TY  - GEN
N2  - The centromere is an essential locus for chromosome segregation. It serves as a platform for kinetochore assembly and subsequent spindle microtubule attachment during cell division. Histone H3 variant CENP-A, or CID in Drosophila, is incorporated into centromeric chromatin to epigenetically mark its location. To maintain the centromere, new CID-containing nucleosomes are loaded by the chaperone Cal1, and targeted to the centromere by the inner kinetochore protein CENP-C. Even though centromeres are epigenetically defined and thus independent of the underlying DNA sequences, most are localized on repetitive DNA. In Drosophila, the centromere of each chromosome has a unique combination of transposable elements (TEs), flanked by both simple and complex satellite repeats. In all eukaryotes studied so far, centromeric transcription and the derived centromeric RNAs (cenRNAs) seem to be a structural or regulatory component of the centromere. In Drosophila, the (peri)centromeric RNA SATIII is a part of centromeric chromatin and important for CID loading. 
Due to the wide variety of repeats and TEs present at Drosophila centromeric DNA, it is likely that we do not have the full picture of the RNAs that are associated with centromeres. Therefore, I set out to identify all centromere-associated RNAs using RNA chromatin immunoprecipitation followed by deep sequencing (RNA-ChIP-seq) in embryos. RNAs from several TEs were enriched in this centromeric pull down, most significantly gypsy element Blastopia. I validated the centromeric localization of Blastopia with single molecule RNA fluorescent in situ hybridization (smRNA-FISH), but was unable to observe pronounced mitotic defects after knockdown. 
Additionally, I set out to identify which of the factors involved in the CID loading machinery are RNA-binding proteins. Using an RNA-protein complex isolation method called XRNAX, I showed that Cal1 and CID, but not CENP-C, directly bind RNA in S2 cells. By sequencing the RNAs linked to Cal1, I identified the centromere-associated TE Copia as the interaction RNA of Cal1. smRNA-FISH confirmed the presence of Copia at centromeres. 
Taken together, I identified novel centromere-associated RNAs as a structural or regulatory component of centromeric chromatin, some of which directly interact with the key centromeric protein Cal1.
A1  - Valent, Iris Anne
UR  - https://archiv.ub.uni-heidelberg.de/volltextserver/32271/
ID  - heidok32271
AV  - public
CY  - Heidelberg
TI  - Centromere-Associated RNAs in Drosophila melanogaster
Y1  - 2022///
ER  -