Roles of origin recognition complex subunit 6 (Orc6) in DNA replication and damage response

Well-regulated replication and timely repair of DNA is required for genome stability maintenance, and failing to do so leads to tumorigenesis. This thesis will focus on how human Orc6, the smallest subunit of Origin Recognition Complex (ORC) which shows dispensable role in origin licensing, functions in DNA replication progression and regulates chromatin remodeler binding. In Chapter 1, I review the following three aspects: 1) protein complexes involved in DNA replication licensing and initiation, represented by ORC; 2) DNA damage response regulated by ATM/ATR-signaling; 3) ATP-dependent chromatin remodeling complexes and their association with DNA replication or repair. In Chapter 2, I investigate the functional relevance of Orc6-T229 phosphorylation in replication fork progression and cancer cell migration & invasion. Orc6-pT229 is an S-phase-specific, oxidative damage-triggered and ATR-dependent modification. I further confirm that Orc6-pT229 localizes at replication forks and reduces fork progression rate. Cell cycle progression upon release from oxidative damage is impacted with the presence of the Orc6-T229 phosphomimic mutant. I therefore propose that Orc6-pT229 is a replication fork pausing signal. Further, I showed that Orc6-pT229 as a signal of tumor suppression since it was lacking from colorectal cancer tissue, and the phosphomimic mutant reduced the tumorigenic behaviors (such as migration and invasion) of cancer cell lines. A pan-cancer analysis further revealed the high Orc6 expressions in various cancers. In Chapter 3, I further investigate the role of Orc6 upon oxidative damage. The genomic binding of Orc6 was characterized using CUT&RUN method, and I found that Orc6 was enriched at open chromatin regions and at active replication origins. Oxidative damage leads to a more widespread genomic binding of Orc6. Interestingly, I noticed that Orc6, but not other ORC components, shows potential of association with high-molecular-weight, non-ORC protein complexes. With proteomic method, I identified and characterized the S-phase specific, oxidative damage-triggered interaction between Orc6 and ISWI family chromatin remodeler SMARCA1. Genomic mapping of SMARCA1 showed that when Orc6 is depleted, SMARCA1 binds to original Orc6-enriched sites, and this causes an increase of local chromatin accessibility. Low Orc6 presence also facilitates the remodeling of SMARCA1, while increased Orc6 presence may negatively regulate it. The chromatin remodeling at Orc6-depleted sites could correlate with local DNA damage, active replication fork/origin remodeling or dormant origin firing, which requires further investigation. Finally, in Chapter 4, I concluded my work in Chapters 2 & 3 and discussed the gaps in knowledge and limitations of the work. Meanwhile, I discussed some connections between these two chapters.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Cell and Developmental Biology

Grantor

University of Illinois Urbana-Champaign

Year dc:date

2025

Author dc:creator

• Liu, Dazhen

Contributors dc:contributor

• Prasanth, Supriya
• Belmont, Andrew S
• Chen, Jie
• Madak-Erdogan, Zeynep

• Dna Replication
• Dna Damage
• Orc6
• Chromatin Remodeling
• Iswi
• Smarca1

Statement dc:rights

• Copyright 2025 Dazhen Liu

Language dc:language

en, eng

Handle dc:identifier

https://hdl.handle.net/2142/130142

OAI record oai:identifier

https://www.ideals.illinois.edu/oai-pmh?verb=GetRecord&metadataPrefix=etdms&identifier=oai%3Awww.ideals.illinois.edu%3A2142%2F130142