The Osiris family genes function as novel regulators of the tube maturation process in the Drosophila trachea.

Drosophila trachea is a premier model to study tube morphogenesis. After the formation of continuous tubes, tube maturation follows. Tracheal tube maturation starts with an apical secretion pulse that deposits extracellular matrix components to form a chitin-based apical luminal matrix (aECM). This aECM is then cleared and followed by the maturation of taenidial folds. Finally, air fills the tubes. Meanwhile, the cellular junctions are maintained to ensure tube integrity. Previous research has identified several key components (ER, Golgi, several endosomes) of protein trafficking pathways that regulate the secretion and clearance of aECM, and the maintenance of cellular junctions. The Osiris (Osi) gene family is located at the Triplo-lethal (Tpl) locus on chromosome 3R 83D4-E3 and exhibits dosage sensitivity. Here, we show that three Osi genes (Osi9, Osi15, Osi19), function redundantly to regulate adherens junction (AJ) maintenance, luminal clearance, taenidial fold formation, tube morphology, and air filling during tube maturation. The localization of Osi proteins in endosomes (Rab7-containing late endosomes, Rab11-containing recycling endosomes, Lamp-containing lysosomes) and the reduction of these endosomes in Osi mutants suggest the possible role of Osi genes in tube maturation through endosome-mediated trafficking. We analyzed tube maturation in zygotic rab11 and rab7 mutants, respectively, to determine whether endosome-mediated trafficking is required. Interestingly, similar tube maturation defects were observed in rab11 but not in rab7 mutants, suggesting the involvement of Rab11-mediated trafficking, but not Rab7-mediated trafficking, in this process. To investigate whether Osi genes regulate tube maturation primarily through the maintenance of Rab11-containing endosomes, we overexpressed rab11 in Osi mutant trachea. Surprisingly, no obvious rescue was observed. Thus, increasing endosome numbers is not sufficient to rescue tube maturation defects in Osi mutants. These results suggest that Osi genes regulate other aspects of endosome-mediated trafficking, or regulate an unknown mechanism that converges or acts in parallel with Rab11-mediated trafficking during tube maturation.

Long-Term Safety and Efficacy of Elexacaftor/Tezacaftor/Ivacaftor in Children Aged ⩾6 Years with Cystic Fibrosis and at Least One F508del Allele: A Phase 3, Open-Label Clinical Trial.

Rationale: A 24-week, phase 3, open-label study showed elexacaftor/tezacaftor/ivacaftor (ELX/TEZ/IVA) was safe and efficacious in children aged 6-11 years with cystic fibrosis (CF) and one or more F508del-CFTR alleles. Objectives: To assess long-term safety and efficacy of ELX/TEZ/IVA in children who completed the pivotal 24-week phase 3 trial. Methods: In this phase 3, two-part (part A and part B), open-label extension study, children aged ⩾6 years with CF heterozygous for F508del and a minimal function CFTR mutation (F/MF genotypes) or homozygous for F508del (F/F genotype) who completed the 24-week parent study received ELX/TEZ/IVA based on weight. Children weighing <30 kg received ELX 100 mg once daily/TEZ 50 mg once daily/IVA 75 mg every 12 hours, whereas children weighing ⩾30 kg received ELX 200 mg once daily/TEZ 100 mg once daily/IVA 150 mg every 12 hours (adult dose). The 96-week analysis of part A of this extension study is reported here. Measurements and Main Results: Sixty-four children (F/MF genotypes, n = 36; F/F genotype, n = 28) were enrolled and received one or more doses of ELX/TEZ/IVA. Mean (SD) period of exposure to ELX/TEZ/IVA was 93.9 (11.1) weeks. The primary endpoint was safety and tolerability. Adverse events and serious adverse events were consistent with common manifestations of CF disease. Overall, exposure-adjusted rates of adverse events and serious adverse events (407.74 and 4.72 events per 100 patient-years) were lower than in the parent study (987.04 and 8.68 events per 100 patient-years). One child (1.6%) had an adverse event of aggression that was moderate in severity and resolved after study drug discontinuation. From parent study baseline at Week 96 of this extension study, the mean percent predicted FEV1 increased (11.2 [95% confidence interval (CI), 8.3 to 14.2] percentage points), sweat chloride concentration decreased (-62.3 [95% CI, -65.9 to -58.8] mmol/L), Cystic Fibrosis Questionnaire-Revised respiratory domain score increased (13.3 [95% CI, 11.4 to 15.1] points), and lung clearance index 2.5 decreased (-2.00 [95% CI, -2.45 to -1.55] units). Increases in growth parameters were also observed. The estimated pulmonary exacerbation rate per 48 weeks was 0.04. The annualized rate of change in percent predicted FEV1 was 0.51 (95% CI, -0.73 to 1.75) percentage points per year. Conclusions: ELX/TEZ/IVA continued to be generally safe and well tolerated in children aged ⩾6 years through an additional 96 weeks of treatment. Improvements in lung function, respiratory symptoms, and CFTR function observed in the parent study were maintained. These results demonstrate the favorable long-term safety profile and durable clinical benefits of ELX/TEZ/IVA in this pediatric population. Clinical trial registered with www.clinicaltrials.gov (NCT04183790).

