A5060073964- CRISPR and Genetic Engineering
- Cytomegalovirus and herpesvirus research
- DNA Repair Mechanisms
- Virus-based gene therapy research
- Mosquito-borne diseases and control
- RNA regulation and disease
- PARP inhibition in cancer therapy
- BRCA gene mutations in cancer
- Genetics, Aging, and Longevity in Model Organisms
- RNA Interference and Gene Delivery
- Cancer-related Molecular Pathways
- Ubiquitin and proteasome pathways
- Genetic factors in colorectal cancer
- HIV Research and Treatment
- Protein Degradation and Inhibitors
University of Massachusetts Chan Medical School
2019-2024
Worcester State University
2022
Dana-Farber Cancer Institute
2018-2020
Abstract Prime editors (PEs) mediate genome modification without utilizing double-stranded DNA breaks or exogenous donor as a template. PEs facilitate nucleotide substitutions local insertions deletions within the based on template sequence encoded prime editing guide RNA (pegRNA). However, efficacy of in adult mice has not been established. Here we report an NLS-optimized SpCas9-based editor that improves efficiency both fluorescent reporter cells and at endogenous loci cultured cell lines....
Type V CRISPR-Cas12a systems provide an alternate nuclease platform to Cas9, with potential advantages for specific genome editing applications. Here we describe improvements the Cas12a system that facilitate efficient targeted mutagenesis in mammalian cells and zebrafish embryos. We show engineered variants of two different nuclear localization sequences (NLS) on C terminus increased efficiency cells. Additionally, find pre-crRNAs comprising a full-length direct repeat (full-DR-crRNA)...
Abstract Prime editing systems have enabled the incorporation of precise edits within a genome without introducing double strand breaks. Previous studies defined an optimal primer binding site (PBS) length for pegRNA ∼13 nucleotides depending on sequence composition. However, PBS characterization has been based prime outcomes using plasmid or lentiviral expression systems. In this study, we demonstrate that editor (PE) ribonucleoprotein complexes, auto-inhibitory interaction between and...
Type V CRISPR–Cas12a systems are an attractive Cas9-alternative nuclease platform for specific genome-editing applications. However, previous studies demonstrate that there is a gap in overall activity between Cas12a and Cas9 primary cells. Here we describe optimization to the nuclear localization signal (NLS) composition architecture of facilitate highly efficient targeted mutagenesis human transformed cell lines (HEK293T, Jurkat, K562 cells) cells (natural killer [NK] CD34+ hematopoietic...
Abstract Prime editing efficiency is modest in cells that are quiescent or slowly proliferating where intracellular dNTP levels tightly regulated. MMLV-reverse transcriptase - the prime editor polymerase subunit requires high dNTPs for efficient polymerization. We report primary and vivo increased by mutations enhance enzymatic properties of can be further complemented targeting SAMHD1 degradation.
Abstract Prime editors (PEs) mediate genome modification without utilizing double-stranded DNA breaks or exogenous donor as a template. PEs facilitate nucleotide substitutions local insertions deletions within the based on template sequence encoded prime editing guide RNA (pegRNA). However, efficacy of in adult mice has not been established. Here we report an NLS-optimized SpCas9-based editor that improves efficiency both fluorescent reporter cells and at endogenous loci cultured cell lines....
Abstract Type V CRISPR–Cas12a systems are an attractive alternative nuclease platform for specific genome editing applications. However, previous studies demonstrate that there is a gap in overall activity between Cas12a and Cas9 primary cells. Here we describe optimization to the nuclear localization signal composition architecture of facilitate highly efficient targeted mutagenesis mammalian cell lines (HEK293T, Jurkat, K562 cells) cells (NK CD34+ HSPCs), regardless ortholog. A 3xNLS...
Abstract Homologous-recombination (HR) deficient tumors with BRCA1 and BRCA2 mutations exhibit replication fork stability defects. To date, PARP inhibitors are the only targeted therapy available in clinic against HR tumors, alternative therapies needed. In this study, we found a deubiquitinase, USP1, to be significantly upregulated BRCA1. SiRNA mediated silencing or small molecule inhibition of USP1 activity resulted destabilization decreased viability cells, revealing synthetic lethal...