COVID Virologist Aspects, Life-cycle and Targets for Drug Development
CoVs have a long, capped and poly-adenylated RNA genome, which contains between 8 to 10 open-reading-frames (ORFs), allowing structural, non-structural and accessory viral protein synthesis [87]. SARS-CoV2 is 29,903 base-long and contains 6 majors ORFs, as well as additional accessory genes; the reference sequence is registered in GenBank with ID: MN908947.3 [1]. (A and B Up to 28 different polypeptides are potentially produced in fine from the different ORFs and after polyprotein processing by viro-encoded proteases [87]. If the RNA genome contained in virions can already serve, after cell entry, as a template for the synthesis of non-structural proteins, which are involved in the early phase of virus replication (mainly by forming the replicase complex), subgenomic messenger RNAs (sgmRNAs) are also produced in the late phase of the cycle to allow the synthesis of structural proteins (e.g. spike (S), envelope (E), membrane (M) and nucleocapsid proteins), as well as other accessory polypeptides.
Another main replication intermediate is the complementary minus-sens RNA, which is used by the viro-encoded RNA-dependent RNA-polymerase (RdRp), within replicase complex, to amplify the full-length genome, which is then capped and polyadenylated by both viral and host enzymes before being incorporated into virus progeny. (B) After entry into ACE2-positive (entry receptor) and TMPRSS2-positive (co-factor for entry) cells, and the membrane fusion (i.e. uncoating process), a full-length genome is released in the cytoplasm of cells. This full-length polycistronic RNA is directly used to efficiently encode a polyprotein from the first ORFs present on the molecule, starting from 5’ extremity, i.e. ORF1a and ORF1b; the latter is read after a frame-shift from ribosomal scanning of ORF1a. (A and B The polyprotein is then processed by two viro-encoded proteases, a papain-like cysteine protease (PLpro/Nsp3) and a chemotrypsin-like protease (3CLpro/Nsp5; also known as main protease (Mpro)), into 16 proteins/polypeptides (Nsp1 to 16). (B) These non-structural proteins/polypeptides are important for the early stage of infection, as they allow in particular the formation of the replicase complex around the RdRp enzymatic activity, which is involved in the synthesis of negative-sense full-length RNA, as well as sgmRNAs by a discontinuous transcription strategy [87]. The latters enable an efficient and stochiometric synthesis of all other viral proteins/polypeptides, which are important for virus assembly and release of progeny virions.
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