Selenite Inhibits Notch Signaling in Cells and Mice.

Selenium is an essential micronutrient with a wide range of biological effects in mammals. The inorganic form of selenium, selenite, is supplemented to relieve individuals with selenium deficiency and to alleviate associated symptoms. Additionally, physiological and supranutritional selenite have shown selectively higher affinity and toxicity towards cancer cells, highlighting their potential to serve as chemotherapeutic agents or adjuvants. At varying doses, selenite extensively regulates cellular signaling and modulates many cellular processes. In this study, we report the identification of Delta-Notch signaling as a previously uncharacterized selenite inhibited target. Our transcriptomic results in selenite treated primary mouse hepatocytes revealed that the transcription of Notch1, Notch2, Hes1, Maml1, Furin and c-Myc were all decreased following selenite treatment. We further showed that selenite can inhibit Notch1 expression in cultured MCF7 breast adenocarcinoma cells and HEPG2 liver carcinoma cells. In mice acutely treated with 2.5 mg/kg selenite via intraperitoneal injection, we found that Notch1 expression was drastically lowered in liver and kidney tissues by 90% and 70%, respectively. Combined, these results support selenite as a novel inhibitor of Notch signaling, and a plausible mechanism of inhibition has been proposed. This discovery highlights the potential value of selenite applied in a pathological context where Notch is a key drug target in diseases such as cancer, fibrosis, and neurodegenerative disorders.

Two contemporaneous mitogenomes from terminal Pleistocene burials in eastern Beringia.

Pleistocene residential sites with multiple contemporaneous human burials are extremely rare in the Americas. We report mitochondrial genomic variation in the first multiple mitochondrial genomes from a single prehistoric population: two infant burials (USR1 and USR2) from a common interment at the Upward Sun River Site in central Alaska dating to ∼11,500 cal B.P. Using a targeted capture method and next-generation sequencing, we determined that the USR1 infant possessed variants that define mitochondrial lineage C1b, whereas the USR2 genome falls at the root of lineage B2, allowing us to refine younger coalescence age estimates for these two clades. C1b and B2 are rare to absent in modern populations of northern North America. Documentation of these lineages at this location in the Late Pleistocene provides evidence for the extent of mitochondrial diversity in early Beringian populations, which supports the expectations of the Beringian Standstill Model.

Computational LADAR imaging.

Most LADAR (laser radar, LIDAR) imaging systems use pixel-basis sampling, where each azimuth and elevation resolution element is uniquely sampled and recorded. We demonstrate and examine alternative sampling and post-detection processing schemes where recorded measurements are made in alternative bases that are intended to reduce system power consumption and laser emissions. A prototype of such a sensor having the capability to generate arbitrary illumination beam patterns rather than spot, line scanning, or flash techniques is described along with computational imaging algorithms to reduce speckle and identify scene objects in a low-dimensional compressed basis rather than in the pixel basis. Such techniques yield considerable energy savings and prove valuable when used on platforms with severe limitations on sensor size, weight, and power, and in particular as part of autonomous systems where image output for human interpretation is unnecessary.

The Skin Cancer Objective Structured Clinical Examination (SCOSCE): A multi-institutional collaboration to develop and validate a clinical skills assessment for melanoma.

BACKGROUND: Assessing medical students on core skills related to melanoma detection is challenging in the absence of a well-developed instrument. OBJECTIVE: We sought to develop an objective structured clinical examination for the detection and evaluation of melanoma among medical students. METHODS: This was a prospective cohort analysis of student and objective rater agreement on performance of clinical skills and assessment of differences in performance across 3 schools. RESULTS: Kappa coefficients indicated excellent agreement for 3 of 5 core skills including commenting on the presence of the moulage (k = 0.87, 95% confidence interval 0.77-0.96), obtaining a history for the moulage (k = 0.84, 95% confidence interval 0.74-0.94), and making a clinical impression (k = 0.80, 95% confidence interval 0.68-0.92). There were no differences in performance across schools with respect to 3 of 5 core skills: commenting on the presence of the moulage (P = .15), initiating a history (P = .53), and managing the suspicious lesion (P value range .07-.17). Overall, 54.2% and 44.7% of students commented on the presence of the moulage and achieved maximum performance of core skills, respectively, with no difference in performance across schools. LIMITATIONS: Limitations include overall sample size of students and schools. CONCLUSION: The Skin Cancer Objective Structured Clinical Examination represents a potentially important instrument to measure students' performance on the optimal step-by-step evaluation of a melanoma.

