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Nucleic Acids Research, 1994, Vol. 22, No. 8 1346-1353
© 1994

RNA

The 3'-untranslated region of alfalfa mosaic virus RNA 3 contains at least two independent binding sites for viral coat protein

Chantal B.E.M. Reusken, Lyda Neeleman and John F. Bol*

Institute of Molecular Plant Sciences, Gorlaeus Laboratories, Leiden University PO Box 9502, 2300 RA Leiden, The Netherlands

*To whom correspondence should be addressed

Received February 17, 1994. Revised March 22, 1994. Accepted March 22, 1994.

The 3'-termini of the three genomlc RNAs of alfalfa mosaic virus contain a common sequence of 145 nucleotldes (nt) with a specific binding site for coat protein (CP). This sequence consists of several stem/loop structures interspersed with single-stranded AUGC-motifs; in RNA 3 this folding pattern is extended to a region upstream of the homologous sequence. By band-shift assays a minimum of two specific binding sites for CP were Identified near the 3'-end of RNA 3. Site 1 consists of the region between nt 11 and 127 from the 3'-end and contains two AUGC-motifs. Site 2 is located between nt 133 and 208 from the 3'-end in a sequence that is largely unique to RNA 3 and contains also two AUGC-motifs. Deletion studies revealed that the two sites could bind CP independently of each other and permitted the identification of sequence elements that are essential for the activity of each site. By site-directed mutagenesis it was shown that the AUGC-motifs are important for binding of CP to both sites. These binding sites may play a role in the phenomenon that each genomic RNA has to be complexed with a few CP molecules to initiate Infection. Later in the replication cycle they may act as origins for the assembly of virus particles.


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