Biology subjects

Schatz, C.

Publications and source records attributed to Schatz, C..

3 recordsLinked to original sources

bioRxiv2025

Dual E3 ligase recruitment by monovalent degraders enables redundant and tuneable degradation of SMARCA2/4

Proteolysis-Targeting Chimeras (PROTACs) and Molecular Glue Degraders (MGDs) canonically target proteins for degradation by recruiting them to a single E3 ligase complex. While heterotrivalent PROTACs that can co-opt multiple E3 ligase complexes have been described, to our knowledge all MGDs reported to date are dependent on a single E3. Here, using orthogonal genetic screening, biophysical and structural analyses, we show that a monovalent MGD can covalently recruit CUL4DCAF16 and CRL1FBXO22 in a parallel and redundant manner to degrade SMARCA2/4. Deep mutational scanning identifies a single cysteine (Cys173) in DCAF16 essential for degrader activity, and intact protein MS confirms covalent adduct at this site. The cryo-EM structure of the DCAF16:SMARCA2:degrader ternary complex reveals a unique binding mode and a distinct interface of neo-interactions, providing insights into degrader specificity. We demonstrate that E3 ligase dependency can be tuned both chemically and genetically. Minimal alterations to the compounds "degradation tail" switches ligase preference from DCAF16 to FBXO22, while a single L59W mutation on DCAF16 is sufficient to drive DCAF16 engagement for otherwise FBXO22-dependent compounds. These results establish a molecular and structural framework for the design of tuneable dual glue degraders that could mitigate challenges from resistance mechanisms in degrader therapies.

biochemistry↗

bioRxiv2024

Insight Into the Molecular Parameters of PEI Promoting an Efficient Gene Delivery into Cells

The development of natural or synthetic polycations able to interact with nucleic acids and condense them into nanoparticles known as polyplexes, faces several unresolved challenges at the cellular level. Key issues include the intracellular trafficking of polyplexes, the endosomal escape and the release of nucleic acids into the cytosol, which are considered major bottlenecks for efficient protein expression. Here, we aim at gaining fundamental insights into the stability of polyplexes in biological media and their uptake and intracellular trafficking, while correlating data of the expression of reporter protein with both the molecular characteristics of various poly(ethylenimines) (PEI) and the physicochemical characteristics of PEI/peGFP-C3 polyplexes. For this, we chosen four samples of PEI, selected as a model polycation, with different molecular weights (Mw = 0.8, 20, 25 and 60 kg/mol) and structures (linear and branched). We found that the in vitro and in vivo stability of PEI/peGFP-C3 polyplexes, their cell internalization and transfection efficiency is dependent on the variation of polycation Mw and structure, as well as the intrinsic properties of polyplexes, such as the charge ratio (R=[N+]/[P-]). A relation between the percentage of positive cells to green fluorescent protein (GFP) and the amount of internalized nucleic acid (cyanine 5-peGFP-C3) allowed revealing the molecular characteristics of PEI promoting both higher both cell internalization and GFP expression on HEK293T cells. In the long term, the outcome of this work will be to propose guidelines to help design more effective, and less cytotoxic non-viral gene carriers with a great potential for new therapeutic applications.

biophysics↗

bioRxiv2023

Comparative Analysis of Whole Transcriptome Single-Cell Sequencing Technologies in Complex Tissues

The development of single-cell omics tools has enabled scientists to study the tumor microenvironment (TME) in unprecedented detail. However, each of the different techniques may have its unique strengths and limitations. Here we directly compared two commercially available high-throughput single-cell RNA sequencing (scRNA-seq) technologies - droplet-based 10X Chromium vs. microwell-based BD Rhapsody - using paired samples from patients with localized prostate cancer (PCa) undergoing a radical prostatectomy. Although high technical consistency was observed in unraveling the whole transcriptome, the relative abundance of cell populations differed. Cells with low-mRNA content such as T cells were underrepresented in the droplet-based system, at least partly due to lower RNA capture rates. In contrast, microwell based scRNA-seq recovered less cells of epithelial origin. Moreover, we discovered platform-dependent variabilities in mRNA quantification and cell-type marker annotation. Overall, our study provides important information for selection of the appropriate scRNA-seq platform and for the interpretation of published results. SYNOPSISO_LIComparison of scRNA-seq protocols uncovers disparities in RNA-to-library conversion C_LIO_LIMicrowell-based scRNA-seq technology excels in capturing low-mRNA content cells C_LIO_LIBiased transcriptomes due to gene specific RNA detection efficacies by both platforms C_LIO_LIThe study guides in informed scRNA-seq platform selection and data interpretation C_LI

cancer biology↗