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Published on: March 31, 2019
Structure-function relationships explain CTCF zinc finger mutation phenotypes in cancer
Charles G Bailey1,2,3, Shailendra Gupta4,5, Cynthia Metierre1,2
1Cancer and Gene Regulation Laboratory Centenary Institute, The University of Sydney, Camperdown, NSW, 2050, Australia.
Abstract:
CCCTC-binding factor (CTCF) plays fundamental roles in transcriptional regulation and chromatin architecture maintenance. CTCF is also a tumour suppressor frequently mutated in cancer, however, the structural and functional impact of mutations have not been examined. We performed molecular and structural characterisation of five cancer-specific CTCF missense zinc finger (ZF) mutations occurring within key intra- and inter-ZF residues. Functional characterisation of CTCF ZF mutations revealed a complete (L309P, R339W, R377H) or intermediate (R339Q) abrogation as well as an enhancement (G420D) of the anti-proliferative effects of CTCF. DNA binding at select sites was disrupted and transcriptional regulatory activities abrogated. Molecular docking and molecular dynamics confirmed that mutations in residues specifically contacting DNA bases or backbone exhibited loss of DNA binding. However, R339Q and G420D were stabilised by the formation of new primary DNA bonds, contributing to gain-of-function. Our data confirm that a spectrum of loss-, change- and gain-of-function impacts on CTCF zinc fingers are observed in cell growth regulation and gene regulatory activities. Hence, diverse cellular phenotypes of mutant CTCF are clearly explained by examining structure-function relationships.
Insights
Cancer-associated mutations in CCCTC-binding factor (CTCF) zinc fingers can disrupt DNA binding and gene regulation. Some mutations lead to loss-of-function, while others cause gain-of-function, explaining diverse cellular effects.
Area of Science:
- Molecular biology
- Structural biology
- Cancer genetics
Background:
- CCCTC-binding factor (CTCF) is crucial for gene regulation and chromatin structure.
- CTCF mutations are frequent in cancer, but their structural and functional impacts are poorly understood.
Purpose of the Study:
- To investigate the molecular and structural effects of cancer-specific CTCF zinc finger mutations.
- To correlate these structural changes with functional alterations in DNA binding, gene regulation, and cell proliferation.
Main Methods:
- Molecular and structural characterization of five cancer-associated CTCF missense zinc finger mutations.
- Functional assays assessing anti-proliferative effects, DNA binding affinity, and transcriptional regulation.
- Molecular docking and molecular dynamics simulations to analyze mutation impacts on protein-DNA interactions.
Main Results:
- Mutations L309P, R339W, and R377H completely abrogated CTCF's anti-proliferative effects, while R339Q showed intermediate abrogation and G420D enhanced it.
- DNA binding at specific sites was disrupted, leading to abrogated transcriptional regulation for some mutants.
- Molecular dynamics revealed that mutations affecting direct DNA contact caused loss-of-function, whereas R339Q and G420D gained stability via new DNA bonds.
Conclusions:
- Cancer-associated CTCF zinc finger mutations exhibit a range of functional impacts, including loss-, change-, and gain-of-function.
- These diverse effects on cell growth and gene regulation are explained by structure-function relationships.
- Understanding these mutation impacts is key to explaining diverse cellular phenotypes in mutant CTCF-driven cancers.
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