Biology subjects

Schafer, Z. T.

Publications and source records attributed to Schafer, Z. T..

9 recordsLinked to original sources

bioRxiv2025

Synthetic essentiality of TRAIL/TNFSF10 in VHL-deficient renal cell carcinoma

Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive subtype of kidney cancer. Loss of von Hippel-Lindau (VHL) and the consequent activation of hypoxia-inducible factor- (HIF, especially HIF2) plays an essential role in ccRCC initiation and progression. The approved HIF2 inhibitor belzutifan faces the challenge of resistance, presenting an opportunity of co-targeting HIF2 and another vulnerability. This study elucidates the synthetic essentiality of TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) in VHL-deficient ccRCC, uncovering a novel reciprocal regulation between HIF2 and TRAIL. TRAIL was identified as a direct transcriptional target of HIF2 and paradoxically found to be crucial for cell proliferation, primarily by activating the p38 MAPK pathway and facilitating G1/S phase transition. Depletion of endogenous TRAIL or inhibition of HIF2 with belzutifan sensitizes ccRCC cells to recombinant TRAIL, presenting a promising avenue for combination therapy to overcome both TRAIL resistance and belzutifan resistance in treating ccRCC. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/621197v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1d603b9org.highwire.dtl.DTLVardef@18e6dc7org.highwire.dtl.DTLVardef@1c953cdorg.highwire.dtl.DTLVardef@9cd144_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

Recharacterization of RSL3 reveals that the selenoproteome is a druggable target in colorectal cancer

Ferroptosis is a non-apoptotic form of cell death resulting from the iron-dependent accumulation of lipid peroxides. Colorectal cancer (CRC) cells accumulate high levels of intracellular iron and reactive oxygen species (ROS) and are thus particularly sensitive to ferroptosis. The compound (S)-RSL3 ([1S,3R]-RSL3) is a commonly used ferroptosis inducing compound that is currently characterized as a selective inhibitor of the selenocysteine containing enzyme (selenoprotein) Gluathione Peroxidase 4 (GPx4), an enzyme that utilizes glutathione to directly detoxify lipid peroxides. However, through chemical controls utilizing the (R) stereoisomer of RSL3 ([1R,3R]-RSL3) that does not bind GPx4, combined with inducible genetic knockdowns of GPx4 in CRC cell lines, we revealed that GPx4 dependency does not always align with (S)-RSL3 sensitivity, questioning the current characterization of GPx4 as the central regulator of ferroptosis. Utilizing affinity pull-down mass spectrometry with chemically modified (S)-RSL3 probes we discovered that the effects of (S)-RSL3 extend far beyond GPx4 inhibition, revealing that (S)-RSL3 is a broad and non-selective inhibitor of selenoproteins. To further investigate the therapeutic potential of broadly disrupting the selenoproteome as a therapeutic strategy in CRC, we employed additional chemical and genetic approaches. We found that the selenoprotein inhibitor auranofin, an FDA approved gold-salt, chemically induced oxidative cell death and ferroptosis in both in-vitro and in-vivo models of CRC. Consistent with these data, we found that AlkBH8, a tRNA-selenocysteine methyltransferase required for the translation of selenoproteins, is essential for the in-vitro growth and xenograft survival of CRC cell lines. In summary, these findings recharacterize the mechanism of action of the most commonly used ferroptosis inducing molecule, (S)-RSL3, and reveal that broad inhibition of selenoproteins is a promising novel therapeutic angle for the treatment of CRC.

cancer biology↗

bioRxiv2023

Acod1 Expression in Cancer Cells Promotes Immune Evasion through the Generation of Inhibitory Peptides

Targeting PD-1 is an important component of many immune checkpoint blockade (ICB) therapeutic approaches. However, ICB is not an efficacious strategy in a variety of cancer types, in part due to immunosuppressive metabolites in the tumor microenvironment (TME). Here, we find that PD-1-resistant cancer cells produce abundant itaconate (ITA) due to enhanced levels of aconitate decarboxylase (Acod1). Acod1 has an important role in the resistance to PD-1, as decreasing Acod1 levels in PD-1 resistant cancer cells can sensitize tumors to PD-1 therapy. Mechanistically, cancer cells with high Acod1 inhibit the proliferation of naive CD8+ T cells through the secretion of inhibitory factors. Surprisingly, inhibition of CD8+ T cell proliferation is not dependent on secretion of ITA, but is instead a consequence of the release of small inhibitory peptides. Our study suggests that strategies to counter the activity of Acod1 in cancer cells may sensitize tumors to ICB therapy.

