Interaction of RNA polymerase II and the RNA silencing machinery in heterochromatin formation of Drosophila

Abstract

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Heterochromatin is thought to be formed by transcription via RNA Pol II of aberrant/repeated regions such as those found in the centromeric regions that enter into the RNAi silencing pathway and guide chromatin modifications for gene silencing. We examined this issue in Drosophila. Our initial results indicate genetic and biochemical interactions between RNAi related components in Drosophila (dcr2, aub, ago2, hls and piwi) and the second largest subunit of RNA Pol II. The interaction between mutations in the RNAi silencing machinery and RNA pol II results in a strong suppression of PEV (Position Effect Variegation) indicating a possible effect on heterochromatin formation in Drosophila. The analysis of polytene chromosomes revealed a decrease in the H3-K9 me2 modification at the chromocenter in the double heterozygous (hlsE616, hls125 , dcr2 and RNA Pol II A5) mutants when compared with wild type and single heterozygotes. In addition the double heterozygotes also showed mislocalisation of Heterochromatin Protein 1(HP1). The results also indicated that the RNA Pol II and RNA silencing machinery components do not affect post-transcriptional gene silencing. In addition co-immunoprecipitation data indicate that RNA Pol II biochemically interacts with components of the RNA silencing machinery.