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Wang, Alexander B.

University of British Columbia

Zika virus (ZIKV) was first isolated in Uganda in 1947 and circulated quietly throughout Africa and Southeast Asia during the 20th century, sporadically causing mild febrile disease. However, recent ZIKV outbreaks have been associated with severe neurological complications. Along with these dramatic phenotypic changes came viral genetic evolution, leading to the formation of distinct African and Asian lineages of the virus. Much work has been done characterizing strain level and amino acid level differences, but the relationship between ZIKV genetic evolution and viral phenotype has remained unclear. Surprisingly, nucleotide polymorphisms themselves, particularly in the viral untranslated regions (UTRs), which play crucial roles in coordinating several stages of the viral life cycle, have been understudied to date. In particular, the 5′ terminal structure in the ZIKV genome, termed stem-loop A (SLA), is essential in promoting viral RNA replication and capping, yet little is known regarding its evolution. Herein, we investigate ZIKV SLA evolution both structurally and functionally. We describe four main evolutionarily-conserved SLA variants that show differences in RNA structure and/or base pairing, termed ancestral (Anc), intermediate (Int), contemporary (Con) and alternative (Alt). Within the African ZIKV lineage, SLAInt is the most common, while the Asian lineage predominantly encoded SLACon. SLAAnc, SLAInt, and SLACon displayed very similar predicted structure and base-pairing, but evolutionarily-acquired nucleotide polymorphisms resulted in a stepwise reduction in thermodynamic stability of SLA over time. However, SLAAlt exhibited a drastically modified RNA structure and a higher overall thermodynamic stability. Examination of viral RNA accumulation of SLA variants in a subgenomic replicon model suggested similar overall viral RNA accumulation across SLAAnc, SLAInt and SLACon, while the SLAAlt variant was not replication-competent. Nonetheless, subsequent rapid amplification of cDNA ends (RACE) analysis on a natural ZIKV isolate that phylogenetically clusters with SLAAlt-encoding strains confirmed that SLAAlt exists in nature. This work emphasizes the importance of considering evolution within noncoding regions and illustrates the power of using evolution to inform our understanding of structured RNA function.

Text

2024-08-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/89118

Microbiology and Immunology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/