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Human C2orf69 is an evolutionarily conserved gene whose function is unknown. Here, we report eight unrelated families from which 20 children presented with a fatal syndrome consisting of severe autoinflammation and progredient leukoencephalopathy with recurrent seizures; 12 of these subjects, whose DNA was available, segregated homozygous loss-of-function C2orf69 variants. C2ORF69 bears homology to esterase enzymes, and orthologs can be found in most eukaryotic genomes, including that of unicellular phytoplankton. We found that endogenous C2ORF69 (1) is loosely bound to mitochondria, (2) affects mitochondrial membrane potential and oxidative respiration in cultured neurons, and (3) controls the levels of the glycogen branching enzyme 1 (GBE1) consistent with a glycogen-storage-associated mitochondriopathy. We show that CRISPR-Cas9-mediated inactivation of zebrafish C2orf69 results in lethality by 8 months of age due to spontaneous epileptic seizures, which is preceded by persistent brain inflammation. Collectively, our results delineate an autoinflammatory Mendelian disorder of C2orf69 deficiency that disrupts the development/homeostasis of the immune and central nervous systems.

Activation of coagulation factor (F) XI promotes multiorgan failure in rodent models of sepsis and in a baboon model of lethal systemic inflammation induced by infusion of heat-inactivated Staphylococcus aureus. Here we used the anticoagulant FXII-neutralizing antibody 5C12 to verify the mechanistic role of FXII in this baboon model. Compared with untreated control animals, repeated 5C12 administration before and at 8 and 24 hours after bacterial challenge prevented the dramatic increase in circulating complexes of contact system enzymes FXIIa, FXIa, and kallikrein with antithrombin or C1 inhibitor, and prevented cleavage and consumption of high-molecular-weight kininogen. Activation of several coagulation factors and fibrinolytic enzymes was also prevented. D-dimer levels exhibited a profound increase in the untreated animals but not in the treated animals. The antibody also blocked the increase in plasma biomarkers of inflammation and cell damage, including tumor necrosis factor, interleukin (IL)-1 beta, IL-6, IL-8, IL-10, granulocyte-macrophage colony-stimulating factor, nucleosomes, and myeloperoxidase. Based on clinical presentation and circulating biomarkers, inhibition of FXII prevented fever, terminal hypotension, respiratory distress, and multiorgan failure. All animals receiving 5C12 had milder and transient clinical symptoms and were asymptomatic at day 7, whereas untreated control animals suffered irreversible multiorgan failure and had to be euthanized within 2 days after the bacterial challenge. This study confirms and extends our previous finding that at least 2 enzymes of the contact activation complex, FXIa and FXIIa, play critical roles in the development of an acute and terminal inflammatory response in baboons challenged with heat-inactivated S aureus.

DOT1L, the only H3K79 methyltransferase in human cells and a homolog of the yeast Dot1, normally forms a complex with AF10, AF17, and ENL or AF9, is dysregulated in most cases of mixed-lineage leukemia (MLLr), and has been believed to regulate transcriptional elongation on the basis of its colocalization with RNA polymerase II (Pol II), the sharing of subunits (AF9 and ENL) between the DOT1L and super elongation complexes, and the distribution of H3K79 methylation on both promoters and transcribed regions of active genes. Here we show that DOT1L depletion in erythroleukemic cells reduces its global occupancy without affecting the traveling ratio or the elongation rate (assessed by 4sUDRB-seq) of Pol II, suggesting that DOT1L does not play a major role in elongation in these cells. In contrast, analyses of transcription initiation factor binding reveal that DOT1L and ENL depletions each result in reduced TATA binding protein (TBP) occupancies on thousands of genes. More importantly, DOT1L and ENL depletions concomitantly reduce TBP and Pol II occupancies on a significant fraction of direct (DOT1L-bound) target genes, indicating a role for the DOT1L complex in transcription initiation. Mechanistically, proteomic and biochemical studies suggest that the DOT1L complex may regulate transcriptional initiation by facilitating the recruitment or stabilization of transcription factor IID, likely in a monoubiquitinated H2B (H2Bub1)-enhanced manner. Additional studies show that DOT1L enhances H2Bub1 levels by limiting recruitment of the Spt-Ada-Gcn5-acetyltransferase (SAGA) complex. These results advance our understanding of roles of the DOT1L complex in transcriptional regulation and have important implications for MLLr leukemias.

