24K Citrus psorosis virus protein interferes with plant miRNA biogenesis and function

CARINA ANDREA REYES; ELIANA EVELINA OCOLOTOBICHE; FACUNDO MARMISOLLÉ; CECILIA BAEZA; ARIEL BAZZINI; SEBASTIÁN ASURMENDI; MARÍA LAURA GARCÍA

Keystone, Colorado

Simposio; Small RNA Silencing: Little Guides, Big Biology; 2016

Keystone Symposia on Molecular and Celular Biology

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) genome consists of three single-stranded RNAs of negative polarity encoding four proteins: the RNA-dependent RNA polymerase, the movement protein (54K), the coat protein and a small protein of 24 kDa (24K). Here we demonstrate that infection of sweet orange plants with CPsV alters processing of a group of conserved miRNA precursors (pre-miRNAs) leading to a reduction in mature miRNA species. miR156 and miR171 were the strongest down-regulated in infected samples with a concomitant up-regulation of some of its targets. Association between the viral 24K protein and pre-miR156a or pre-miR171a was evidenced by co-immunoprecipitation, suggesting that processing alteration might be due to direct or indirect interaction of pre-miRNAs with 24K in the nucleus. On the other hand, an inhibitory effect of this viral protein over the effector function of the RNA-induced Silencing Complex (RISC) was also shown using a transient expression assay of a reporter construct consisting in a miRNA target sequence fused to GFP together with 24K. The co-expression of 54K with 24K increases the mentioned inhibitory effect. This work contributes to the understanding of how virus can alter host regulatory mechanisms at different levels, particularly miRNAs biogenesis in the nucleus and their function in the cytoplasm.