Biology subjects

Woodruff, K. A.

Publications and source records attributed to Woodruff, K. A..

5 recordsLinked to original sources

bioRxiv2026

Dfm1 promotes Ste24-dependent translocon quality control

Proteins stalled during endoplasmic reticulum translocation clog Sec61 channels and require translocon quality control (TQC) mechanisms for clearance. Genetic screens implicated the derlin Dfm1 in TQC; however, its mechanistic role remained unclear. Using engineered and disease-relevant translocon-clogging substrates in Saccharomyces cerevisiae, we show Dfm1 functions in the Ste24-dependent branch of TQC, but not Hrd1- or Ltn1-dependent pathways. Loss of DFM1 increased accumulation and toxicity of translocon-clogging proteins. Increased dosage of wild type or catalytically inactive Hrd1 or Ste24 rescued toxicity, supporting a non-enzymatic buffering function for these factors in TQC. These findings identify Dfm1 as a key TQC factor whose critical role in clearing clogged translocons can be suppressed in multiple ways. Our results suggest enhancing TQC may protect cells from proteotoxic stress caused by disease-relevant translocon-clogging substrates, with potential implications for pancreatic {beta}-cell dysfunction in diabetes. Significance StatementO_LICells rely on translocon quality control (TQC) to clear proteins that clog the Sec61 translocon, but how the derlin Dfm1 contributes to this process has remained unclear. C_LIO_LIWe demonstrate that Dfm1 functions specifically in the Ste24-dependent TQC pathway and show that increased levels of the TQC factors Hrd1 or Ste24 rescues Dfm1 deficiency even without catalytic activity, revealing an unexpected non-enzymatic buffering function. C_LIO_LIThis work defines Dfm1s mechanistic role in TQC, provides new insight into how cells preserve endoplasmic reticulum function during translocation stress, and establishes a framework for investigating conserved pathways that protect against diseases associated with translocon clogging. C_LI

cell biology↗

bioRxiv2025

Depletion of microenvironmental syndecan-2 impairs hematopoietic stem cell self-renewal and cytokine responses

Syndecan-2 is a heparan sulfate proteoglycan highly enriched on murine bone marrow hematopoietic stem cells (HSCs) compared to terminally differentiated hematopoietic cells. Syndecan-2 binds growth factors via its heparan sulfate glycosaminoglycan chains to coordinate cell signaling. Knockdown of syndecan-2 reduces HSC self-renewal ability and promotes cell cycling via Cdkn1c. In this study, we analyzed the function of syndecan-2 expressed by bone marrow niche cells in hematopoiesis and HSC self-renewal. We determined that syndecan-2 is highly expressed by bone marrow mesenchymal stromal cells and moderately expressed by endothelial cells. To test the function of niche-expressed syndecan-2 in hematopoiesis, we generated transgenic mice depleted of Sdc2 in Lepr-targeted mesenchymal stromal cells (Sdc2{Delta}MSC mice) or Cdh5-targeted endothelial cells (Sdc2{Delta}EC mice). Loss of syndecan-2 from endothelial or mesenchymal stromal cells did not change bone marrow HSC frequencies or numbers. However, depletion of syndecan-2 from Lepr-targeted mesenchymal stromal cells, but not Cdh5-targeted endothelial cells, diminishes HSC self-renewal ability analyzed by competitive transplants into lethally irradiated mice. Ex vivo studies further show that HSCs co-cultured with HS-5 stromal cells depleted of SDC2 exhaust more rapidly than HSCs cultured with control HS-5 cells. Single-cell RNA sequencing analyses reveal that the depletion of Sdc2 from mesenchymal stromal cells significantly remodels the HSC transcriptome by enriching for pathways associated with excessive growth factor signaling. Together, our findings suggest that HSC self-renewal is supported by cell-extrinsic mechanisms enacted by syndecan-2 from the MSC niche, highlighting the importance of the niche proteoglycome in HSC functions. KEY POINTSO_LIThe heparan sulfate proteoglycan syndecan-2 expressed by mesenchymal stromal cells but not endothelial cells regulates HSC self-renewal C_LIO_LIDepletion of syndecan-2 from Lepr-targeted mesenchymal stromal cells remodels the hematopoietic stem cell transcriptional landscape C_LI

cell biology↗

bioRxiv2025

The bone marrow niche and hematopoietic system are distinctly remodeled by CD45-targeted astatine-211 radioimmunotherapy

