furthermore, analysis of the host cell transcriptome suggests that the mutant N protein results in dysregulated interferon response genes. We provide crucial information in linking the R203K/G204R mutations in the N protein as a major modulator of host-virus interactions and increased viral load and underline the potential of the nucleocapsid protein as a drug target during infection.

Cytosine deamination in SARS-CoV-2 leads to progressive CpG depletion.

RNA viruses use CpG reduction to evade the host cell defense, but the driving mechanisms are still largely unknown. In an attempt to address this we used a rapidly growing genomic dataset of SARS-CoV-2 with relevant metadata information. Remarkably, by simply ordering SARS-CoV-2 genomes by their date of collection, we find a progressive increase of C-to-U substitutions resulting in 5'-UCG-3' motif reduction that in turn have reduced the CpG frequency over just a few months of observation. This is consistent with APOBEC-mediated RNA editing resulting in CpG reduction, thus allowing the virus to escape ZAP-mediated RNA degradation. Our results thus link the dynamics of target sequences in the viral genome for two known host molecular defense mechanisms, mediated by the APOBEC and ZAP proteins.

The genomic variation landscape of globally-circulating clades of SARS-CoV-2 defines a genetic barcoding scheme

We describe fifteen major mutation events from 2,058 high-quality SARS-CoV-2 genomes deposited up to March 31st, 2020. These events define five major clades (G, I, S, D and V) of globally-circulating viral populations, representing 85.7% of all sequenced cases, which we can identify using a 10 nucleotide genetic classifier or barcode. We applied this barcode to 4,000 additional genomes deposited between March 31st and April 15th and classified successfully 95.6% of the clades demonstrating the utility of this approach. An analysis of amino acid variation in SARS-CoV-2 ORFs provided evidence of substitution events in the viral proteins involved in both host-entry and genome replication. The systematic monitoring of dynamic changes in the SARS-CoV-2 genomes of circulating virus populations over time can guide therapeutic and prophylactic strategies to manage and contain the virus and, also, with available efficacious antivirals and vaccines, aid in the monitoring of circulating genetic diversity as we proceed towards elimination of the agent. The barcode will add the necessary genetic resolution to facilitate tracking and monitoring of infection clusters to distinguish imported and indigenous cases and thereby aid public health measures seeking to interrupt transmission chains without the requirement for real-time complete genomes sequencing.