Medicine and Health Sciences Commons^™

Genetic Separation Of Brca1 Functions Reveal Mutation-Dependent Polθ Vulnerabilities, John J. Krais, David J. Glass, Ilse Chudoba, Yifan Wang, Wanjuan Feng, Dennis Simpson, Pooja Patel, Zemin Liu, Ryan Neumann-Domer, Robert G. Betsch, Andrea J. Bernhardy, Alice M. Bradbury, Jason Conger, Wei-Ting Yueh, Joseph Nacson, Richard T. Pomerantz, Gaorav P. Gupta, Joseph R. Testa, Neil Johnson

Department of Biochemistry and Molecular Biology Faculty Papers

Homologous recombination (HR)-deficiency induces a dependency on DNA polymerase theta (Polθ/Polq)-mediated end joining, and Polθ inhibitors (Polθi) are in development for cancer therapy. BRCA1 and BRCA2 deficient cells are thought to be synthetic lethal with Polθ, but whether distinct HR gene mutations give rise to equivalent Polθ-dependence, and the events that drive lethality, are unclear. In this study, we utilized mouse models with separate Brca1 functional defects to mechanistically define Brca1-Polθ synthetic lethality. Surprisingly, homozygous Brca1 mutant, Polq−/− cells were viable, but grew slowly and had chromosomal instability. Brca1 mutant cells proficient in DNA end resection were …

Candidate Variants In Dna Replication And Repair Genes In Early-Onset Renal Cell Carcinoma Patients Referred For Germline Testing, Elena V. Demidova, Ilya G. Serebriiskii, Ramilia Vlasenkova, Simon Kelow, Mark D. Andrake, Tiffiney R. Hartman, Tatiana Kent, James Virtucio, Gail L. Rosen, Richard T. Pomerantz, Roland L. Dunbrack, Erica A. Golemis, Michael J. Hall, David Y.T. Chen, Mary B. Daly, Sanjeevani Arora

Department of Biochemistry and Molecular Biology Faculty Papers

Background: Early-onset renal cell carcinoma (eoRCC) is typically associated with pathogenic germline variants (PGVs) in RCC familial syndrome genes. However, most eoRCC patients lack PGVs in familial RCC genes and their genetic risk remains undefined.

Methods: Here, we analyzed biospecimens from 22 eoRCC patients that were seen at our institution for genetic counseling and tested negative for PGVs in RCC familial syndrome genes.

Results: Analysis of whole-exome sequencing (WES) data found enrichment of candidate pathogenic germline variants in DNA repair and replication genes, including multiple DNA polymerases. Induction of DNA damage in peripheral blood monocytes (PBMCs) significantly elevated numbers of …

Semi-Quantitative Detection Of Pseudouridine Modifications And Type I/Ii I/Ii Hypermodifications In Human Mrnas Using Direct Long-Read Sequencing, Sepideh Tavakoli, Mohammad Nabizadeh, Amr Makhamreh, Howard Gamper, Caroline A Mccormick, Neda K Rezapour, Ya-Ming Hou, Meni Wanunu, Sara H Rouhanifard

Department of Biochemistry and Molecular Biology Faculty Papers

Here, we develop and apply a semi-quantitative method for the high-confidence identification of pseudouridylated sites on mammalian mRNAs via direct long-read nanopore sequencing. A comparative analysis of a modification-free transcriptome reveals that the depth of coverage and specific k-mer sequences are critical parameters for accurate basecalling. By adjusting these parameters for high-confidence U-to-C basecalling errors, we identify many known sites of pseudouridylation and uncover previously unreported uridine-modified sites, many of which fall in k-mers that are known targets of pseudouridine synthases. Identified sites are validated using 1000-mer synthetic RNA controls bearing a single pseudouridine in the center position, demonstrating systematic …