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small RNA mediated heterochromatin formation in fission yeast

Final Report Summary - SMALLRNAHET (small RNA mediated heterochromatin formation in fission yeast)

Regulation of genome expression is essential for many cellular processes including cell proliferation, differentiation, development and viability. Changes in genome expression and genomic instability are allowing cancer cells to acquire their characteristics. It has become clear over the past decades that histone modifications are essential in the cellular control of gene expression. In this project we have combined molecular biology, genetics,biochemistry and cryo-EM to dissect requirements for small RNA generation and heterochromatin formation in fission yeast. Our work has contributed to the understanding of fundamental process of how genomes recognize repetitive and transposable elements and target them into heterochromatin (Marasovic et al, Mol Cell, 2013). To achieve this, the host cells generate Dicer-independent small RNAs that scan the transcriptome to distinguish self from non-self (Marasovic et al, Mol Cell, 2013). We have also shown that degradation of Argonaute-bound small RNAs is essential in providing fidelity in RNAi mediated silencing (Pisacane and Halic, Nature Communications, 2017) and that retention of lncRNAs on chromatin perturbs heterochromatin organization and silencing (Broenner et al, Genome Research, 2017). We also use cryo-EM to understand mechanisms of RNAi and heterochromatin formation. We have solved cryo-EM structure of a chromodomain, a reader of H3K9 methylation mark, bound to the H3K9 methylated nucleosome (Zocco et al, Cell Discovery, 2016). Our recent cryo-EM structures show how nucleosomes open and unwind DNA and how histone octamer adapts to DNA unwinding (Bilokapic et al, NSMB, 2018). We have also solved several structures that reveal how histone octamer core rearranges and deforms the nuclesome to translocate the DNA. These structures unveil first mechanistic insights into nucleosome remodeling. Nucleosome plasticity observed in our structures is likely exploited by other chromatin machineries beyond chromatin remodeling (Bilokapic et al, Nature Communications, 2018). Our recent cryo-EM data show that two nucleosomes can interact in many different ways (Bilokapic et al, Sci Rep, 2018) indicating that chromatin structure is very flexible.