STAG2 Promotes Error Correction in Mitosis by Regulating Kinetochore–Microtubule Attachments

Mutations in the STAG2 gene are present in ∼20% of tumors from different tissues of origin. STAG2 encodes a subunit of the cohesin complex, and tumors with loss-of-function mutations are usually aneuploid and display elevated frequencies of lagging chromosomes during anaphase. Lagging chromosomes are a hallmark of chromosomal instability (CIN) arising from persistent errors in kinetochore-microtubule (kMT) attachment. To determine whether the loss of STAG2 increases the rate of formation of kMT attachment errors or decreases the rate of their correction, we examined mitosis in STAG2-deficient cells. STAG2 depletion does not impair bipolar spindle formation or delay mitotic progression. Instead, loss of STAG2 permits excessive centromere stretch along with hyperstabilization of kMT attachments. STAG2-deficient cells display mislocalization of Bub1 kinase, Bub3 and the chromosome passenger complex. Importantly, strategically destabilizing kMT attachments in tumor cells harboring STAG2 mutations by overexpression of the microtubule-destabilizing enzymes MCAK (also known as KIF2C) and Kif2B decreased the rate of lagging chromosomes and reduced the rate of chromosome missegregation. These data demonstrate that STAG2 promotes the correction of kMT attachment errors to ensure faithful chromosome segregation during mitosis

B Cells Promote Pancreatic Tumorigenesis

Summary: Three recent studies, approaching the question from different angles and using different and/or overlapping models, provide compelling evidence for the involvement of tumor-infiltrating B cells in the initiation and progression of pancreatic ductal adenocarcinoma. These studies highlight the need for a better understanding of pancreatic tumor–immune system interactions and the immunologic mechanisms that promote or inhibit tumorigenesis, paving the way for better treatment strategies.Bloodwise (UK) (Visiting Fellowship Grant (14043)

MFSD7C switches mitochondrial ATP synthesis to thermogenesis in response to heme

© 2020, The Author(s). ATP synthesis and thermogenesis are two critical outputs of mitochondrial respiration. How these outputs are regulated to balance the cellular requirement for energy and heat is largely unknown. Here we show that major facilitator superfamily domain containing 7C (MFSD7C) uncouples mitochondrial respiration to switch ATP synthesis to thermogenesis in response to heme. When heme levels are low, MSFD7C promotes ATP synthesis by interacting with components of the electron transport chain (ETC) complexes III, IV, and V, and destabilizing sarcoendoplasmic reticulum Ca2+-ATPase 2b (SERCA2b). Upon heme binding to the N-terminal domain, MFSD7C dissociates from ETC components and SERCA2b, resulting in SERCA2b stabilization and thermogenesis. The heme-regulated switch between ATP synthesis and thermogenesis enables cells to match outputs of mitochondrial respiration to their metabolic state and nutrient supply, and represents a cell intrinsic mechanism to regulate mitochondrial energy metabolism

Anticancer immunity induced by a synthetic tumor-targeted CD137 agonist

Background In contrast to immune checkpoint inhibitors, the use of antibodies as agonists of immune costimulatory receptors as cancer therapeutics has largely failed. We sought to address this problem using a new class of modular synthetic drugs, termed tumor-targeted immune cell agonists (TICAs), based on constrained bicyclic peptides (Bicycles).Methods Phage libraries displaying Bicycles were panned for binders against tumor necrosis factor (TNF) superfamily receptors CD137 and OX40, and tumor antigens EphA2, Nectin-4 and programmed death ligand 1. The CD137 and OX40 Bicycles were chemically conjugated to tumor antigen Bicycles with different linkers and stoichiometric ratios of binders to obtain a library of low molecular weight TICAs (MW <8 kDa). The TICAs were evaluated in a suite of in vitro and in vivo assays to characterize their pharmacology and mechanism of action.Results Linking Bicycles against costimulatory receptors (e.g., CD137) to Bicycles against tumor antigens (e.g., EphA2) created potent agonists that activated the receptors selectively in the presence of tumor cells expressing these antigens. An EphA2/CD137 TICA (BCY12491) efficiently costimulated human peripheral blood mononuclear cells in vitro in the presence of EphA2 expressing tumor cell lines as measured by the increased secretion of interferon γ and interleukin-2. Treatment of C57/Bl6 mice transgenic for the human CD137 extracellular domain (huCD137) bearing EphA2-expressing MC38 tumors with BCY12491 resulted in the infiltration of CD8+ T cells, elimination of tumors and generation of immunological memory. BCY12491 was cleared quickly from the circulation (plasma t1/2 in mice of 1–2 hr), yet intermittent dosing proved effective.Conclusion Tumor target-dependent CD137 agonism using a novel chemical approach (TICAs) afforded elimination of tumors with only intermittent dosing suggesting potential for a wide therapeutic index in humans. This work unlocks a new path to effective cancer immunotherapy via agonism of TNF superfamily receptors