BACKGROUND: Pregnant women and their neonates represent 2 vulnerable populations with an interdependent immune system that are highly susceptible to viral infections. The immune response of pregnant women to severe acute respiratory syndrome coronavirus 2 and the interplay of how the maternal immune response affects the neonatal passive immunity have not been studied systematically. OBJECTIVE: We characterized the serologic response in pregnant women and studied how this serologic response correlates with the maternal clinical presentation and with the rate and level of passive immunity that the neonate received from the mother. STUDY DESIGN: Women who gave birth and who tested positive for immunoglobulin M or immunoglobulin G against severe acute respiratory syndrome coronavirus 2 using semiquantitative detection in a New York City hospital between March 22, 2020, and May 31, 2020, were included in this study. A retrospective chart review of the cases that met the inclusion criteria was conducted to determine the presence of coronavirus disease 2019 symptoms and the use of oxygen support. Serology levels were compared between the symptomatic and asymptomatic patients using a Welch 2 sample t test. Further chart review of the same patient cohort was conducted to identify the dates of self-reported onset of coronavirus disease 2019 symptoms and the timing of the peak immunoglobulin M and immunoglobulin G antibody levels after symptom onset was visualized using local polynomial regression smoothing on log(2)-scaled serologic values. To study the neonatal serology response, umbilical cord blood samples of the neonates born to the subset of serology positive pregnant women were tested for serologic antibody responses. The maternal antibody levels of serology positive vs the maternal antibody levels of serology negative neonates were compared using the Welch 2 sample t test. The relationship between the quantitative maternal and quantitative neonatal serologic data was studied using a Pearson correlation and linear regression. A multiple linear regression analysis was conducted using maternal symptoms, maternal serology levels, and maternal use of oxygen support to determine the predictors of neonatal immunoglobulin G levels. RESULTS: A total of 88 serology positive pregnant women were included in this study. The antibody levels were higher in symptomatic pregnant women than in asymptomatic pregnant women. Serology studies in 34 women with symptom onset data revealed that the maternal immunoglobulin M and immunoglobulin G levels peak around 15 and 30 days after the onset of coronavirus disease 2019 symptoms, respectively. Furthermore, studies of 50 neonates born to this subset of serology positive women showed that passive immunity in the form of immunoglobulin G is conferred in 78% of all neonates. The presence of passive immunity is dependent on the maternal antibody levels, and the levels of neonatal immunoglobulin G correlate with maternal immunoglobulin G levels. The maternal immunoglobulin G levels and maternal use of oxygen support were predictive of the neonatal immunoglobulin G levels. CONCLUSION: We demonstrated that maternal serologies correlate with symptomatic maternal infection, and higher levels of maternal antibodies are associated with passive neonatal immunity. The maternal immunoglobulin G levels and maternal use of oxygen support, a marker of disease severity, predicted the neonatal immunoglobulin G levels. These data will further guide the screening for this uniquely linked population of mothers and their neonates and can aid in developing maternal vaccination strategies.

Measurements of fluorescence intensity of the hydrophobic pyridinium salt (DTPSH) remaining in the organic phase after partition experiments in the DCM/H2O system allowed an approximate method to be developed to estimate the mean number of molecules (N = 942) on the surface of 22.8 nm gold nanoparticles and the separation (1.89 nm) between these organic molecules. This protocol is based on the ability that the organic molecules possess to coat the surface of the nanoparticle, which can migrate from the organic to the aqueous phase as a result of the driving force of the strong binding of sulfur to gold. To validate our estimation, we used a projection of the results obtained by Wales and Ulker to solve the Thomson problem, a mathematicians' challenge, used as a model to calculate the mean distance (1.82 nm) separating particles on the surface, in excellent agreement with the results obtained by our method. The quality of results, the simplicity of calculations, the low fluorescence detection limit, and the inexpensive materials, recommend this procedure for rapid estimates of the mean number of molecules on the surface of nanoparticles.

The way in which interactions between mechanics and biochemistry lead to the emergence of complex cell and tissue organization is an old question that has recently attracted renewed interest from biologists, physicists, mathematicians and computer scientists. Rapid advances in optical physics, microscopy and computational image analysis have greatly enhanced our ability to observe and quantify spatiotemporal patterns of signalling, force generation, deformation, and flow in living cells and tissues. Powerful new tools for genetic, biophysical and optogenetic manipulation are allowing us to perturb the underlying machinery that generates these patterns in increasingly sophisticated ways. Rapid advances in theory and computing have made it possible to construct predictive models that describe how cell and tissue organization and dynamics emerge from the local coupling of biochemistry and mechanics. Together, these advances have opened up a wealth of new opportunities to explore how mechanochemical patterning shapes organismal development. In this roadmap, we present a series of forward-looking case studies on mechanochemical patterning in development, written by scientists working at the interface between the physical and biological sciences, and covering a wide range of spatial and temporal scales, organisms, and modes of development. Together, these contributions highlight the many ways in which the dynamic coupling of mechanics and biochemistry shapes biological dynamics: from mechanoenzymes that sense force to tune their activity and motor output, to collectives of cells in tissues that flow and redistribute biochemical signals during development.

