Structural domains within the 3' untranslated region of Turnip crinkle virus.
The genomes of positive-strand RNA viruses undergo conformational shifts that complicate efforts to equate structures with function. We have initiated a detailed analysis of secondary and tertiary elements within the 3' end of Turnip crinkle virus (TCV) that are required for viral accumulation in vivo. MPGAfold, a massively parallel genetic algorithm, suggested the presence of five hairpins (H4a, H4b, and previously identified hairpins H4, H5, and Pr) and one H-type pseudoknot (Psi(3)) within the 3'-terminal 194 nucleotides (nt). In vivo compensatory mutagenesis analyses confirmed the existence of H4a, H4b, Psi(3) and a second pseudoknot (Psi(2)) previously identified in a TCV satellite RNA. In-line structure probing of the 194-nt fragment supported the coexistence of H4, H4a, H4b, Psi(3) and a pseudoknot that connects H5 and the 3' end (Psi(1)). Stepwise replacements of TCV elements with the comparable elements from Cardamine chlorotic fleck virus indicated that the complete 142-nt 3' end, and subsets containing Psi(3), H4a, and H4b or Psi(3), H4a, H4b, H5, and Psi(2), form functional domains for virus accumulation in vivo. A new 3-D molecular modeling protocol (RNA2D3D) predicted that H4a, H4b, H5, Psi(3), and Psi(2) are capable of simultaneous existence and bears some resemblance to a tRNA. The related Japanese iris necrotic ring virus does not have comparable domains. These results provide a framework for determining how interconnected elements participate in processes that require 3' untranslated region sequences such as translation and replication.
McCormack
John C
JC
Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, Maryland 20742, USA.
Yuan
Xuefeng
X
Yingling
Yaroslava G
YG
Kasprzak
Wojciech
W
Zamora
Rodolfo E
RE
Shapiro
Bruce A
BA
Simon
Anne E
AE
D020413:3' Untranslated Regions / Q000737:chemistry* / Q000235:genetics; D000465:Algorithms; D020137:Base Pair Mismatch; D001483:Base Sequence; D019184:Carmovirus / Q000737:chemistry* / Q000235:genetics; D003198:Computer Simulation; D005814:Genes, Viral; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D010957:Plasmids; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D019244:RNA, Satellite / Q000737:chemistry; D012367:RNA, Viral / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D014779:Virus Replication
D020413:3' Untranslated Regions; D019244:RNA, Satellite; D012367:RNA, Viral; D012313:RNA