2 DOMAINS OF THE AL1 PROTEIN MEDIATE GEMINIVIRUS ORIGIN RECOGNITION

The geminiviruses tomato golden mosaic virus (TGMV) and bean golden mo saic virus (BGMV) have bipartite genomes. Their A and B DNA components contain cis-acting sequences that function as origins of replication, while their A components encode the trans-acting replication proteins - AL1 and AL3. Earlier experiments demonstrated that virus-specific i nteractions between the cis-and trans-acting functions are required fo r TGMV and BGMV replication and that the ALI proteins of the two virus es specifically bind their respective origins. In the current study, c haracterization of AL1 and AL3 proteins produced from plant expression cassettes in transient replication assays revealed that interaction b etween ALI and the origin is responsible for virus-specific replicatio n. The AL3 protein does not contribute to specificity but can he prefe rred by its cognate ALI protein when replication is impaired. Analysis of chimeric proteins showed that two regions of ALI act as specificit y determinants during replication. The first domain is located between amino acids 1 and 116 and recognizes the ALI origin binding site. The second region, which is between amino acids 12.1 and 209, is not depe ndent on the known ALI DNA binding site. Analysis oi wild type and chi meric proteins in transient transcription assays showed that ALI also represses its own promoter in a virus-specific manner. Transcriptional specificity is conferred primarily by ALI amino acids 1-93 with amino acids 121-209 making a smaller contribution. Together, these results demonstrated that the virus-specific interactions of ALI. during repli cation and transcription are complex, involving at least two discreet domains of the protein. (C) 1997 Academic Press.