cancer biology↗

bioRxiv2023

A Cell Cycle-Dependent Ferroptosis Sensitivity Switch Governed by EMP2

Ferroptosis is a non-apoptotic form of cell death characterized by iron-dependent lipid peroxidation. Ferroptosis can be induced by system xc- cystine/glutamate antiporter inhibition or by direct inhibition of the phospholipid hydroperoxidase glutathione peroxidase 4 (GPX4). The regulation of ferroptosis in response to system xc- inhibition versus direct GPX4 inhibition may be distinct. Here, we show that cell cycle arrest enhances sensitivity to ferroptosis triggered by GPX4 inhibition but not system xc- inhibition. Arrested cells have increased levels of oxidizable polyunsaturated fatty acid-containing phospholipids, which drives sensitivity to GPX4 inhibition. Epithelial membrane protein 2 (EMP2) expression is reduced upon cell cycle arrest and is sufficient to enhance ferroptosis in response to direct GPX4 inhibition. An orally bioavailable GPX4 inhibitor increased markers of ferroptotic lipid peroxidation in vivo in combination with a cell cycle arresting agent. Thus, responses to different ferroptosis-inducing stimuli can be regulated by cell cycle state.

cancer biology↗

bioRxiv2023

A paradox promoted by microglia cannibalism shortens the lifespan of developmental microglia

The overproduction of cells and subsequent production of debris is a universal principle of neurodevelopment. Here we show an additional feature of the developing nervous system that causes neural debris - promoted by the sacrificial nature of embryonic microglia that irreversibly become phagocytic after clearing other neural debris. Described as long-lived, microglia colonize the embryonic brain and persist into adulthood. Using transgenic zebrafish to investigate the microglia debris during brain construction, we identified that unlike other neural cell-types that die in developmental stages after they have expanded, necroptosis-dependent microglial debris is prevalent when microglia are expanding in the zebrafish brain. Time-lapse imaging of microglia demonstrates that this debris is cannibalized by other microglia. To investigate features that promote microglia death and cannibalism, we used time-lapse imaging and fate-mapping strategies to track the lifespan of individual developmental microglia. These approaches revealed that instead of embryonic microglia being long-lived cells that completely digest their phagocytic debris, once most developmental microglia in zebrafish become phagocytic they eventually die, including ones that are cannibalistic. These results establish a paradox -- which we tested by increasing neural debris and manipulating phagocytosis -- that once most microglia in the embryo become phagocytic, they die, create debris and then are cannibalized by other microglia, resulting in more phagocytic microglia that are destined to die.

developmental biology↗

bioRxiv2023

A role for fibroblast-derived SASP factors in the activation of pyroptotic cell death in mammary epithelial cells

In normal tissue homeostasis, bidirectional communication between different cell types can shape numerous biological outcomes. Many studies have documented instances of reciprocal communication between fibroblasts and cancer cells that functionally change cancer cell behavior. However, less is known about how these heterotypic interactions shape epithelial cell function in the absence of oncogenic transformation. Furthermore, fibroblasts are prone to undergo senescence, which is typified by an irreversible cell cycle arrest. Senescent fibroblasts are also known to secrete various cytokines into the extracellular space; a phenomenon that is termed the senescence-associated secretory phenotype (SASP). While the role of fibroblast derived SASP factors on cancer cells has been well studied, the impact of these factors on normal epithelial cells remains poorly understood. We discovered that treatment of normal mammary epithelial cells with conditioned media (CM) from senescent fibroblasts (SASP CM) results in a caspase-dependent cell death. This capacity of SASP CM to cause cell death is maintained across multiple senescence-inducing stimuli. However, the activation of oncogenic signaling in mammary epithelial cells mitigates the ability of SASP CM to induce cell death. Despite the reliance of this cell death on caspase activation, we discovered that SASP CM does not cause cell death by the extrinsic or intrinsic apoptotic pathway. Instead, these cells die by an NLRP3, caspase-1, and gasdermin D (GSDMD)-dependent induction of pyroptosis. Taken together, our findings reveal that senescent fibroblasts can cause pyroptosis in neighboring mammary epithelial cells, which has implications for therapeutic strategies that perturb the behavior of senescent cells.

