Biology subjects

Merchant, S.

Publications and source records attributed to Merchant, S..

9 recordsLinked to original sources

bioRxiv2026

Glutathione Protects Hematopoietic Stem Cells from Proteotoxic Stress

Glutathione is a redox buffer, best known for preventing oxidative stress and ferroptosis, but rarely studied in stem cells. We studied glutathione function in hematopoietic stem cells (HSCs) by deleting the glutamate-cysteine ligase catalytic subunit (GCLC), which synthesizes glutathione, or glutathione peroxidase 4 (GPX4), which employs glutathione to prevent ferroptosis. GCLC deficiency eliminated glutathione from hematopoietic cells, severely impairing HSC function and hematopoiesis. In contrast, GPX4 deficiency did not affect HSC function and had little effect on hematopoiesis, even after vitamin E depletion, suggesting glutathione has ferroptosis-independent functions in HSCs. Consistent with this, GCLC deficiency increased reactive oxygen species levels in hematopoietic cells. But treatment with antioxidants or ferroptosis inhibitors gave little or no rescue of HSC function despite reducing ROS levels. Rather, GCLC-deficient, but not GPX4-deficient, HSCs exhibited unfolded protein accumulation, proteostatic stress, and cell death marked by elevated p53 levels and Annexin V staining. Deficiency for p53 prolonged the function of GCLC-deficient HSCs after transplantation. Glutathione is thus necessary in HSCs for proteostasis but may be dispensable for ferroptosis inhibition in vivo.

cell biology↗

bioRxiv2026

Estimation of chloroplast macromolecular complex copy numbers and subunit stoichiometries during the Chlamydomonas reinhardtii cell cycle

An unbiased, quantitative view of biomolecules in a living cell is a prerequisite for accurate modeling approaches and informs our understanding of cellular metabolism at scale. In this work, we used the total protein approach (TPA), in which the total protein mass of a given proteomics sample is used as a calibrator for absolute protein quantification, to determine protein abundances during the Chlamydomonas reinhardtii diurnal cycle. We use external, independently measured quantitative markers (metals, pigments) to assess the absolute protein abundances in unlabeled whole cell extracts. We calculate protein abundances in fg / cell of 7322 Chlamydomonas proteins, 2266 of which were captured in every time point, including the major proteins involved in the light reactions, photoprotection, proteostasis and fatty acid metabolism during a cell cycle. As expected, Rubisco large and small subunits are present in a 1:1 stoichiometry, with the large subunit being the most abundant protein in our data set, averaging 5.05 x 106 molecules per cell, reflecting 2.7% of the total protein mass. We noticed that PSII is the most abundant complex involved in the light reactions with 2.08 x 106 complexes per cell. PSI averages 1.75 x 106 complexes per cell and cytochrome b6f averages 0.77 x 106 complexes per cell. The TPA is a robust tool to study proteome dynamics quantitatively, while avoiding artefacts due to biochemical fractionation. Our proteome data set with an unprecedented temporal resolution is a valuable resource to assess protein abundances during the cell cycle in the reference alga Chlamydomonas.

systems biology↗

bioRxiv2026

Leptin Receptor+ cells create a perisinusoidal niche for thrombopoiesis in the bone marrow by synthesizing CXCL14

Leptin Receptor-expressing (LepR+) stromal cells in the bone marrow are a critical source of growth factors for the maintenance of hematopoietic stem cells (HSCs) and most restricted hematopoietic progenitors. An important unresolved question is whether they also regulate terminal differentiation in some hematopoietic cells. We found that LepR+ cells promote thrombopoiesis by synthesizing the chemokine CXCL14, which is expressed in the bone marrow by a subset of LepR+ cells. Cxcl14-expressing LepR+ cells extend fine processes that wrap around perisinusoidal megakaryocytes. Deletion of Cxcl14 from LepR+ cells did not significantly alter HSC function or most aspects of bone marrow hematopoiesis, including megakaryocyte generation; however, it significantly reduced the numbers of proplatelet-forming megakaryocytes in the bone marrow and platelets in the blood. CXCL14 promoted platelet formation by remodeling lipid metabolism in megakaryocytes, increasing fatty acid transporter expression and enabling megakaryocytes to take up more polyunsaturated fatty acids from circulation. A high fat diet rescued the formation of proplatelet-forming megakaryocytes and platelets in Lepr-cre; Cxcl14 fl/fl mice. CXCL14 protein was sufficient to promote platelet formation by megakaryocytes in vitro and in vivo. LepR+ cells thus create a perisinusoidal niche for thrombopoiesis by producing CXCL14, which regulates lipid metabolism and terminal differentiation in megakaryocytes. Key pointsO_LILeptin Receptor+ stromal cells regulate terminal differentiation in megakaryocytes in addition to maintaining stem and progenitor cells C_LIO_LICXCL14 from Leptin Receptor+ cells promotes the formation of platelets by remodeling lipid metabolism in megakaryocytes in the bone marrow C_LI

