The budding yeast heterochromatic protein Sir3 modulates genome-wide gene expression through transient direct contacts with euchromatin

Abstract

The budding yeast SIR complex (Silent Information Regulator) is the principal actor in heterochromatin formation, which causes epigenetically regulated gene silencing phenotypes.The maternal chromatin bound SIR complex is disassembled during replication.Consequently, if heterochromatin is to be restored on both daughter strands, the SIR complex has to be reformed on both strands to pre-replication levels. The dynamics of SIR complex maintenance and re-formation during the cell-cycle and in different growth conditions are however not clear. Understanding exchange rates of SIR subunits during the cell cycle and their distribution pattern to daughter chromatids after replication has important implications for how heterochromatic states may be inherited and therefore how epigenetic states are maintained from one cellular generation to the next. We used the tag switch RITE system to measure genome wide turnover rates of the SIR subunit Sir3 before and after exit from stationary phase and show that maternal Sir3 subunits are completely replaced with newly synthesized Sir3 at subtelomeric regions during the first cell cycle after release from stationary phase. The SIR complex is therefore not "inherited" and the silenced state has to be established de novo upon exit from stationary phase. Additionally, our analysis of genomewide transcription dynamics shows that precise Sir3 dosage is needed for the optimal upregulation of "growth" genes during the first cell-cycle after release from stationary phase.

Application of Recombination -Induced Tag Exchange (RITE) to study histone dynamics in human cells

“…RITE is a versatile method to monitor protein replacement that has been applied to histones [ 53 – 55 , 57 , 59 – 63 , 31 ] Ramakrishnan et al, 2016 and non-histone proteins [ 58 , 71 – 73 ] Menendez-Benito et al 2012 [ 74 ]; in budding and fission yeast. RITE is compatible with pull-down strategies, which enables genome-wide mapping or determining modifications of old and new histones by mass-spectrometry [ 56 ].…”