Biology subjects

Schroeter, C.

Publications and source records attributed to Schroeter, C..

4 recordsLinked to original sources

bioRxiv2025

HM-DyadCap - Capture and Mapping of 5-Hydroxymethyl-cytosine/5-Methylcytosine CpG Dyads in Mammalian DNA

5-Methylcytosine (mC) and 5-hydroxymethylcytosine (hmC) are the main epigenetic modifications of mammalian DNA, and play crucial roles in cell differentiation, development, and tumorigenesis. Both modifications co-exist with unmodified cytosine in palindromic CpG dyads in different symmetric and asymmetric combinations across the two DNA strands, each having unique regulatory potential. To facilitate investigating the individual functions of such dyad modifications, we report HM-DyadCap. This method employs an evolved methyl-CpG-binding domain (MECP2 HM) for the direct capture and sequencing of DNA fragments containing the CpG dyad hmC/mC. Binding studies reveal a high discrimination of MECP2 HM against off-target dinucleotides. We conduct comparative mapping experiments for mESC genomes with HM-DyadCap, standard MethylCap employing wild type MECP2, as well as MeDIP and hMeDIP protocols. We find that MECP2 HM is blocked by hmC glucosylation, and conduct control enrichments with glucosylated genomes that indicate highly selective enrichment of hmC/mC dyads by MECP2 HM. Metagene profiles correlate hmC/mC marks with actively transcribed genes, and reveal global enrichment in gene bodies as well as depletion at transcription start sites. We anticipate that HM-DyadCap will enable effective enrichment and mapping of hmC/mC marks with broad applicability for unravelling the function of this dyad in chromatin biology and cancer.

genomics↗

bioRxiv2024

Welding PROxAb Shuttles: A modular approach for generating bispecific antibodies via site-specific protein-protein conjugation

Targeted protein degradation is an innovative therapeutic strategy to selectively eliminate disease-causing proteins. Exemplified by proteolysis-targeting chimeras (PROTACs), it has shown promise in overcoming drug resistance and targeting previously undruggable proteins. However, PROTACs face challenges such as low oral bioavailability and limited selectivity. The recently published PROxAb Shuttle technology offers a solution enabling targeted delivery of PROTACs using antibodies fused with PROTAC-binding domains derived from camelid single-domain antibodies (VHHs). Here, a modular approach to quickly generate PROxAb Shuttles by enzymatically coupling PROTAC-binding VHHs to off- the-shelf antibodies was developed. The resulting conjugates retained their target binding and internalization properties and incubation with BRD4-targeting PROTACs resulted in formation of defined PROxAb-PROTAC complexes. These complexes selectively induced degradation of the BRD4 protein, resulting in cytotoxicity specifically in cells expressing the antibodys target. The chemoenzymatic approach described here provides a versatile and efficient solution for generating antibody-VHH conjugates for targeted protein degradation applications but could also be used to combine antibody and VHH binders to generate bispecific antibodies for further applications.

biochemistry↗

bioRxiv2023

PROxAb Shuttle: A non-covalent plug-and-play platform for the rapid generation of tumor-targeting antibody-PROTAC conjugates

Proteolysis-targeting chimeras (PROTACs) have evolved in recent years from an academic idea to a therapeutic modality with more than 25 active clinical programs. However, achieving oral bioavailability and cell-type specificity remains a challenge, especially for PROTACs recruiting the von Hippel-Lindau (VHL) E3 ligase. Herein, we present an unprecedented, plug- and-play platform for VHL-recruiting PROTACs to overcome these limitations. Our platform allows for the generation of non-covalent antibody-PROTAC complexes within minutes and obviates the need for prior PROTAC modification, antibody-drug linker chemistry optimization or bioconjugation. Our technology relies on camelid-derived antibody domains (VHHs) which can easily be engineered into existing therapeutic antibody scaffolds. The resulting targeted, bispecific fusion proteins can be complexed with PROTACs at defined PROTAC-to-antibody ratios and have been termed PROxAb Shuttles. PROxAb Shuttles can prolong the half-life of PROTACs from hours to days, demonstrate anti-tumor efficacy in vivo and have the potential for reloading in vivo to further boost efficacy.

biochemistry↗

bioRxiv2019

Robustness and timing of cellular differentiation through population-based symmetry breaking

During mammalian development, cell types expressing mutually exclusive genetic markers are differentiated from a multilineage primed state. These observations have invoked single-cell multistability view as the dynamical basis of differentiation. However, the robust regulative nature of mammalian development is not captured therein. Considering the well-established role of cell-cell communication in this process, we propose a fundamentally different dynamical treatment in which cellular identities emerge and are maintained on population level, as a novel unique solution of the coupled system. Subcritical systems organization here enables symmetry-breaking to be triggered by cell number increase in a timed, self-organized manner. Robust cell type proportions are thereby an inherent feature of the resulting inhomogeneous solution. This framework is generic, as exemplified for early embryogenesis and neurogenesis cases. Distinct from mechanisms that rely on pre-existing asymmetries, we thus demonstrate that robustness and accuracy necessarily emerge from the cooperative behaviour of growing cell populations during development.

systems biology↗