Biology subjects

Witus, S. R.

Publications and source records attributed to Witus, S. R..

5 recordsLinked to original sources

bioRxiv2026

Systemic degradation of repressive transcription factors gates gene expression and cell fate specification

While proteasomes are best known for eliminating defective proteins or turning off signaling pathways, they also enable crucial cellular activities. Critical among these, proteasomes allow cells to initiate gene expression, but underlying targets and regulatory mechanisms remain poorly understood. Here, we report that proteasomes drive the systemic degradation of repressive trans-cription factors to eject TLE/Groucho-family co-repressors from chromatin and thereby constantly free transcription start sites for activator binding. This circuitry requires the E3 ligase SCFFBXL14, which modifies its targets dependent on presentation by TLEs, but independently of their identity. The continuous cycling of co-repressors off chromatin, as achieved by systemic turnover of a protein family, is essential for stem cells to translate developmental cues into lineage-specific gene expression, and it is disrupted by cancer mutations in TLE1 that impair SCFFBXL14-recruit-ment. We conclude that systemic degradation of repressive transcription factors establishes co-repressor dynamics required for genes expression and cell fate specification.

biochemistry↗

bioRxiv2026

Metabolic glues as a means of purine sensing and chemotherapeutic response

Molecular glues stabilize weak interactions to impart novel functionalities onto complexes. While plant hormones or drugs are known molecular glues, it is still unknown whether this modality provides endogenous regulation in human cells. Here, we show that purine nucleotides are molecular glues that tether the rate-limiting enzyme of purine biosynthesis, phosphoribosyl-pyrophosphate-amidotransferase (PPAT), to its inhibitor NUDT5. This mechanism allows cells to sense purine levels and establish essential feedback control of their synthesis. Thiopurine chemotherapeutics, in clinical use since the 1950s, act as molecular glues of the same complex, but adopt unique orientations for enhanced function. Distinct from the recognition of many therapeutic glues, metabolic glue pockets can adjust their conformation to significant compound alterations and thereby enable increasing glue potency without sacrificing specificity. Our findings therefore identify endogenous metabolic glues as a mode of nutrient sensing that can be exploited to obtain compounds that rewire metabolic pathways for therapeutic benefit.

biochemistry↗

bioRxiv2024

The molecular basis of integrated stress response silencing

Chronic stress response activation impairs cell survival and causes devastating degenerative diseases. To counteract this, cells deploy dedicated silencing factors, such as the E3 ligase SIFI that terminates the mitochondrial stress response. How a single enzyme can sense stress across cells and elicit timely stress response inactivation is poorly understood. Here, we report the structure of human SIFI, which revealed how this 1.3MDa complex can target hundreds of proteins for accurate stress response silencing. SIFI attaches the first ubiquitin to substrates using flexible domains within an easily accessible scaffold, yet builds linkage-specific ubiquitin chains at distinct, sterically restricted elongation modules in its periphery. Ubiquitin handover via a ubiquitin-like domain couples versatile substrate modification to precise chain elongation. Stress response silencing therefore exploits a catalytic mechanism that is geared to process many diverse proteins and hence allows a single enzyme to monitor and, if appropriate, modulate a complex cellular state.

biochemistry↗

bioRxiv2024

Force transmission through the inner kinetochore is enhanced by centromeric DNA sequences

Previously, we reconstituted a minimal functional kinetochore from recombinant Saccharomyces cerevisiae proteins that was capable of transmitting force from dynamic microtubules to nucleosomes containing the centromere-specific histone variant Cse4 (Hamilton et al. 2020). This work revealed two paths of force transmission through the inner kinetochore: through Mif2 and through the Okp1/Ame1 complex (OA). Here, using a chimeric DNA sequence that contains crucial centromere-determining elements of the budding yeast point centromere, we demonstrate that the presence of centromeric DNA sequences in Cse4-containing nucleosomes significantly strengthens OA-mediated linkages. Our findings indicate that centromeric sequences are important for the transmission of microtubule-based forces to the chromosome.

biochemistry↗

bioRxiv2022

Multivalent BARD1-nucleosome interactions facilitate H2A ubiquitylation by BRCA1/BARD1

BRCA1/BARD1 is a tumor suppressor E3 ubiquitin (Ub) ligase with roles in DNA damage repair and in transcriptional regulation. BRCA1/BARD1 RING domains interact with nucleosomes to facilitate mono-ubiquitylation of distinct residues on the C-terminal tail of histone H2A. These enzymatic domains constitute a small fraction of the heterodimer, raising the possibility of functional chromatin interactions involving other regions such as the BARD1 C-terminal domains that bind nucleosomes containing the DNA damage signal H2A K15-Ub and H4 K20me0, or portions of the expansive intrinsically disordered regions found in both subunits. Herein, we reveal novel interactions that support robust H2A ubiquitylation activity mediated through a high-affinity, intrinsically disordered DNA-binding region of BARD1. These interactions support BRCA1/BARD1 recruitment to chromatin and sites of DNA damage in cells and contribute to their survival. We also reveal distinct BRCA1/BARD1 complexes that depend on the presence of H2A K15-Ub, including a complex where a single BARD1 subunit spans adjacent nucleosome units. Our findings identify an extensive network of multivalent BARD1- nucleosome interactions that serve as a platform for BRCA1/BARD1-associated functions on chromatin.

biochemistry↗