Recombinant ADAMTS13 for Immune Thrombotic Thrombocytopenic Purpura.

In patients with immune thrombotic thrombocytopenic purpura (iTTP), autoantibodies against the metalloprotease ADAMTS13 lead to catastrophic microvascular thrombosis. However, the potential benefits of recombinant human ADAMTS13 (rADAMTS13) in patients with iTTP remain unknown. Here, we report the clinical use of rADAMTS13, which resulted in the rapid suppression of disease activity and complete recovery in a critically ill patient whose condition had proved to be refractory to all available treatments.

Hypoxia extends lifespan and neurological function in a mouse model of aging.

There is widespread interest in identifying interventions that extend healthy lifespan. Chronic continuous hypoxia delays the onset of replicative senescence in cultured cells and extends lifespan in yeast, nematodes, and fruit flies. Here, we asked whether chronic continuous hypoxia is beneficial in mammalian aging.

Computer vision quantitation of erythrocyte shape abnormalities provides diagnostic, prognostic, and mechanistic insight.

Examination of red blood cell (RBC) morphology in peripheral blood smears can help diagnose hematologic diseases, even in resource-limited settings, but this analysis remains subjective and semiquantitative with low throughput. Prior attempts to develop automated tools have been hampered by their poor reproducibility and limited clinical validation. Here, we present a novel, open-source machine-learning approach (denoted as RBC-diff) to quantify abnormal RBCs in peripheral smear images and generate an RBC morphology differential.

Red blood cells contain enzymatically active GPx4 whose abundance anticorrelates with hemolysis during blood bank storage.

The antioxidant function of the phospholipid hydroperoxide glutathione peroxidase (GPx4) is vital for the homeostasis of many cell types, from neoplastic cells to normal erythroid precursors. However, some functional proteins in erythroid precursors are lost during the development of red blood cells (RBCs); whether GPx4 is maintained as an active enzyme in mature RBCs has remained unclear. Our meta-analyses of existing RBC proteomics and metabolomics studies revealed the abundance of GPx4 to be correlated with lipid-anchored proteins.

Marked factor V activity elevation in severe COVID-19 is associated with venous thromboembolism.

Coagulopathy causes morbidity and mortality in patients with coronavirus disease 2019 (COVID-19) due to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. Yet, the mechanisms are unclear and biomarkers are limited. Early in the pandemic, we observed markedly elevated factor V activity in a patient with COVID-19, which led us to measure factor V, VIII, and X activity in a cohort of 102 consecutive inpatients with COVID-19.

Retrospective cohort studies of repeat donors reveal donor-dependent variability in the recovery of transfused platelets.

Background: The in vivo recovery of transfused platelets is variable and often unpredictable. Although many recipient-dependent factors are well described, donor-dependent variables remain poorly understood.

Study Design And Methods: To explore donor-dependent variables we conducted 2 retrospective studies of platelet transfusion outcomes in repeat donors.

Chemotherapy-sparing treatment of haemophagocytic lymphohistiocytosis with intravenous immunoglobulins and corticosteroids.

Haemophagocytic lymphohistiocytosis (HLH) can be a rapidly fatal disease. Current treatment in adults is extrapolated from the HLH-2004 protocol that specifies a regimen of etoposide, dexamethasone and cyclosporine. However, HLH presents as a spectrum of disease severity.

Mass spectrometry proteomics reveals a function for mammalian CALCOCO1 in MTOR-regulated selective autophagy.

Macroautophagy/autophagy is suppressed by MTOR (mechanistic target of rapamycin kinase) and is an anticancer target under active investigation. Yet, MTOR-regulated autophagy remains incompletely mapped. We used proteomic profiling to identify proteins in the MTOR-autophagy axis.

A physician-scientist preceptorship in clinical and translational research enhances training and mentorship.

Background: Dual degree program MD/PhD candidates typically train extensively in basic science research and in clinical medicine, but often receive little formal experience or mentorship in clinical and translational research.

Methods: To address this educational and curricular gap, the University of Wisconsin Medical Scientist Training Program partnered with the University of Wisconsin Institute for Clinical and Translational Research to create a new physician-scientist preceptorship in clinical and translational research. This six-week apprentice-style learning experience-guided by a physician-scientist faculty mentor-integrates both clinical work and a translational research project, providing early exposure and hands-on experience with clinically oriented research and the integrated career of a physician-scientist.

Mitochondrial Complex I Activity Is Required for Maximal Autophagy.

