Abstracts Category : Other

Chromatin association of UHRF1 during the cell cycle

by Bothayna Al-Gashgari

Ubiquitin-like with PHD and RING Finger domains 1 (UHRF1) is a nuclear proteinthat associates with chromatin. Regardless of the various functions of UHRF1 in thecell, one of its more important functions is its role in the maintenance of DNAmethylation patterns by the recruitment of DNMT1. Studies on UHRF1 based on thisfunction have revealed the importance of UHRF1 during the cell cycle. Moreover,based on different studies various factors were described to be involved in theregulation of UHRF1 with different functionalities that can control its binding affinityto different targets on chromatin. These factors are regulated differently in a cellcycle specific manner. In light of this, we propose that UHRF1 has different bindingbehaviors during the cell cycle in regard to its association with chromatin. In thisproject, we first analyzed the binding behavior of endogenous UHRF1 from differentunsynchronized cell systems in pull-down assays with peptides and oligonucleotides.Moreover, to analyze UHRF1 binding behavior during the cell cycle, weused two different approaches. First we sorted Jurkat and HT1080 cells based ontheir cell cycle stage using FACS analysis. Additionally, we synchronized HeLa cells todifferent stages of the cell cycle by chemical treatments, and used extracts from cellsortingand cell synchronization experiments for pull-down assays. We observed thatUHRF1 in different cell systems has different preferences in regard to its binding toH3 unmodified and H3K9me3. Moreover, we detected that UHRF1, in general,displays different patterns between different stages of cell cycle; however, we cannotdraw a final model for UHRF1 binding pattern during cell cycle. Ubiquitin-like with PHD and RING Finger domains 1 (UHRF1) is a nuclear proteinthat associates with chromatin. Regardless of the various functions of UHRF1 in thecell, one of its more important functions is its role in the maintenance of DNAmethylation patterns by the recruitment of DNMT1. Studies on UHRF1 based on thisfunction have revealed the importance of UHRF1 during the cell cycle. Moreover,based on different studies various factors were described to be involved in theregulation of UHRF1 with different functionalities that can control its binding affinityto different targets on chromatin. These factors are regulated differently in a cellcycle specific manner. In light of this, we propose that UHRF1 has different bindingbehaviors during the cell cycle in regard to its association with chromatin. In thisproject, we first analyzed the binding behavior of endogenous UHRF1 from differentunsynchronized cell systems in pull-down assays with peptides and oligonucleotides.Moreover, to analyze UHRF1 binding behavior during the cell cycle, weused two different approaches. First we sorted Jurkat and HT1080 cells based ontheir cell cycle stage using FACS analysis. Additionally, we synchronized HeLa cells todifferent stages of the cell cycle by chemical treatments, and used extracts fromAdvisors/Committee Members: Fischle, Wolfgang, Biological and Environmental Sciences and Engineering (BESE) Division, Merzaban, Jasmeen, Mahfouz, Magdy M..