The Integrated Skin Exam film: an educational intervention to promote early detection of melanoma by medical students.

BACKGROUND: Knowledge of the skin cancer examination (SCE) and its practice remain relevant competency gaps among medical students. OBJECTIVE: We elaborate on a method of SCE known as the Integrated Skin Exam and discuss the development of an instructional film that illustrates its principles. We assess the tool's effect on knowledge, attitudes, and perceptions related to the SCE. METHODS: Second-year students among 8 randomized schools viewed the film and completed pre-post questionnaires. RESULTS: After viewing The Integrated Skin Exam film, students demonstrated improved melanoma knowledge, including identification of high-risk demographic groups (61% vs 42.9%, P < .001), high-risk anatomic sites in women (88.6% vs 46.5%, P < .001) and men (92.1% vs 34.8%, P < .001), and the ABCDEs of melanoma (98.4% vs 91.2%, P < .001). Students demonstrated increased confidence in the SCE (66.93% vs 16.40%, P < .001) and augmented intentions to practice it (99.05% vs 13.9%, P < .001). A greater proportion (70.4% vs 41.9%, P < .001) of students thought less than 3 minutes were required to integrate SCE into the routine examination. LIMITATIONS: Longitudinal impact of the film was not assessed. CONCLUSION: The Integrated Skin Exam film introduces an integrated approach to the SCE that addresses knowledge gaps, mitigates perceived barriers, and augments intention related to practice of the SCE.

N-naphthoyl-beta-naltrexamine (NNTA), a highly selective and potent activator of µ/kappa-opioid heteromers.

Numerous G protein-coupled receptors (GPCRs) have been shown to form heteromeric receptors in cell-based assays. Among the many heteromers reported in the opioid receptor family are µ/κ, κ/δ, and µ/δ. However, the in vivo physiological and behavioral relevance for the proposed heteromers have not yet been established. Here we report a unique example of a ligand, N-naphthoyl-ß-naltrexamine (NNTA) that selectively activates heteromeric µ/κ-opioid receptors in HEK-293 cells and induces potent antinociception in mice. NNTA was an exceptionally potent agonist in cells expressing µ/κ-opioid receptors. Intriguingly, it was found to be a potent antagonist in cells expressing only µ-receptors. In the mouse tail-flick assay, intrathecal (i.t.) NNTA produced antinociception that was ~100-fold greater than by intracerebroventricular (i.c.v.) administration. The κ-antagonist, norBNI, decreased the i.t. potency, and the activity was virtually abolished in µ-opioid receptor knockout mice. No tolerance was induced i.t., but marginal tolerance (3-fold) was observed via the i.c.v. route. Moreover, NNTA produced neither significant physical dependence nor place preference in the ED50 dose range. Taken together, this work provides an important pharmacologic tool for investigating the in vivo functional relevance of heteromeric µ/κ-opioid receptors and suggests an approach to potent analgesics with fewer deleterious side effects.

ADAM10 mediates vascular injury induced by Staphylococcus aureus α-hemolysin.

Staphylococcus aureus is a leading cause of bacteremia and sepsis. The interaction of S. aureus with the endothelium is central to bloodstream infection pathophysiology yet remains ill-understood. We show herein that staphylococcal α-hemolysin, a pore-forming cytotoxin, is required for full virulence in a murine sepsis model. The α-hemolysin binding to its receptor A-disintegrin and metalloprotease 10 (ADAM10) upregulates the receptor's metalloprotease activity on endothelial cells, causing vascular endothelial-cadherin cleavage and concomitant loss of endothelial barrier function. These cellular injuries and sepsis severity can be mitigated by ADAM10 inhibition. This study therefore provides mechanistic insight into toxin-mediated endothelial injury and suggests new therapeutic approaches for staphylococcal sepsis.

Hypometabolism, Alzheimer's Disease, and Possible Therapeutic Targets: An Overview.