Cells adapt to nutrient and energy deprivation by inducing autophagy, which is regulated by the mammalian target of rapamycin (mTOR) and AMP-activated protein kinases (AMPKs). We found that cell metabolism significantly influences the ability to induce autophagy, with mitochondrial complex I function being an important factor in the initiation, amplitude, and duration of the response. We show that phenformin or genetic defects in complex I suppressed autophagy induced by mTOR inhibitors, whereas autophagy was enhanced by strategies that increased mitochondrial metabolism.

Conserved Lipid and Small-Molecule Modulation of COQ8 Reveals Regulation of the Ancient Kinase-like UbiB Family.

Human COQ8A (ADCK3) and Saccharomyces cerevisiae Coq8p (collectively COQ8) are UbiB family proteins essential for mitochondrial coenzyme Q (CoQ) biosynthesis. However, the biochemical activity of COQ8 and its direct role in CoQ production remain unclear, in part due to lack of known endogenous regulators of COQ8 function and of effective small molecules for probing its activity in vivo. Here, we demonstrate that COQ8 possesses evolutionarily conserved ATPase activity that is activated by binding to membranes containing cardiolipin and by phenolic compounds that resemble CoQ pathway intermediates.

Biochemistry of Mitochondrial Coenzyme Q Biosynthesis.

Coenzyme Q (CoQ, ubiquinone) is a redox-active lipid produced across all domains of life that functions in electron transport and oxidative phosphorylation and whose deficiency causes human diseases. Yet, CoQ biosynthesis has not been fully defined in any organism. Several proteins with unclear molecular functions facilitate CoQ biosynthesis through unknown means, and multiple steps in the pathway are catalyzed by currently unidentified enzymes.

Mitochondrial protein functions elucidated by multi-omic mass spectrometry profiling.

Mitochondrial dysfunction is associated with many human diseases, including cancer and neurodegeneration, that are often linked to proteins and pathways that are not well-characterized. To begin defining the functions of such poorly characterized proteins, we used mass spectrometry to map the proteomes, lipidomes, and metabolomes of 174 yeast strains, each lacking a single gene related to mitochondrial biology. 144 of these genes have human homologs, 60 of which are associated with disease and 39 of which are uncharacterized.

Cerebellar Ataxia and Coenzyme Q Deficiency through Loss of Unorthodox Kinase Activity.

The UbiB protein kinase-like (PKL) family is widespread, comprising one-quarter of microbial PKLs and five human homologs, yet its biochemical activities remain obscure. COQ8A (ADCK3) is a mammalian UbiB protein associated with ubiquinone (CoQ) biosynthesis and an ataxia (ARCA2) through unclear means. We show that mice lacking COQ8A develop a slowly progressive cerebellar ataxia linked to Purkinje cell dysfunction and mild exercise intolerance, recapitulating ARCA2.

Mitochondrial ADCK3 employs an atypical protein kinase-like fold to enable coenzyme Q biosynthesis.

The ancient UbiB protein kinase-like family is involved in isoprenoid lipid biosynthesis and is implicated in human diseases, but demonstration of UbiB kinase activity has remained elusive for unknown reasons. Here, we quantitatively define UbiB-specific sequence motifs and reveal their positions within the crystal structure of a UbiB protein, ADCK3. We find that multiple UbiB-specific features are poised to inhibit protein kinase activity, including an N-terminal domain that occupies the typical substrate binding pocket and a unique A-rich loop that limits ATP binding by establishing an unusual selectivity for ADP.

A Gly-zipper motif mediates homodimerization of the transmembrane domain of the mitochondrial kinase ADCK3.

Interactions between α-helices within the hydrophobic environment of lipid bilayers are integral to the folding and function of transmembrane proteins; however, the major forces that mediate these interactions remain debated, and our ability to predict these interactions is still largely untested. We recently demonstrated that the frequent transmembrane association motif GASright, the GxxxG-containing fold of the glycophorin A dimer, is optimal for the formation of extended networks of Cα-H hydrogen bonds, supporting the hypothesis that these bonds are major contributors to association. We also found that optimization of Cα-H hydrogen bonding and interhelical packing is sufficient to computationally predict the structure of known GASright dimers at near atomic level.

Amine-reactive neutron-encoded labels for highly plexed proteomic quantitation.

We describe a novel amine-reactive chemical label that exploits differential neutron-binding energy between (13)C and (15)N isotopes. These neutron-encoded (NeuCode) chemical labels enable up to 12-plex MS1-based protein quantification. Each structurally identical, but isotopically unique, tag is encoded with a 12.