Citrus psorosis virus 24K protein interacts with citrus miRNA precursors, affects their processing and subsequent miRNA accumulation and target expression (disertación)

REYES CA

Simposio; Plant biology symposium: Learning from infections; 2015

Sweet orange (Citrus sinensis) is one of the most important fruit crops worldwide. Disease symptoms caused by virus infections can interfere with cellular processes causing dramatic economic losses. Citrus psorosis virus (CPsV) is the type member of the genus Ophiovirus. Its genome consists of three ssRNAs of negative polarity that encodes four proteins: the RNA-dependent RNA polymerase of 280 kDa, the movement protein of 54 kDa (54K), the coat protein and a small protein of 24 kDa (24K). Here we demonstrate that infection of sweet orange plants with Citrus psorosis virus (CPsV) alters accumulation of a set of microRNAs (miRNAs). miR156, miR167 and miR171 were the strongest down- regulated reaching levels of almost 3 folds reduction in infected samples and leading to a concomitant up-regulation of some of its targets: Squamosa promoter binding protein like (SPL) 9 and 13 and Scarecrow-like 6 (SCL6). The most severe symptom, expressed as necrosis of young shoots (shock), showed the strongest decrease in miRNA levels, particularly miR156. Secondary structure of sweet orange pre-miR156 family of precursors (a to g) was predicted and analysis is consistent with a loop to base model of processing by DCL1. Pre-miR156 rocessing is affected in sweet orange by the virus as manifested by increment in the level of unprocessed pre-miR156 in relation to mature species in Northern blot assays. Co-immunoprecipitation of viral proteins and pre-miRNAs were performed and an association between pre-miR156a and the viral 24K protein was evidenced. These results let us conclude that processing alteration might be due to direct or indirect interaction of miRNA precursors with viral proteins, coincident also with their nuclear localization by confocal microscope observations. This work contributes to the understanding of how virus can alter host regulatory mechanisms, particularly miRNAbiogenesis and function.