ASSOCIATION OF 2 BARLEY YELLOW MOSAIC-VIRUS (RNA-2) ENCODED PROTEINS WITH CYTOPLASMIC INCLUSION-BODIES REVEALED BY IMMUNOGOLD LOCALIZATION

Antisera were raised against the RNA 2-encoded proteins of 28 kDa and 70 kDa of barley yellow mosaic virus (BaYMV) by using the correspondin g cDNA sequences of a German isolate for protein overexpression in Esc herichia coli BL 21 and subsequent purification. The proposed processi ng of a 98 kDa precursor polyprotein encoded by the long open reading frame of RNA2 to two proteins of 28 kDa and 70 kDa could be confirmed by immunoprecipitation of the in vitro transcribed and translated cDNA -clone of RNA 2 and Western blot analysis of fragmentated protein extr acts of BaYMV-infected winter barley plants. In situ localisation stud ies of infected leaf tissue using immunogold labeling techniques for e lectron microscopy revealed that both viral proteins of BaYMV (RNA 2) were associated with the crystal-like cytoplasmic inclusion bodies. No other parts of the cells and no other inclusions (pinwheel-structures or aggregated virus particles) showed any gold labeling when the 28 k Da and 70 kDa antisera were used. We suppose that both RNA 2-encoded p roteins take part in the formation of the crystal-like cytoplasmic inc lusion bodies which are the most dominant structures in the cytoplasm of BaYMV-infected tissue. Possible functions of the 28 kDa and 70 kDa protein of BaYMV (RNA 2) are discussed.