RNA sequestration in P-bodies sustains myeloid leukaemia.

Post-transcriptional mechanisms are fundamental safeguards of progenitor cell identity and are often dysregulated in cancer. Here, we identified regulators of P-bodies as crucial vulnerabilities in acute myeloid leukaemia (AML) through genome-wide CRISPR screens in normal and malignant haematopoietic progenitors. We found that leukaemia cells harbour aberrantly elevated numbers of P-bodies and show that P-body assembly is crucial for initiation and maintenance of AML. Notably, P-body loss had little effect upon homoeostatic haematopoiesis but impacted regenerative haematopoiesis. Molecular characterization of P-bodies purified from human AML cells unveiled their critical role in sequestering messenger RNAs encoding potent tumour suppressors from the translational machinery. P-body dissolution promoted translation of these mRNAs, which in turn rewired gene expression and chromatin architecture in leukaemia cells. Collectively, our findings highlight the contrasting and unique roles of RNA sequestration in P-bodies during tissue homoeostasis and oncogenesis. These insights open potential avenues for understanding myeloid leukaemia and future therapeutic interventions.

Lysosomal storage disease proteo/lipidomic profiling using nMOST links ferritinophagy with mitochondrial iron deficiencies in cells lacking NPC2.

Lysosomal storage diseases (LSDs) comprised ~50 monogenic diseases characterized by the accumulation of cellular material in lysosomes and associated defects in lysosomal function, but systematic molecular phenotyping is lacking. Here, we develop a nanoflow-based multi-omic single-shot technology (nMOST) workflow allowing simultaneously quantify HeLa cell proteomes and lipidomes from more than two dozen LSD mutants, revealing diverse molecular phenotypes. Defects in delivery of ferritin and its autophagic receptor NCOA4 to lysosomes (ferritinophagy) were pronounced in NPC2-/- cells, which correlated with increased lyso-phosphatidylcholine species and multi-lamellar membrane structures visualized by cryo-electron-tomography. Ferritinophagy defects correlated with loss of mitochondrial cristae, MICOS-complex components, and electron transport chain complexes rich in iron-sulfur cluster proteins. Strikingly, mitochondrial defects were alleviated when iron was provided through the transferrin system. This resource reveals how defects in lysosomal function can impact mitochondrial homeostasis in trans and highlights nMOST as a discovery tool for illuminating molecular phenotypes across LSDs.

PKA regulatory subunit Bcy1 couples growth, lipid metabolism, and fermentation during anaerobic xylose growth in Saccharomyces cerevisiae.

Organisms have evolved elaborate physiological pathways that regulate growth, proliferation, metabolism, and stress response. These pathways must be properly coordinated to elicit the appropriate response to an ever-changing environment. While individual pathways have been well studied in a variety of model systems, there remains much to uncover about how pathways are integrated to produce systemic changes in a cell, especially in dynamic conditions. We previously showed that deletion of Protein Kinase A (PKA) regulatory subunit BCY1 can decouple growth and metabolism in Saccharomyces cerevisiae engineered for anaerobic xylose fermentation, allowing for robust fermentation in the absence of division. This provides an opportunity to understand how PKA signaling normally coordinates these processes. Here, we integrated transcriptomic, lipidomic, and phospho-proteomic responses upon a glucose to xylose shift across a series of strains with different genetic mutations promoting either coupled or decoupled xylose-dependent growth and metabolism. Together, results suggested that defects in lipid homeostasis limit growth in the bcy1Δ strain despite robust metabolism. To further understand this mechanism, we performed adaptive laboratory evolutions to re-evolve coupled growth and metabolism in the bcy1Δ parental strain. The evolved strain harbored mutations in PKA subunit TPK1 and lipid regulator OPI1, among other genes, and evolved changes in lipid profiles and gene expression. Deletion of the evolved opi1 gene partially reverted the strain's phenotype to the bcy1Δ parent, with reduced growth and robust xylose fermentation. We suggest several models for how cells coordinate growth, metabolism, and other responses in budding yeast and how restructuring these processes enables anaerobic xylose utilization.

A role for cytoglobin in regulating intracellular hydrogen peroxide and redox signals in the vasculature.

The oxidant hydrogen peroxide serves as a signaling molecule that alters many aspects of cardiovascular functions. Recent studies suggest that cytoglobin - a hemoglobin expressed in the vasculature - may promote electron transfer reactions with proposed functions in hydrogen peroxide decomposition. Here, we determined the extent to which cytoglobin regulates intracellular hydrogen peroxide and established mechanisms. We found that cytoglobin decreased the hyperoxidation of peroxiredoxins and maintained the activity of peroxiredoxin 2 following challenge with exogenous hydrogen peroxide. Cytoglobin promoted a reduced intracellular environment and facilitated the reduction of the thiol-based hydrogen peroxide sensor Hyper7 after bolus addition of hydrogen peroxide. Cytoglobin also limited the inhibitory effect of hydrogen peroxide on glycolysis and reversed the oxidative inactivation of the glycolytic enzyme GAPDH. Our results indicate that cytoglobin in cells exists primarily as oxyferrous cytoglobin (CygbFe 2+ -O 2 ) with its cysteine residues in the reduced form. We found that the specific substitution of one of two cysteine residues on cytoglobin (C83A) inhibited the reductive activity of cytoglobin on Hyper7 and GAPDH. Carotid arteries from cytoglobin knockout mice were more sensitive to glycolytic inhibition by hydrogen peroxide than arteries from wildtype mice. Together, these results support a role for cytoglobin in regulating intracellular redox signals associated with hydrogen peroxide through oxidation of its cysteine residues, independent of hydrogen peroxide reaction at its heme center.