Defining the functional bases within centromeric promoters

Abstract

During cellular division, DNA is duplicated and partitioned such that the two new cells inherit the same genetic information. Failure to form a centromere, a complex of DNA and proteins, causes errors in DNA division and can cause developmental defects in humans. Heterochromatin, a condensed form of DNA and DNA-associated proteins, is necessary for centromere formation. Previous work has shown that RevCen, a transcribed DNA sequence present in multiple copies at the centromere, is sufficient to recruit heterochromatin and silence nearby genes. To test whether RevCen transcription is important for silencing, we engineered versions of RevCen without a promoter. We show that RevCen-mediated gene silencing is partially dependent on the presence of its promoter. Future work will confirm that the loss of gene silencing is accompanied by a loss of heterochromatin and will determine whether RevCen transcript levels decrease when the promoter is absent. To define the functional bases within the RevCen promoter, we will create a series of promoter deletion fragments and measure their ability to initiate transcription and establish heterochromatin. This work demonstrates that the RevCen promoter is important for gene silencing and will identify specific sequences within the promoter that are necessary for transcription and heterochromatin establishment.