Functional study of chromatin modulators-histone H1 and HP1 in "Drosophila melanogaster"

Abstract

The impact of chromatin structure on transcriptional gene activity, and many other nuclear events, has become increasingly apparent over the past few decades. It is known that eukaryotic DNA in the cell nucleus is packaged into periodic nuclear proteins known as nucleosomes, the basic units of chromatin. Within each nucleosome, about 146 bp of DNA is wrapped around a core histone particle consisting of two molecules each of histones H2A, H2B, H3 and H4. It is believed that linker histone H1 binds to the linker DNA between nucleosomes, to stabilise the nucleosome and protect an additional 20 bp of DNA from nuclease digestion. Histone H1 promotes or facilitates the condensation of nucleosome filaments into supercoiled chromatin fibres, then further forms chromosomes, which can normally be seen under a microscope. Studies in vitro have shown that H1 is a transcriptional repressor, while the effect of histone H1 on transcription in vivo is rather gene-specific. Linker histone H1 inhibits DNA repair and homologous recombination in unicellular and simple multicellular organisms. In higher multicellular organisms, H1 appears to play a key role in apoptosis and cell differentiation. However, the dynamics of histone H1 in higher-order chromatin packaging, and its role in transcriptional gene regulation, remain largely unknown. The eukaryotic linker histone H1 has a typical structure consisting of a tripartite structure of a trypsin-resistant central globular domain flanked by basic N- and C-terminal tails. It has been proposed that the globular domain binds the DNA where it enters and exits the nucleosome, while the C-terminal tail binds to the linker DNA and facilitates condensation of chromatin. Several models have been suggested, based on indirect biochemical evidence, for the location of H1 in nucleosomes. However, the precise location of H1 in the nucleosome and how it is involved in higher-order chromatin packaging still remain debated issues. Unlike mammalian cells which have many H1 variants, Drosophila melanogaster contains about 100 copies of histone H1 genes but these encode only a single type of H1 protein with a structure typical of linker histone H1 in higher eukaryotes, and thus provides us with an ideal model system to address the function of H1 in chromatin and its impact on development. Using in vitro and in vivo biochemical and genetic approaches, we have tried to investigate the role of H1 in nucleosome dynamics and chromatin transcriptional gene silencing. Besides linker histone H1 and core histones on chromatin, a large number of non-histone proteins, such as polycomb group protein, trithorax protein and HMG protein, are also associated with chromatin and play important roles in gene transcription. Another molecule, which we are interested in, is heterochromatin protein 1 (HP1): this is of the key components of condensed chromatin, and is primarily localised at heterochromatic domains. Our study showed that a number of regions in euchromatin also contain HP1, indicating that HP1 plays a genome-wide role in chromatin organization. Other recent papers have described the interaction of HP1 with both methylated histone H3 at lysine 9 and the methyltransferase enzyme (Su(var)3-9), and have further proposed a mechanism for maintenance and spreading of heterochromatin. To access the role of HP1 in cell proliferation and development, we conditionally deplete HP1 using the RNA interference (RNAi) approach. The effects of HP1 on chromatin structure, cell cycle regulation, genome-wide gene expression and late-stage development are being studied.