Radioimmunotherapy (RIT) is used to treat patients with hematological malignancies known to infiltrate the bone marrow (BM) microenvironment. RIT uses target-specific monoclonal antibodies stably conjugated to radionuclides to deliver cytotoxic radiation to cells of interest. While RIT is effective at delivering radiation to cancer cells, normal tissue is also exposed to radiation upon RIT, the consequences of which are largely unknown. Here, we studied the cellular and molecular effects of CD45-targeted astatine-211 (211At) RIT, IgG non-targeted 211At RIT, and Cesium-137 total-body irradiation (TBI) on hematopoietic cells and their BM niche in wild-type immunocompetent mice. Relative to non-targeted RIT or TBI, CD45-targeted RIT significantly delayed hematopoietic regeneration overall in the peripheral blood and BM and reduced hematopoietic stem/progenitor cell recovery and colony-forming ability. While BM endothelial cells (ECs) do not express the CD45 antigen, CD45-targeted RIT significantly depleted BM ECs compared to non-targeted RIT or TBI. RNA sequence analysis revealed significantly different transcriptomic profiles of BM ECs from CD45-RIT-treated mice compared to non- targeted RIT or TBI. ECs from CD45-RIT-treated mice, but not TBI or IgG-RIT-treated mice, were transcriptionally enriched for TGF{beta}, NOTCH, and IFN signaling pathways compared to untreated mice. Collectively, our study indicates that CD45-targeted RIT severely impacts hematopoietic and EC niche recovery compared to non- targeted approaches. Future studies are required to determine the long-term consequences of such RIT-driven effects on BM niche physiology and how BM niche reprogramming by RIT affects cancer cells. KEY POINTSO_LICD45-targeted radioimmunotherapy more effectively suppresses the hematopoietic system than non- targeted radiation delivery. C_LIO_LIThe bone marrow vascular niche is differentially reprogrammed by CD45-targeted radioimmunotherapy compared to non-targeted radiation delivery. C_LI

cell biology↗

bioRxiv2023

"Important enough to show the world": Using Authentic Research Opportunities and Micropublications to Build Students' Science Identities

Primarily undergraduate institutions (PUIs) often struggle to provide authentic research opportunities that culminate in peer-reviewed publications due to "recipe-driven" lab courses and the comprehensive body of work necessary for traditional scientific publication. However, the advent of short-form, single-figure "micropublications" has created novel opportunities for early-career scientists to make and publish authentic scientific contributions on a scale and in a timespan compatible with their training periods. The purpose of this qualitative case study is to explore the benefits accrued by eight undergraduate and masters students who participated in authentic, small-scale research projects and disseminated their work as coauthors of peer-reviewed micropublications at a PUI. In these interviews, students reported that through the process of conducting and publishing their research, they developed specific competencies: reading scientific literature, proposing experiments, and collecting/interpreting publication-worthy data. Further, they reported this process enabled them to identify as contributing members of the greater scientific community.

scientific communication and education↗

bioRxiv2022

Lipid Biosynthesis Perturbation Impairs Endoplasmic Reticulum-Associated Degradation

The relationship between lipid homeostasis and protein homeostasis (proteostasis) is complex and remains incompletely understood. We conducted a screen for genes required for efficient degradation of Deg1-Sec62, a model aberrant translocon-associated substrate of the endoplasmic reticulum (ER) ubiquitin ligase Hrd1, in Saccharomyces cerevisiae. This screen revealed that INO4 is required for efficient Deg1-Sec62 degradation. INO4 encodes one subunit of the Ino2/Ino4 heterodimeric transcription factor, which regulates expression of genes required for lipid biosynthesis. Deg1-Sec62 degradation was also impaired by mutation of genes encoding several enzymes mediating phospholipid and sterol biosynthesis. The degradation defect in ino4{Delta} yeast was rescued by supplementation with metabolites whose synthesis and uptake are mediated by Ino2/Ino4 targets. Stabilization of a panel of substrates of the Hrd1 and Doa10 ER ubiquitin ligases by INO4 deletion indicates ER protein quality control is generally sensitive to perturbed lipid homeostasis. Further, loss of INO4 sensitized yeast to proteotoxic stress, suggesting a broad requirement for lipid homeostasis in maintaining proteostasis. Abundance of the ER ubiquitin-conjugating enzyme Ubc7 was reduced in the absence of INO4, consistent with a model whereby perturbed lipid biosynthesis alters the abundance of critical protein quality control mediators, with broad consequences for ER proteostasis. A better understanding of the dynamic relationship between lipid homeostasis and proteostasis may lead to improved understanding and treatment of several human diseases associated with altered lipid biosynthesis.

cell biology↗