cell biology↗

bioRxiv2025

Investigating Overflow Metabolism in Heterotrophic Cultures of the Green Alga Chromochloris zofingiensis

Chromochloris zofingiensis is of interest for its ability to perform a reversible trophic switch in the presence of glucose that is characterized by a shutdown of photosynthesis and an accumulation of energy storage metabolites. Previous work has shown that this trophic switch is accompanied by overflow metabolism and the production of lactate in aerobic conditions. This trophic switch is not observed in nutrient replete media. We utilized isotopically assisted metabolic flux analysis to characterize intracellular flux distributions that are associated with different metabolic phenotypes observed in this organism in different media formulations in light and dark conditions. The results of this analysis showed differences in flux through carbon fixation reactions, the TCA cycle and through the reaction catalyzed by pyruvate kinase. This analysis was complemented with transcriptomics data collected for C. zofingiensis grown in iron limited conditions to provide further evidence towards the negative impact of iron limitation on both photosynthetic and respiratory activity. Overflow metabolism allows this alga to compensate for the lower energy production that results from iron limitation. This work highlights how nutrient availability can lead to drastic changes in the metabolism of C. zofingiensis.

systems biology↗

bioRxiv2024

Fe starvation induces a second LHCI tetramer to photosystem I in green algae

Iron (Fe) availability limits photosynthesis at a global scale where Fe-rich photosystem (PS) I abundance is drastically reduced in Fe-poor environments. We used single-particle cryo-electron microscopy to reveal a unique Fe starvation-dependent arrangement of light-harvesting chlorophyll (LHC) proteins where Fe starvation-induced TIDI1 is found in an additional tetramer of LHC proteins associated with PSI in Dunaliella tertiolecta and Dunaliella salina. These cosmopolitan green algae are resilient to poor Fe nutrition. TIDI1 is a distinct LHC protein that co- occurs in diverse algae with flavodoxin (an Fe-independent replacement for the Fe-containing ferredoxin). The antenna expansion in eukaryotic algae we describe here is reminiscent of the iron-starvation induced (isiA-encoding) antenna ring in cyanobacteria, which typically co-occurs with isiB, encoding flavodoxin. Our work showcases the convergent strategies that evolved after the Great Oxidation Event to maintain PSI capacity.

plant biology↗

bioRxiv2023

Iron rescues glucose-mediated photosynthesis repression during lipid accumulation in the green alga Chromochloris zofingiensis

Energy status and nutrients regulate photosynthetic protein expression. The unicellular green alga Chromochloris zofingiensis switches off photosynthesis in the presence of exogenous glucose (+Glc) in a process that depends on hexokinase (HXK1). Here, we show that this response requires that cells lack sufficient iron (-Fe). Cells grown in -Fe+Glc accumulate triacylglycerol (TAG) while losing photosynthesis and thylakoid membranes. However, cells with an iron supplement (+Fe+Glc) maintain photosynthesis and thylakoids while still accumulating TAG. Proteomic analysis shows that known photosynthetic proteins are most depleted in heterotrophy, alongside hundreds of uncharacterized, conserved proteins. Photosynthesis repression is associated with enzyme and transporter regulation that redirects iron resources to (a) respiratory instead of photosynthetic complexes and (b) a ferredoxin-dependent desaturase pathway supporting TAG accumulation rather than thylakoid lipid synthesis. Combining insights from diverse organisms from green algae to vascular plants, we show how iron and trophic constraints on metabolism aid gene discovery for photosynthesis and biofuel production.

