SARS-CoV-2 Orf1ab Genome Mutations, the Driving Force for Virus Pathogenicity.
Authors: Cobar Oscar , Cóbar Stella 
Published: December 2023

Background: The proteins codified in the Open Reading Frame 1ab -Orf1ab- region of the SARS-CoV-2 genome are the main responsible of the virus transcription, replication, and translation processes inside the human cell. Once inside the cell, the viral mRNA encode structural and nonstructural proteins, that direct virus assembly, transcription, replication and host control, and the accessory proteins whose function has not been determined. The largest gene, Orf1ab, contains overlapping open reading frames that encode polyproteins PP1ab and PP1a. These polyproteins are cleaved to yield 16 nonstructural proteins, NSP1-16. Production of the longer (PP1ab) or shorter protein (PP1a) depends on a ribosomal frameshifting event. The proteins, include the papain-like proteinase (NSP3), 3C-like proteinase (NSP5), RNA-dependent RNA polymerase (NSP12, RdRp), helicase (NSP13, HEL), endoRNAse (NSP15), 2'-O-Ribose-Methyltransferase (NSP16) and other nonstructural proteins. The SARS-CoV-2 nonstructural proteins are responsible for viral transcription, replication, proteolytic processing, suppression of host immune responses, and suppression of host gene expression. The purpose of the manuscript is to present a systematic review as of September 30, 2023, on the Orf1ab region mutations of the SARS-CoV-2 genome as of September 30, 2023, with the aim to predict, through the mutations profile on that region, the severity of an infection for a new SARS-CoV-2 variant that could emerge in the near future. Material and Methods: Original scientific articles published in Medline, Pubmed, Science Direct, Web of Science, Scopus, EBSCO and BioMed Central databases, official health organizations (WHO, CDC, ECDEC, NIH) electronic publications, and specialized media in the subject, were electronically searched to accomplish the aim of the study. Articles published in any language were included from 2020 to present using a variety of keywords in combination. The studies relevant to our review were analysed and compared. Results and discussion: The NIH “National Human Genome Research Institute” define the Open Reading Frames (ORFs) as a portion of a DNA sequence that does not include a stop codon. The Open Reading Frames encode accessory proteins transcribed from the 3′ one-third of the genome to form a set of subgenomic mRNAs (sg mRNAs). Following entry on the human cell, the viral particle release the genomic RNA molecule that is translated on two large open reading frames, ORF1a and ORF1b. The resulting polyproteins pp1a and pp1ab are co-translationally and post-translationally processed into the individual non-structural proteins (NSPs) that form the viral replication and transcription complex (RTC). Translated structural proteins translocate into endoplasmic reticulum (ER) membranes and transit through the ER-to-Golgi intermediate compartment (ERGIC), where interaction with N-encapsidated, newly produced genomic RNA results in budding into the lumen of secretory vesicular compartments. Two viral proteases, Plpro -NSP3- and 3Cl-pro -NSP5-, process the polyproteins and generate the nonstructural proteins NSP1-NSP16 that directs the transcription, replication and construction of new virions that are secreted by exocytosis from the infected cell. The evolution of key proteins in viral transcription and replication is clearly observed by carefully studying the structure, function, and evolution of RdRp, Mpro or 3Clpro, and NSP13 proteins directed by the Orf1a and Orf1ab genome mutations. Conclusions: ORF1ab is cleaved into 16 non-structural proteins involved in SARS-CoV-2 transcription and genome replication. P323L, P227L, G671S, V776L and A185S are the first five frequent mutations of RdRp (NSP12), the mutations P227L and G671S might have functional consequences in the viral transcription and replication. Mutations in residues D499 to L514, K545, R555, T611 to M626, G678 to T710, S759 to D761 are directly implicated with the transcription-replication capability of the virus by RdRp. In Mpro (NSP5) the mutation of residues H41, P132, C145, S145, L226, T234, R298, S301, F305, and Q306 may increase the efficiency of proteolytic cleavage of proteins such as NEMO, thereby improving the ability of the omicron series of viruses to suppress the immune system and accelerate the viral replication. In Helicase (NSP13) the mutations of residues E261, K218, K288, S289, H290, D374, E375, Q404, K460, R567, and A598 are involved in the separation of the double-stranded RNA or DNA with a 5′→3′ polarity as well as 5′ mRNA capping activity in the virus transcription-replication process. In the Orf1ab gene, ORF1b:V2354F mutation, corresponding to NSP15:V303F, may induce a conformational change and result in a disruption to a flanking beta-sheet structure. The premature stop codon ORF7a:Q94*, truncates the transmembrane protein and cytosolic tail used to mediate protein transport, may affect protein localization to the ER-Golgi. The analyses of Orf1ab genome mutations, allows us to predict, through the mutations profile on that region, the severity of an infection for a new SARS-CoV-2 variant that could emerge in the near future.