DNA inter-strand crosslinks (ICLs) threaten genomic stability by creating a physical barrier to DNA replication and transcription. ICLs can be caused by endogenous reactive metabolites or from chemotherapeutics. ICL repair in humans depends heavily on the Fanconi Anaemia (FA) pathway. A key signalling step of the FA pathway is the mono-ubiquitination of Fanconi Anaemia Complementation Group D2 (FANCD2), which is achieved by the multi-subunit E3 ligase complex. FANCD2 mono-ubiquitination leads to the recruitment of DNA repair proteins to the site of the ICL. The loss of FANCD2 mono-ubiquitination is a common clinical feature of FA patient cells. Therefore, molecules that restore FANCD2 mono-ubiquitination could lead to a potential drug for the management of FA. On the other hand, in some cancers, FANCD2 mono-ubiquitination has been shown to be essential for cell survival. Therefore, inhibition of FANCD2 mono-ubiquitination represents a possible therapeutic strategy for cancer specific killing. We transferred an 11-protein FANCD2 mono-ubiquitination assay to a high-throughput format. We screened 9,067 compounds for both activation and inhibition of the E3 ligase complex. The use of orthogonal assays revealed that candidate compounds acted via non-specific mechanisms. However, our high-throughput biochemical assays demonstrate the feasibility of using sophisticated and robust biochemistry to screen for small molecules that modulate a key step in the FA pathway. The future identification of FA pathway modulators is anticipated to guide future medicinal chemistry projects with drug leads for human disease.
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http://dx.doi.org/10.1038/s41598-020-64868-7 | DOI Listing |
Cell
October 2024
Department of Pathology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA; Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA; Harold C. Simmons Comprehensive Cancer Center, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA. Electronic address:
Chromothripsis describes the catastrophic shattering of mis-segregated chromosomes trapped within micronuclei. Although micronuclei accumulate DNA double-strand breaks and replication defects throughout interphase, how chromosomes undergo shattering remains unresolved. Using CRISPR-Cas9 screens, we identify a non-canonical role of the Fanconi anemia (FA) pathway as a driver of chromothripsis.
View Article and Find Full Text PDFFront Endocrinol (Lausanne)
June 2024
College of Basic Medicine, Jining Medical University, Jining, China.
Non-obstructive azoospermia (NOA) is a disease characterized by spermatogenesis failure and comprises phenotypes such as hypospermatogenesis, mature arrest, and Sertoli cell-only syndrome. Studies have shown that FA cross-linked anemia (FA) pathway is closely related to the occurrence of NOA. There are FA gene mutations in male NOA patients, which cause significant damage to male germ cells.
View Article and Find Full Text PDFCell Rep
December 2023
Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, VA 22903, USA. Electronic address:
J Mol Biol
November 2023
Department of Genetics, Cell Biology and Development, Medical School, University of Minnesota at Twin Cities, Minneapolis, MN 55455, USA; Masonic Cancer Center, Minneapolis, MN 55455, USA. Electronic address:
Faithful genome duplication is a challenging task for dividing mammalian cells, particularly under replication stress where timely resolution of late replication intermediates (LRIs) becomes crucial prior to cell division. In human cancer cells, mitotic DNA repair synthesis (MiDAS) is described as a final mechanism for the resolution of LRIs to avoid lethal chromosome mis-segregation. RAD52-driven MiDAS achieves this mission in part by generating gaps/breaks on metaphase chromosomes, which preferentially occur at common fragile sites (CFS).
View Article and Find Full Text PDFMol Cell Biochem
July 2023
Department of Oncogene Regulation, Chittaranjan National Cancer Institute, 37 SPMukherjee Road, Kolkata, West Bengal, 700026, India.
We aimed to understand the crosstalk between mismatch repair (MMR) and FA-BRCA pathway in primary bladder carcinoma (BlCa) samples as well as in chemotolerant cell line. We analysed the genetic alterations of MLH1 and MSH2 (MMR-related genes) and after that we correlated it with the nuclear translocation of FANCD2 protein. Next, we evaluated this crosstalk in T24 BlCa cell line in response to doxorubicin treatment.
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