The brain is a highly dynamic organ that requires a constant energy source to function normally. This energy is mostly supplied by glucose, a simple sugar that serves as the brain's principal fuel source. Glucose transport across the blood-brain barrier (BBB) is primarily controlled via sodium-independent facilitated glucose transport, such as by glucose transporter 1 (GLUT1) and 3 (GLUT3). However, other glucose transporters, including GLUT4 and the sodium-dependent transporters SGLT1 and SGLT6, have been reported in vitro and in vivo. When the BBB endothelial layer is crossed, neurons and astrocytes can absorb the glucose using their GLUT1 and GLUT3 transporters. Glucose then enters the glycolytic pathway and is metabolized into adenosine triphosphate (ATP), which supplies the energy to support cellular functions. The transport and metabolism of glucose in the brain are impacted by several medical conditions, which can cause neurological and neuropsychiatric symptoms. Alzheimer's disease (AD), Parkinson's disease (PD), epilepsy, traumatic brain injury (TBI), schizophrenia, etc., are a few of the most prevalent disorders, characterized by a decline in brain metabolism or hypometabolism early in the course of the disease. Indeed, AD is considered a metabolic disorder related to decreased brain glucose metabolism, involving brain insulin resistance and age-dependent mitochondrial dysfunction. Although the conventional view is that reduced cerebral metabolism is an effect of neuronal loss and consequent brain atrophy, a growing body of evidence points to the opposite, where hypometabolism is prodromal or at least precedes the onset of brain atrophy and the manifestation of clinical symptoms. The underlying processes responsible for these glucose transport and metabolic abnormalities are complicated and remain poorly understood. This review article provides a comprehensive overview of the current understanding of hypometabolism in AD and potential therapeutic targets.

Loss of hepatic manganese transporter ZIP8 disrupts serum transferrin glycosylation and the glutamate-glutamine cycle.

BACKGROUND: ZIP8, encoded by SLC39A8, is a membrane transporter that facilitates the cellular uptake of divalent biometals including zinc (Zn), manganese (Mn), and iron (Fe). The hepatic system has long been accepted as the central modulator for whole-body biometal distribution. Earlier investigations suggest the propensity of ZIP8 to prioritize Mn influx, as opposed to Fe or Zn, in hepatocytes. Hepatic ZIP8 Mn transport is crucial for maintaining homeostasis of various Mn-dependent metalloenzymes and their associated pathways. Herein, we hypothesize that a drastic decrease in systemic Mn, via the loss of hepatic ZIP8, disrupts two unique cellular pathways, post-translational glycosylation and the glutamate-glutamine cycle. METHODS: ZIP8 liver-specific knockout (LSKO) mice were chosen in an attempt to substantially decrease whole-body Mn levels. To further elucidate the role of Mn in serum glycosylation, a Mn-deficient diet was adopted in conjunction with the LSKO mice to model a near-complete loss of systemic Mn. After the treatment course, transferrin sialylation profiles were determined using imaged capillary isoelectric focusing (icIEF). We also investigated the role of Mn in the glutamate-glutamine cycle; the conversion of glutamate to glutamine in F/F and LSKO mice was assessed by the glutamine/glutamate ratio in cerebrospinal fluid (CSF) via HPLC-MS. An open-field study was ultimately conducted to check if these mice displayed atypical behavior. RESULTS: Two major biological pathways were found to be significantly altered due to the loss of hepatic ZIP8. We identified a disparity between F/F and LSKO transferrin sialylation profiles that were exacerbated under a Mn-deficient diet. Additionally, we discovered a neurotransmitter imbalance between the levels of glutamine and glutamate, exclusive to LSKO mice. This was characterized by the decreased glutamine/glutamate ratio in CSF. Secondary to the neurotransmitter alteration, LSKO mice exhibited an increase in locomotor activity in an open-field. CONCLUSION: Our model successfully established a connection between the loss of hepatic ZIP8 and two Mn-dependent cellular pathways, namely, protein glycosylation and the glutamate-glutamine cycle.

Establishing a Cystic Fibrosis Learning Network: Interventions to promote collaboration and data-driven improvement at scale.

