Localization of multiple TMV encapsidation initiation sites on rbcL gene transcripts

Area of Science:

• Molecular Biology
• Virology
• Plant Science

Background:

• Tobacco mosaic virus (TMV) capsid protein is known to encapsidate various RNA molecules.
• Pseudovirions containing chloroplast DNA transcripts are formed in vivo by TMV capsid protein.

Purpose of the Study:

• To investigate the encapsidation initiation reaction of chloroplast-encoded rbcL mRNA with TMV capsid protein.
• To identify and characterize the sites on rbcL mRNA responsible for initiating encapsidation.

Main Methods:

• In vitro reaction assays between TMV capsid protein oligomers and rbcL mRNA.
• Nucleotide sequence analysis and secondary structure prediction of rbcL mRNA regions.
• Comparison of rbcL mRNA initiation site efficiency with viral RNA initiation sites.

Main Results:

• At least three distinct sites on rbcL mRNA independently initiate encapsidation by TMV capsid protein.
• These rbcL mRNA initiation sites exhibit lower in vitro reaction efficiency compared to the viral RNA initiation site (ei).
• The most reactive rbcL site (ei-3) shares sequence homology with TMV U1 and Cc strains' initiation sites and can form similar secondary structures.

Conclusions:

• Chloroplast rbcL mRNA contains functional encapsidation initiation sites for TMV capsid protein.
• The identified rbcL sites represent a novel mechanism for non-viral RNA encapsidation by TMV.
• A potential transcription termination signal downstream of the rbcL coding region may also influence encapsidation.