cell biology↗

bioRxiv2022

Reprogramming of Iron Metabolism Confers Ferroptosis Resistance in ECM-Detached Cells

Cancer cells often acquire resistance to cell death programs induced by loss of integrin-mediated attachment to extracellular matrix (ECM). Given that adaptation to ECM-detached conditions can facilitate tumor progression and metastasis, there is significant interest in effective elimination of ECM-detached cancer cells. Here, we find that ECM-detached cells are remarkably resistant to the induction of ferroptosis. While alterations in membrane lipid content are observed during ECM-detachment, it is instead fundamental changes in iron metabolism that underlie resistance of ECM-detached cells to ferroptosis. More specifically, our data demonstrate that levels of free iron are low during ECM-detachment due to changes in both iron uptake and iron storage. In addition, we establish that lowering the levels of iron storage proteins sensitizes ECM-detached cells to death by ferroptosis. Taken together, our data suggest that therapeutics designed to kill cancer cells by ferroptosis may be hindered by lack of efficacy towards ECM-detached cells.

cell biology↗

bioRxiv2021

Oncogenic signaling inhibits c-FLIP expression and promotes cancer cell survival during ECM-detachment

Inhibition of programmed cell death pathways is frequently observed in cancer cells where it functions to facilitate tumor progression. However, some proteins involved in the regulation of cell death function dichotomously to both promote and inhibit cell death depending on the cellular context. As such, understanding how cell death proteins are regulated in a context-dependent fashion in cancer cells is of utmost importance. We have uncovered evidence that cellular FLICE-like Inhibitory Protein (c-FLIP), a well-known anti-apoptotic protein, is often downregulated in tumor tissue when compared to adjacent normal tissue. These data argue that c-FLIP may have activity distinct from its canonical role in antagonizing cell death. Interestingly, we have discovered that detachment from extracellular matrix (ECM) serves as a signal to elevate c-FLIP transcription and that oncogenic signaling blocks ECM-detachment-induced c-FLIP elevation. In addition, our data reveal that downregulation of c-FLIP promotes the survival of ECM-detached cells and that c-FLIP overexpression in cancer cells restricts the viability of cancer cells grown in anchorage-independent conditions. Taken together, our study reveals an unexpected role for c-FLIP in constraining the viability of cancer cells during ECM-detachment and raises the idea that c-FLIP may have context-dependent pro- and anti-cell death roles during tumorigenesis.

cell biology↗

bioRxiv2020

SGK1 Signaling Promotes Glucose Metabolism and Survival in Extracellular Matrix Detached Cells

Loss of integrin-mediated attachment to extracellular matrix (ECM) proteins can trigger a variety of cellular changes that impact cell viability. Foremost among these is the activation of anoikis, caspase-mediated cell death induced by ECM-detachment. In addition to anoikis, loss of ECM-attachment causes profound alterations in cellular metabolism that can lead to anoikis-independent cell death. Here, we describe a surprising role for serum and glucocorticoid kinase-1 (SGK1) in the promotion of energy production when cells are detached. Our data demonstrate that SGK1 activation is necessary and sufficient for ATP generation during ECM-detachment and anchorage-independent growth. More specifically, SGK1 promotes a substantial elevation in glucose uptake due to elevated GLUT1 transcription. In addition, carbon flux into the pentose phosphate pathway (PPP) is necessary to accommodate elevated glucose uptake and PPP-mediated glyceraldehyde-3-phosphate (G3P) is necessary for ATP production. Thus, our data unmask SGK1 as master regulator of glucose metabolism and cell survival during ECM-detached conditions.

cell biology↗