International Laboratory for Tropical Agricultural Biotechnology, Donald Danforth Plant Science Center, 975, N. Warson Road, St. Louis, MO-63132, USA.
Abstracts of the papers presented at the 16th Annual Convention and International Symposium of Indian Virological Society on “Management of Vector-Borne Viruses” at International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru-502324, Hyderabad, India, February 7–10, 2006.
Plant post-transcriptional gene silencing (PTGS) is a natural defense response against viruses, and as a counter-defense strategy, viruses encode PTGS suppressor proteins. MicroRNAs (miRNAs) are endogenous, ~21 nt non-coding RNAs that negatively regulate target mRNAs in eukaryots. Systemic infection by plant viruses frequently results in symptoms that resemble developmental defects caused by improper control of gene expression. Symptoms are often associated with virus-encoded pathogenicity factors, many of which are PTGS suppressors.
Transformation of Arabidopsis thaliana ecotype Columbia was performed by floral dipping method using A. tumefaciens harboring the AC4 gene of ACMV. miRNAs levels of target mRNAs in transgenic plants were analyzed in Northern blot assays. In vitro binding studies were performed using purified AC4 proteins and oligo-nucleotides in gel shift assays. In vivo binding capacity of AC4 to miRNAs was detected using 2’-O-methyl oligo-nucleotide and immunoprecipitation assays. Virus inoculation was performed using the infectious clones by particle bombardment.
The transgenic expression of ACMV-AC4 protein (a PTGS suppressor) shows down-regulated host miRNAs correlated with an up-regulation of the corresponding mRNAs, and induces developmental abnormalities in Arabidopsis. In vitro and in vivo binding assays revealed the ability of ACMV-AC4 to bind miRNAs, but not to miRNA:miRNA* duplexes. A normally labile intermediate in the miRNA biogenesis/RISC assembly, miRNA*, was below the detection level. In addition it was demonstrated that ACMV-AC4 binds to ss-siRNAs but not to siRNAs.
ACMV-AC4 protein has the capacity to bind miRNAs or ss-siRNAs, probably recruiting them by interacting with factors associated with RLC, although such a complex still needs to be unveiled in plants, or by interacting with components associated with the RISC complex. Viruses have evolved a range of PTGS suppressors capable of interfering with gene silencing at different levels, binding with miRNAs or ss-siRNAs is the ultimate step before target mRNAs are cleaved or silenced. It remains to be seen whether animal viruses encode miRNA-binding proteins, which might provide insight, how viruses manipulate the cell system to develop diseases in plants and animals.
