THE SMALL CYSTEINE-RICH PROTEIN P14 OF BEET NECROTIC YELLOW VEIN VIRUS REGULATES ACCUMULATION OF RNA-2 IN CIS AND COAT PROTEIN IN TRANS

The effect of null mutations of the small cysteine-rich protein P14 en coded by RNA 2 of beet necrotic yellow vein virus has been investigate d using in vitro transcripts of viral RNA to infect Chenopodium quinoa protoplasts. The P14 mutations down-regulated RNA 2 accumulation by a pproximately 10- to 50-fold. Accumulation of minus-strand RNA 2 was al so diminished but RNA 1 accumulation was much less affected. The inhib ition of RNA 2 accumulation could not be complemented in trans by prov iding P14 from another source (either a second molecule of RNA 2 or an RNA 3-based replicon) containing and expressing the P14 gene. The P14 null mutations dramatically inhibited accumulation of viral coat prot ein, which is encoded by the 5'-proximal gene on RNA 2, but this effec t could be complemented in trans, indicating that it occurs by a mecha nism distinct from that affecting RNA 2 accumulation. Transient expres sion experiments were also carried out in which a plasmid expressing P 14 and plasmids expressing a reporter gene placed downstream of potent ial translational control sequences (the 5'-noncoding sequences of RNA s 2, 3, or 4) were introduced into C. quinoa or Nicotiana tabacum leav es by microprojectile bombardment. Coexpression of P14 produced a 3- t o 4-fold stimulation of reporter gene expression levels for all the co nstructs. The lack of sequence specificity suggests that this phenomen on is not directly related to the RNA 2-specific stimulation of coat p rotein accumulation observed in a viral infection. (C) 1995 Academic P ress, Inc.