The Highly Conserved Stem-Loop II Motif Is Dispensable for SARS-CoV-2

Area of Science:

• Virology
• Molecular Biology
• RNA Structure

Background:

• The stem-loop II motif (s2m) is an RNA structural element in the 3' untranslated region (UTR) of SARS-CoV-2.
• Its functional significance in viral replication and pathogenesis remains largely unknown despite its discovery over 25 years ago.

Purpose of the Study:

• To investigate the functional importance of the s2m motif in SARS-CoV-2.
• To determine the impact of s2m deletion or mutation on viral growth and fitness.

Main Methods:

• Reverse genetics was used to create SARS-CoV-2 variants with s2m deletions or mutations.
• Viral growth was assessed in vitro (tissue culture) and in vivo (Syrian hamsters).
• RNA secondary structure analysis was performed using SHAPE-MaP and DMS-MaPseq.

Main Results:

• Deletion or mutation of the s2m motif had no discernible effect on SARS-CoV-2 replication in vitro.
• Viral growth and fitness in Syrian hamsters were unaffected by s2m alterations.
• RNA structure analysis confirmed that s2m forms an independent structure and its deletion does not disrupt the overall 3'-UTR RNA structure.

Conclusions:

• The stem-loop II motif (s2m) is dispensable for SARS-CoV-2 replication and fitness.
• The 3'-UTR RNA structure remains largely intact even after s2m deletion.
• These findings challenge the presumed necessity of s2m for viral viability in SARS-CoV-2.