Contributions of chromatin remodelling factors CHRAC/ACF to epigenome programming during oogenesis and early embryogenesis in Drosophila melanogaster
Allgemeine Genetik und funktionelle Genomforschung
Zusammenfassung der Projektergebnisse
We explored the role of the nucleosome sliding factor ACF during Drosophila oogenesis and early embryogenesis. ACF consists of two components, the ATPase ISWI and a larger subunit, ACF1. Since ISWI is found in several remodeling complexes, we focused on ACF1 subunit, as the defining subunit. Coupling chromatin immunoprecipitation to sequencing (ChIP-Seq) we generated genome-wide chromosomal binding profiles of ACF1 in chromatin of embryos and embryonic cell lines and correlated them with published genomic features. ACF1 binds many nucleosome-free regions at transcription start sites of developmental genes. Surprisingly, ACF1 gene mutation or depletion of ACF1 in cells did not lead to a widespread deregulation of ACF1-bound genes. This may be explained by functional redundancy with other nucleosome remodeling factors. Indeed we found that RSF1, which associates with ISWI to form the RSF complex, occupies the very same sites as ACF1. Genetic analyses document strong synthetic phenotypes (lethality or developmental arrest) at all stages of development. How those two factors affect chromatin organisation at their target sites is a focus of current research. We also explored the protein interactors with ACF1 by immune-affinity purification of ACF1 and identification of co-purifying proteins through mass spectrometry. ACF1 was shown to ‘pull down’ a number of proteins involved in transcription and replication. These links are currently being tested for functional relevance. Our biochemical studies also revealed an interaction of ACF with the large Domino/Tip60 complex, a nucleosome remodeling complex involved in the exchange of the histone variant H2A.V. The potentially synergistic actions of a nucleosome sliding factor (ACF) and a histone exchanger (DOM/Tip60) during Drosophila development and in response to DNA damage are the focus of our follow-up studies. Despite the functional redundancy of ACF1 and RSF1 we found that certain acf1 mutant alleles show strong oogenesis phenotypes, including faulty oocyte fate specifications that had not been reported earlier. Our genetic and molecular analyses lead us to conclude that the available acf1 alleles that had been published as loss-of-function alleles are presumably not what they appear. Conceivably, they still express ACF1 C-terminal fragments that disrupt a carefully balanced interplay of local and global chromatin remodeling.
Projektbezogene Publikationen (Auswahl)
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(2010). Developmental role for dACF1-containing nucleosome remodelers in chromatin organisation. Development 137, 3513-3522
Chioda, M., Vengadasalam, S., Kremmer, E., Eberharter, A. & Becker, P.B.
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(2013). The variant histone H2A.V of Drosophila-three roles, two guises. Chromosoma 122, 245-258
Baldi, S. & Becker, P.B.
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(2014). The histone-fold protein CHRAC14 influences chromatin composition in response to DNA damage. Cell Reports 7 321-30
Mathew, V., Pauleau, A.L., Steffen, N., Bergner, A., Becker, P.B. & Erhardt, S.