plant biology↗

bioRxiv2020

Systematic characterization of gene function in a photosynthetic organism

Photosynthetic organisms are essential for human life, yet most of their genes remain functionally uncharacterized. Single-celled photosynthetic model systems have the potential to accelerate our ability to connect genes to functions. Here, using a barcoded mutant library of the model eukaryotic alga Chlamydomonas reinhardtii, we determined the phenotypes of more than 58,000 mutants under more than 121 different environmental growth conditions and chemical treatments. 78% of genes are represented by at least one mutant that showed a phenotype, providing clues to the functions of thousands of genes. Mutant phenotypic profiles allow us to place known and previously uncharacterized genes into functional pathways such as DNA repair, photosynthesis, the CO2-concentrating mechanism, and ciliogenesis. We illustrate the value of this resource by validating novel phenotypes and gene functions, including the discovery of three novel components of a defense pathway that counteracts actin cytoskeleton inhibitors released by other organisms. The data also inform phenotype discovery in land plants: mutants in Arabidopsis thaliana genes exhibit similar phenotypes to those we observed in their Chlamydomonas homologs. We anticipate that this resource will guide the functional characterization of genes across the tree of life.

systems biology↗

bioRxiv2020

Co-expressed subunits of dual genetic origin define a conserved supercomplex mediating essential protein import into chloroplasts

In photosynthetic eukaryotes, thousands of proteins are translated in the cytosol and imported into the chloroplast through the concerted action of two translocons — termed TOC and TIC — located in the outer and inner membranes of the chloroplast envelope, respectively. The degree to which the molecular composition of the TOC and TIC complexes is conserved over phylogenetic distances has remained controversial. Here, we combine transcriptomic, biochemical, and genetic tools in the green alga Chlamydomonas (Chlamydomonas reinhardtii) to demonstrate that, despite a lack of evident sequence conservation for some of its components, the algal TIC complex mirrors the molecular composition of a TIC complex from Arabidopsis thaliana. The Chlamydomonas TIC complex contains three nuclear-encoded subunits, Tic20, Tic56 and Tic100, and one chloroplast-encoded subunit, Tic214, and interacts with the TOC complex, as well as with several uncharacterized proteins to form a stable supercomplex (TicToc), indicating that protein import across both envelope membranes is mechanistically coupled. Expression of the nuclear and chloroplast genes encoding both known and the here newly identified TicToc components is highly coordinated, suggesting that a mechanism for regulating its biogenesis across compartmental boundaries must exist. Conditional repression of Tic214, the only chloroplast-encoded subunit in the TicToc complex, impairs the import of chloroplast proteins with essential roles in chloroplast ribosome biogenesis and protein folding and induces a pleiotropic stress response, including several proteins involved in the chloroplast unfolded protein response. These findings underscore the functional importance of the TicToc supercomplex in maintaining chloroplast proteostasis.Competing Interest StatementThe authors have declared no competing interest.View Full Text

plant biology↗

bioRxiv2020

Vanishing white matter disease expression of truncated EIF2B5 activates induced stress response

Vanishing White Matter disease (VWM) is a severe leukodystrophy of the central nervous system caused by mutations in subunits of the eukaryotic initiation factor 2B complex (eIF2B). Current models only partially recapitulate key disease features, and pathophysiology is poorly understood. Through development and validation of zebrafish (Danio rerio) models of VWM, we demonstrate that zebrafish eif2b mutants phenocopy VWM, including impaired somatic growth, early lethality, impaired myelination, loss of oligodendrocyte precursor cells, increased apoptosis in the CNS, and impaired motor swimming behavior. Expression of human EIF2B2 in the zebrafish eif2b2 mutant rescues lethality and CNS apoptosis, demonstrating conservation of function between zebrafish and human. In the mutants, intron 12 retention leads to expression of a truncated eif2b5 transcript. Expression of the truncated eif2b5 in wild-type larva impairs motor behavior and activates the ISR, suggesting that a feed-forward mechanism in VWM is a significant component of disease pathophysiology.

physiology↗