RICE TUNGRO BACILLIFORM VIRUS ENCODES REVERSE-TRANSCRIPTASE, DNA-POLYMERASE, AND RIBONUCLEASE-H ACTIVITIES

Rice tungro bacilliform virus (RTBV) is a newly described badnavirus a nd proposed member of the plant pararetrovirus group. RTBV open readin g frame 3 is predicted to encode a capsid protein, protease (PR), and reverse transcriptase (RT) and has the capacity to encode other protei ns of as yet unknown function. To study the possible enzymatic activit ies encoded by open reading frame 3, a DNA fragment containing the put ative PR and RT domains was used to construct the recombinant baculovi rus PR/RT-BBac. Trichoplusia ni insect cells infected with PR/RT-BBac were used in pulse-labeling experiments and demonstrated synthesis of an 87-kDa polyprotein that corresponds in molecular mass to that predi cted from the PR/RT DNA coding sequence. The 87-kDa polyprotein was pr ocessed with concomitant accumulation of 62-kDa (p62) and 55-kDa (p55) proteins. Amino-terminal sequencing of p62 and p55 determined that th ey mapped to the PR/RT domain and shared common amino termini. p62 and p55 were purified and exhibited both RT and DNA polymerase activities using synthetic primer/template substrates. Only p55 had detectable r ibonuclease H activity, an activity intrinsic to all reverse transcrip tases studied to date. Characterization of the RTBV RT provides a bioc hemical basis for classifying RTBV as a pararetrovirus and will lead t o further studies of these proteins and their role in virus replicatio n.