title: The histone fold protein CHRAC-14 controls CENP-A loading and   gene expression in Drosophila melanogaster
creator: Doppler, Sarah
subject: 570
subject: 570 Life sciences
description: Centromeres are specialized chromatin domains present on each chromosome, which   determine the location of kinetochore formation – the multiprotein platform for spindle   microtubule attachment during mitosis. Since the underlying centromeric DNA sequence is not   conserved, centromere inheritance and function rely on the conserved epigenetic marker   CENP-A (also known as CID in Drosophila). CENP-A is highly enriched in centromeric   chromatin and partially replaces canonical histone H3. Specific loading and stabilization of   CENP-A at the centromere is ensured by a concert of different factors such as its dedicated   chaperone called CAL1 in Drosophila. Whereas the primary DNA sequence is dispensable for   centromere formation, CENP-A is both absolutely essential and sufficient to maintain and   establish functional centromeres. When overabundant due to transcriptional upregulation for   example in cancer cells, CENP-A escapes the endogenous loading pathway and hijacks other   histone loading machineries, which leads to promiscuous non-centromeric CENP-A   incorporation throughout the genome with detrimental effects for genome stability.   Even though ectopic CENP-A accumulation existentially threatens cellular and   organismal viability, CENP-A is distributed at basal levels throughout the genome under   physiological conditions. The function and mechanistic details of genome-wide CENP-A   loading remain elusive, but the field speculates, that low levels of ectopic CENP-A confer   epigenetic plasticity and prime chromatin for neocentromere formation in the event of ancestral   centromere loss. Previous work in our lab identified the histone fold protein CHRAC-14 as an   important regulator of ectopic CENP-A incorporation and as a new DNA damage factor in   Drosophila melanogaster. CHRAC-14 depletion leads to the increase of CENP-A levels,   increased CENP-A ectopic loading – possibly at telomeric DNA repair sites – and to DNA repair   defects accompanied by G2/M checkpoint failure.   Since we lacked detailed mechanistic insights and clear data whether ectopic CENP-A   localizes to and has a role at DNA lesions, I set out to further elucidate this pathway. Employing   a portfolio of biochemical and molecular biological techniques such as CUT&Tag-Seq, RNA-Seq and immunoprecipitation combined with mass spectrometry analysis, I report in this thesis,   that CHRAC-14 knockdown leads to the accumulation of CENP-A at centromeres and   genome-wide. In addition, altered expression of genes associated with gene ontology   categories such as ‘mitotic progression’ and ‘DNA damage response’ was observed.   Furthermore, I identified candidate genes, which are mis-expressed whilst showing increased   CENP-A binding. Moreover, I found, that both CHRAC-14 and CENP-A interact with Casein   kinase 2 (CK2) and are phosphorylation substrates thereof. Interestingly, CK2 phosphorylation   of CHRAC-14 seems to promote the turnover of a post translationally modified version of   CENP-A, whereas CK2 phosphorylation of endogenous CENP-A itself seems to be essential   for protein stability.   Taken together, my study provides new insights into CHRAC-14 mediated CENP-A   titration in chromatin and opens up a novel direction involving CK2 as a key regulator in this   pathway.
date: 2022
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/31436/1/Dissertation_SDoppler_2022.pdf
identifier: DOI:10.11588/heidok.00031436
identifier: urn:nbn:de:bsz:16-heidok-314363
identifier:   Doppler, Sarah  (2022) The histone fold protein CHRAC-14 controls CENP-A loading and gene expression in Drosophila melanogaster.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/31436/
rights: info:eu-repo/semantics/openAccess
rights: Please see front page of the work (Sorry, Dublin Core plugin does not recognise license id)
language: eng