MOLECULAR-CLONING, SEQUENCING, FUNCTIONAL-ANALYSIS AND EXPRESSION IN ESCHERICHIA-COLI OF MAJOR CODE PROTEIN GENE (S3) OF RICE DWARF VIRUS CHINESE ISOLATE

The complete nucleotide sequence of major core protein gene (segment S 3) of rice dwarf virus (RDV) Chinese isolate was determined after cDNA cloning from the viral genomic RNA. Sequence analysis showed that the cloned fragment is 3195 bp in length and containes a single open read ing frame (ORF), encoding the major core protein (P3) which M-r of 114 K. The nucleotide and deduced amino acid sequences of S3 of this isol ate share significant homology (94.1% and 97%, respectively) with thos e of 83 of the Japanese isolate. At the amino acid level, P3 of RDV Ch inese isolate shares significant homology with P3 of rice gall dwarf v irus (RGDV), significant regional homology with the rotavirus VP4 prot ein which forms spikes on the virus particles and has been identified as a protein involved in the activation of the rotavirus penetration, and homology with spheroidin of amsacta entomopoxvirus (SPH), which is the major protein of the occlusion body, with clp-like ATP-dependent protease binding subunit and with ATP-dependent protease ATP-binding s ubunit. Amino acid sequence analysis also showed that P3 contains RNA- dependent RNA polymerase (RDRP) motif-like elements such as DXXXD, SGX XXXXXN, GDD and ENXXXY. These results may suggest that P3 is a multifu nctional protein which plays very important roles in the virus structu re formation, virus replication and penetration processes. The full le ngth cDNA sequence of RDV S3 and a partial one which covers nt 1004-31 95 were cloned into bacterial expression vector pTrcHisB for expressio n. The full length cDNA sequence failed to be expressed in E. coli, bu t the partial sequence was successfully expressed there as confirmed b y the Western blot analysis. Further analysis of RDV P3 is under way.