Dissecting the Biological Functions of Drosophila Histone Deacetylases by RNA Interference and Transcriptional Profiling

Abstract

Chromatin modifications such as acetylation, methylation, phosphorylation, ubiquitination, and sumoylation of histones play an important role in the regulation of transcription (1-5). Together with DNA methylation, these modifications are part of a broad and multifaceted strategy whereby eukaryotes regulate gene expression at the epigenetic level (6 -8).Acetylation of histones is tightly controlled by the activity of histone acetyltransferases and histone deacetylases (HDACs) 2 (9, 10). Histone deacetylation may repress transcription by different mechanisms. On the one hand, this process increases the charge density on the N termini of the core histones thereby strengthening histone tail-DNA interactions and blocking access of the transcriptional machinery to the DNA template. In addition, histone modifications are specifically recognized by chromatin-interacting proteins thus favoring the formation of higher order chromatin structures (heterochromatin).There is ample evidence that aberrations in the epigenetic regulation of gene expression at the levels of DNA methylation and histone acetylation or histone methylation are an important component in the process of malignant transformation of human cells (11). A number of histone acetyltransferase mutations were identified in cancer, and HDACs were found to be overexpressed or aberrantly recruited in several human malignancies (12). Small molecule HDAC inhibitors induce cell cycle arrest, differentiation, and apoptosis in cancer cells with a significant window over normal cells, and some molecules have already entered clinical trials and show promising results (13).Higher organisms have evolved a considerable complexity in the histone deacetylase family; thus, in mammals, 17 different HDAC subtypes were identified, which were grouped into three classes according to their sequence homologies with the yeast proteins Rpd3 (class I), HDA1 (class II), and Sir (class III) (14 -16). Class I and class II proteins are evolutionarily related and share a common enzymatic mechanism, the Zn-catalyzed hydrolysis of the acetyl-lysine amide bond. The HDAC inhibitors that are presently in clinical trials are rather nonselective and are thought to inhibit most or many of the class I and II proteins. Class III proteins are evolutionarily unrelated to class I or class II and catalyze the transfer of the acetyl group onto the sugar moiety of NAD (16

HDAC3: taking the SMRT-N-CoRrect road to repression

“…Examining the recruitment of different factors on endogenous promoters, it was observed that both N-CoR/HDAC3 and the NuRD complex were recruited by tamoxifen-complexed ER, but in a sequential manner, following a strict choreography of events (Liu and Bagchi, 2004). The distinct functions of different HDACs and HDAC complexes are also suggested by overexpression/downregulation studies as well as developmental profiling of individual HDACs in Drosophila (Cho et al, 2005;Foglietti et al, 2006). Such studies reveal unique sets of genes regulated by each enzyme.…”

“…Such studies reveal unique sets of genes regulated by each enzyme. Surprisingly, similar numbers of genes were found to be repressed and activated in each case, perhaps through both direct and indirect effects (Cho et al, 2005;Foglietti et al, 2006).…”

Widespread regulation of gene expression in the Drosophila genome by the histone acetyltransferase dTip60

“…Foglietti et al performed genome-wide expression analysis upon HDAC1, 2, 3, 4 and X depletion, but surprisingly only found significant changes in gene expression for HDAC1 and HDAC3 knock-down (Foglietti et al, 2006). We therefore compared our expression data to the effect of depleting those two HDACs.…”

“…RNAi and TSA treatment as determined by (Foglietti et al, 2006). We asked if there is an overlap of respective subgroups based on flybase numbers.…”

“…RNAi knock-down of HDAC1-4 and HDAC X (Foglietti et al, 2006). First, we compared the genes upregulated or downregulated upon dTip60 loss to those upregulated or downregulated as a result of treatment with TSA.…”