The mu opioid receptor antagonist/kappa opioid receptor (KOR) partial agonist nalmefene (NMF), a close structural analog of naltrexone (NTX), has been shown to reduce cocaine reward in preclinical models. Given the greater KOR potency and improved bioavailability compared to NTX, NMF may be a promising pharmacotherapeutic for cocaine use disorder (CUD). Here we examine the effects of NMF pretreatment on chronic daily extended access (4h) cocaine intravenous self-administration (IVSA) in adult male C57Bl/6J mice. Methods: separate groups of mice had daily 4h cocaine IVSA sessions (0.25 or 0.5 mg/kg/inf, FR1) for 14 days. Starting on day 8, mice were pretreated with NMF (0, 1, or 10 mg/kg) 30m before each session. A separate group of mice acquired cocaine IVSA [seven days FR1 then four FR3 of 4h daily sessions (0.5 mg/kg/inf)] prior to a single progressive ratio 3 session to examine the effect of 1 mg/kg NMF on cocaine motivation. Results: No significant effect of NMF pretreatment on cocaine intake was observed. Acute pretreatment of 1 mg/kg NMF significantly potentiated cocaine motivation as measured by progressive ratio breakpoint. Conclusions: NMF did not significantly attenuate cocaine intake and increased motivation for cocaine suggesting that NMF may not be suitable for non-abstinent CUD patients. Further research is needed with KOR selective partial or full agonists to determine their effect on cocaine reinforcement.

Portal hypertension is a major contributor to decompensation and death from liver disease, a global health problem. Here, we demonstrate homozygous damaging mutations in GIMAP5, a small organellar GTPase, in four families with unexplained portal hypertension. We show that GIMAP5 is expressed in hepatic endothelial cells and that its loss in both humans and mice results in capillarization of liver sinusoidal endothelial cells (LSECs); this effect is also seen when GIMAP5 is selectively deleted in endothelial cells. Single-cell RNA-sequencing analysis in a GIMAP5-deficient mouse model reveals replacement of LSECs with capillarized endothelial cells, a reduction of macrovascular hepatic endothelial cells, and places GIMAP5 upstream of GATA4, a transcription factor required for LSEC specification. Thus, GIMAP5 is a critical regulator of liver endothelial cell homeostasis and, when absent, produces portal hypertension. These findings provide new insight into the pathogenesis of portal hypertension, a major contributor to morbidity and mortality from liver disease. Despite major advances in the diagnosis and treatment of viral causes of hepatitis (Vilarinho and Lifton, 2016), the incidence of chronic liver disease continues to rise worldwide, affecting up to 1.5 billion people globally (GBD 2017 Disease and Injury Incidence and Prevalence Collaborators, 2018) and leading to similar to 2 million deaths annually (Moon et al., 2020). Because the demand for liver transplantation far exceeds the supply of available donor organs, understanding the pathogenesis of advanced liver disease and its complications will be required to develop new therapies to reduce adverse disease outcomes. Portal hypertension-increased hepatic resistance to blood flow entering the liver-is a major contributor to the morbidity and mortality of liver disease owing to development of

The zinc-finger transcription factor IKAROS is essential for B cell development. Taniuchi, Morio and colleagues identify a human kindred presenting with B cell immunodeficiency that was caused by a heterozygous missense mutation in IKZF3 encoding the related AIOLOS protein. AIOLOS(G159R) is a mutant protein that interferes with both wild-type AIOLOS and IKAROS by forming heterodimers that bind to aberrant DNA-binding sites and prevent normal expression of IKAROS-dependent genes. In the present study, we report a human-inherited, impaired, adaptive immunity disorder, which predominantly manifested as a B cell differentiation defect, caused by a heterozygous IKZF3 missense variant, resulting in a glycine-to-arginine replacement within the DNA-binding domain of the encoded AIOLOS protein. Using mice that bear the corresponding variant and recapitulate the B and T cell phenotypes, we show that the mutant AIOLOS homodimers and AIOLOS-IKAROS heterodimers did not bind the canonical AIOLOS-IKAROS DNA sequence. In addition, homodimers and heterodimers containing one mutant AIOLOS bound to genomic regions lacking both canonical motifs. However, the removal of the dimerization capacity from mutant AIOLOS restored B cell development. Hence, the adaptive immunity defect is caused by the AIOLOS variant hijacking IKAROS function. Heterodimeric interference is a new mechanism of autosomal dominance that causes inborn errors of immunity by impairing protein function via the mutation of its heterodimeric partner.

Streptococcus suis, a ubiquitous bacterial colonizer in pigs, has recently extended host range to humans, leading to a global surge of deadly human infections and three large outbreaks since 1998. To better understand the mechanisms for the emergence of cross-species transmission and virulence in human, we have sequenced 366 S. suis human and pig isolates from 2005 to 2016 and performed a large-scale phylogenomic analysis on 1,634 isolates from 14 countries over 36 years. We show the formation of a novel human-associated clade (HAC) diversified from swine S. suis isolates. Phylogeographic analysis identified Europe as the origin of HAC, coinciding with the exportation of European swine breeds between 1960s and 1970s. HAC is composed of three sub-lineages and contains several healthy-pig isolates that display high virulence in experimental infections, suggesting healthy-pig carriers as a potential source for human infection. New HAC-specific genes are identified as promising markers for pathogen detection and surveillance. Our discovery of a human-associated S. suis clade provides insights into the evolution of this emerging human pathogen and extend our understanding of S. suis epidemics worldwide.