Distinct Traits of Structural and Regulatory Evolutional Conservation of Human Genes with Specific Focus on Major Cancer Molecular Pathways

The evolution of protein-coding genes has both structural and regulatory components. The first can be assessed by measuring the ratio of non-synonymous to synonymous nucleotide substitutions. The second component can be measured as the normalized proportion of transposable elements that are used as regulatory elements. For the first time, we characterized in parallel the regulatory and structural evolutionary profiles for 10,890 human genes and 2972 molecular pathways. We observed a ~0.1 correlation between the structural and regulatory metrics at the gene level, which appeared much higher (~0.4) at the pathway level. We deposited the data in the publicly available database RetroSpect. We also analyzed the evolutionary dynamics of six cancer pathways of two major axes: Notch/WNT/Hedgehog and AKT/mTOR/EGFR. The Hedgehog pathway had both components slower, whereas the Akt pathway had clearly accelerated structural evolution. In particular, the major hub nodes Akt and beta-catenin showed both components strongly decreased, whereas two major regulators of Akt TCL1 and CTMP had outstandingly high evolutionary rates. We also noticed structural conservation of serine/threonine kinases and the genes related to guanosine metabolism in cancer signaling: GPCRs, G proteins, and small regulatory GTPases (Src, Rac, Ras); however, this was compensated by the accelerated regulatory evolution

Cleavage in the γ-subunit of the epithelial sodium channel (ENaC) plays an important role in the proteolytic activation of near-silent channels

The mechanisms by which proteases activate the epithelial sodium channel (ENaC) are not yet fully understood. We investigated the effect of extracellular proteases on rat ENaC heterologously expressed in Xenopus laevis oocytes. Application of trypsin increased ENaC whole-oocyte currents by about 8-fold without a concomitant increase in channel surface expression. The stimulatory effect of trypsin was preserved in oocytes expressing αγ-ENaC, but was abolished in oocytes expressing αβ-ENaC. Thus, the γ-subunit appears to be essential for channel activation by extracellular proteases. Site-directed mutagenesis of a putative prostasin cleavage site in the extracellular loop of the γ-subunit revealed that mutating the 181Lys residue to alanine (γK181A) increases ENaC baseline whole-oocyte currents, decreases channel surface expression, and largely reduces the stimulatory effect of extracellular proteases (trypsin, chymotrypsin and human neutrophil elastase). In single-channel recordings from outside-out patches we demonstrated that the γK181A mutation essentially abolishes the activation of near-silent channels by trypsin, while a stimulatory effect of trypsin on channel gating is preserved. This apparent dual effect of trypsin on channel gating and on the recruitment of near-silent channels was confirmed by experiments using the β518C mutant ENaC which can be converted to a channel with an open probability of nearly one by exposure to a sulfhydryl reagent. Interestingly, the γK181A mutation results in the spontaneous appearance of a 67 kDa fragment of the γ-subunit in the plasma membrane which can be prevented by a furin inhibitor and also occurs after channel activation by extracellular trypsin. This suggests that the mutation promotes channel cleavage and activation by endogenous proteases. This would lower the pool of near-silent channels and explain the constitutive activation and reduced responsiveness of the mutant channel to extracellular proteases. We conclude that the mutated site (K181A) affects a region in the γ-subunit of ENaC that is functionally important for the activation of near-silent channels by extracellular proteases