Biology subjects

Blank, M.

Publications and source records attributed to Blank, M..

4 recordsLinked to original sources

bioRxiv2025

SMURF2/USP7-mediated ubiquitination of KAP1 controls its SUMO E3 ligase activity and chromatin regulation

KRAB-associated protein 1 (KAP1) is a critical nuclear protein that regulates chromatin architecture and gene expression, primarily through its SUMOylation activity. However, the mechanisms controlling KAP1 remain poorly understood. In a recent study, we identified the E3 ubiquitin ligase SMURF2 as a direct interactor and ubiquitin ligase for KAP1. Here, we show that SMURF2-mediated ubiquitination of KAP1 at lysines K254, K319, and K779 regulates its SUMOylation activity. We further demonstrate that SMURF2 collaborates with the deubiquitinase USP7/HAUSP, recruiting it to KAP1, to regulate a dynamic ubiquitination-deubiquitination cycle at these key residues. Cells expressing a KAP1 mutant, resistant to SMURF2-USP7 regulation, exhibit profound alterations in chromatin structure, gene expression, protein-protein interactions, and elevated LINE-1 retrotransposon activity. Notably, disrupting the SMURF2-USP7-KAP1-SUMOylation axis in cancer cells reduces their malignancy and tumor growth in vivo. Collectively, these findings uncover a novel regulatory mechanism for KAP1, highlighting its pivotal role in orchestrating critical cellular processes.

molecular biology↗

bioRxiv2024

Unbiased functional genetic screens reveal essential RNA modifications in human cancer and drug resistance

RNA modification pathways are mis-regulated in multiple types of human cancer. To comprehensively identify cancer-relevant RNA modifications and their regulators, we screened all 150 annotated human RNA modifying proteins across 18 different normal and cancer cell lines using a CRISPR-based genetic knockout system. Fifty RNA modifying proteins were essential for survival of at least one cell type. A third of these essential genes were amplified in 38 different human primary cancer types and potentially drive cancer growth. Unexpectedly, the number of essential genes per cell line varied considerably, and this variation was not due to tissue of origin. Instead, we found that cancer cell-specific mitochondrial metabolic plasticity was responsible for the unique requirement of certain RNA modifications. For example, leukemia cells with high intrinsic drug tolerance required mitochondrial flexibility to survive treatment with the anti-leukemic drugs cytarabine and venetoclax. Synthetic lethality screens revealed that drug-resistance is abolished by deleting the mitochondrial methyltransferase TRMT5, which is responsible for the formation of N1-methylguanosine (m1G) in the tRNA anticodon loop. In summary, our study identifies cancer-relevant RNA modifying enzymes, and reveals a novel promising drug target for therapy-resistant acute myeloid leukemia.

cell biology↗

bioRxiv2023

High yield, low magnesium flexizyme reactions in a water-ice eutectic phase

Flexizymes enable the stoichiometric acylation of tRNAs with a variety of compounds, enabling the in vitro translation of peptides with both non-natural backbones and side chains. However, flexizyme reactions have several drawbacks, including single-turnover kinetics, high Mg(II) carryover inhibiting in vitro translation, and rapid product hydrolysis. Here we present flexizyme reactions utilizing an ice-eutectic phase, with high yields, 30X lower Mg(II), and long-term product stability. The eutectic flexizyme reactions increase the ease of use and flexibility of flexizyme aminoacylation, and increase the in vitro protein production.

synthetic biology↗

bioRxiv2022

The effect of carbamazepine on bone structure and strength in control and Osteogenesis Imperfecta (Col1a2 +/p.G610C) mice

The inherited brittle bone disease osteogenesis imperfecta (OI) is commonly caused by COL1A1 and COL1A2 mutations that disrupt the collagen I triple helix. This causes intracellular endoplasmic reticulum (ER) retention of the misfolded collagen and can result in a pathological ER stress response. A therapeutic approach to reduce this toxic mutant load could be to stimulate mutant collagen degradation by manipulating autophagy and/or ER-associated degradation. Since carbamazepine (CBZ) both stimulates autophagy of misfolded collagen X and improves skeletal pathology in a metaphyseal chondrodysplasia model, we tested the effect of CBZ on bone structure and strength in 3 week-old male OI Col1a2 +/p.G610C and control mice. Treatment for 3 or 6 weeks with CBZ, at the dose effective in metaphyseal chondrodysplasia, provided no therapeutic benefit to Col1a2 +/p.G610C mouse bone structure, strength, or composition, measured by micro-computed tomography, three point bending tests and Fourier-transform infrared microspectroscopy. In control mice however, CBZ treatment for 6 weeks impaired femur growth and led to lower femoral cortical and trabecular bone mass. These data, showing the negative impact of CBZ treatment on the developing mouse bones, raise important issues which must be considered in any human clinical applications of CBZ in growing individuals.

pathology↗