Abstract: 【Objective】Yellow leaf (YL) disease of sugarcane is one of viral diseases in major sugarcane-producing regions in China and worldwide, causing a serious threat to sugarcane industry. Sugarcane yellow leaf virus (SCYLV), a member of the genus Polerovirus and family Luteoviridae, is the causal agent of YL. The aim of this study is to characterize the conserved regions of P0 protein encoded by SCYLV, and to test its functionality as a RNA silencing suppressor in natural host sugarcane (Saccharum spp. hybrids). The findings will contribute to further investigate the molecular mechanism of SCYLV pathogenesis at RNA silencing level. 【Method】P0 and its two deletion mutants, P0^?2-15(15-aa deletion in N terminal) and P0^?155-256(102-aa deletion in C terminal) from SCYLV CHN-FJ4 isolate were obtained by RT-PCR and cloned into an expression vector under control of the maize ubiquitin 1 promoter and Agrobacterium nopaline synthase terminator. Each of P0 or its deletion mutant constructs were co-bombarded with the reporter gene coding for enhance yellow fluorescent protein (EYFP) into sugarcane young leaf segments, and transient EYFP expression was quantified using ImageJ software. The Tomato bushy stunt virus-encoded P19 was included as a model suppressor.【Result】Phylogenetic analysis revealed that the CHN-FJ4 isolate clustered with isolates of the BRA genotype and shared 96.1%-98.4% aa identified with them. MEME software online revealed that the P0 proteins contained three significant conserved regions positioned at 1-60, 76-125, and 161-210 aa residues. Besides, eight positive sites were detected using five different approaches under the datamonkey web-server. P0, P0^?2-15 and P0^?155-256 were co-bombarded with EYFP into sugarcane young leaf segments, respectively. In young leaf segments co-expressing a suppressor P0 or P19, EYFP foci account and EYFP expression level were increased at 48-120 h post-DNA introduction compared with that in the absence of a suppressor. At 120 h post-bombardment, EYFP foci account and of EYFP expression levelincreased by more than 1.6-fold and 4.0-fold, respectively. No significant differences (P>0.05) in EYFP expression were found in presence of P0 or P19. However, EYFP expression was not increased by P0^?2-15 or P0^?155-256. The reason would be 15-aa deletion in N-terminal or 102-aa deletion in C-terminal of P0 protein impaired and even disabled the function of RNA silencing suppressor.【Conclusion】The P0 RNA silencing suppressor proved to be efficient in enhancing reporter gene expression in transient assay using sugarcane young leaf segments. The conserved regions of P0, 15-aa in N terminal and 102-aa in C terminal, are necessary for the RNA silencing suppressor activity of P0.

Key words: Sugarcane yellow leaf virus, P0 protein, RNA silencing suppressor, EYFP