Introduction: A learning health network is a type of learning health system in which stakeholders use network organization to improve health and health care. Building on existing resources in the cystic fibrosis (CF) community, the Cystic Fibrosis Learning Network (CFLN) was designed to improve medical outcomes and quality of life through an intentional focus on achieving reliable evidence-based chronic care delivery and creating a system for data-driven collaborative learning. Methods: We describe the development and growth of the CFLN considering six domains of a Network Maturity Grid: system leadership; governance and policy management; quality improvement (QI); engagement and community building; data and analytics; and research. We illustrate the impact of the CFLN experience on chronic care processes and indicators of collaborative infrastructure. Results: The CFLN represents 36 accredited care centers in the CF Foundation Care Center Network caring for over 6300 patients. Of 6779 patient clinical care visits/quarter, 77% are entered into the CF Foundation Patient Registry within 30 days, providing timely means to track outcomes. Collaborative visit planning is occurring in 93% of clinical care visits to share agenda setting with patients and families. Almost all CFLN teams (94%, n = 34) have a patient/family partner (PFP), and 74% of PFPs indicate they are actively participating, taking ownership of, or leading QI initiatives with the interdisciplinary care team. In 2022, 97% of centers reported completing 1-13 improvement cycles per month, and 82% contributed to monthly QI progress reports to share learning. Conclusion: The CFLN is a maturing, collaborative infrastructure. CFLN centers practice at an advanced level of coproduction. The CFLN fosters interdisciplinary and PFP leadership and the performance of consistent data-driven improvement cycles. CFLN centers are positioned to respond to rapid changes in evidence-based care and advance the practice of QI and implementation science on a broader scale.

The US national registry for childhood interstitial and diffuse lung disease: Report of study design and initial enrollment cohort.

INTRODUCTION: Childhood interstitial and diffuse lung disease (chILD) encompasses a broad spectrum of rare disorders. The Children's Interstitial and Diffuse Lung Disease Research Network (chILDRN) established a prospective registry to advance knowledge regarding etiology, phenotype, natural history, and management of these disorders. METHODS: This longitudinal, observational, multicenter registry utilizes single-IRB reliance agreements, with participation from 25 chILDRN centers across the U.S. Clinical data are collected and managed using the Research Electronic Data Capture (REDCap) electronic data platform. RESULTS: We report the study design and selected elements of the initial Registry enrollment cohort, which includes 683 subjects with a broad range of chILD diagnoses. The most common diagnosis reported was neuroendocrine cell hyperplasia of infancy, with 155 (23%) subjects. Components of underlying disease biology were identified by enrolling sites, with cohorts of interstitial fibrosis, immune dysregulation, and airway disease being most commonly reported. Prominent morbidities affecting enrolled children included home supplemental oxygen use (63%) and failure to thrive (46%). CONCLUSION: This Registry is the largest longitudinal chILD cohort in the United States to date, providing a powerful framework for collaborating centers committed to improving the understanding and treatment of these rare disorders.

Deletion of the monkeypox virus inhibitor of complement enzymes locus impacts the adaptive immune response to monkeypox virus in a nonhuman primate model of infection.

Monkeypox virus (MPXV) is an orthopoxvirus closely related to variola virus, the causative agent of smallpox. Human MPXV infection results in a disease that is similar to smallpox and can also be fatal. Two clades of MPXV have been identified, with viruses of the central African clade displaying more pathogenic properties than those within the west African clade. The monkeypox inhibitor of complement enzymes (MOPICE), which is not expressed by viruses of the west African clade, has been hypothesized to be a main virulence factor responsible for increased pathogenic properties of central African strains of MPXV. To gain a better understanding of the role of MOPICE during MPXV-mediated disease, we compared the host adaptive immune response and disease severity following intrabronchial infection with MPXV-Zaire (n = 4), or a recombinant MPXV-Zaire (n = 4) lacking expression of MOPICE in rhesus macaques (RM). Data presented here demonstrate that infection of RM with MPXV leads to significant viral replication in the peripheral blood and lungs and results in the induction of a robust and sustained adaptive immune response against the virus. More importantly, we show that the loss of MOPICE expression results in enhanced viral replication in vivo, as well as a dampened adaptive immune response against MPXV. Taken together, these findings suggest that MOPICE modulates the anti-MPXV immune response and that this protein is not the sole virulence factor of the central African clade of MPXV.

Spectral LADAR: active range-resolved three-dimensional imaging spectroscopy.

We present the concept and experimental results for Spectral LADAR, an augmented LADAR imager combining three-dimensional (3D) time-of-flight ranging with active multispectral sensing in the shortwave infrared (1080-1620 nm). The demonstrated technique is based on a nanosecond regime pulsed supercontinuum transmitter and spectrally multiplexed receiver that computes a high-resolution range value for each of 25 spectral bands. A low frame-rate prototype unit is described. Results demonstrating 3D imaging and material type classification of objects, especially those obscured by camouflage, are shown at effective stand-off ranges exceeding 40 m. These capabilities and the highly eye safe wavelengths at which the system operates make it suitable for applications in military imaging and robotic perception.

Risk finance for catastrophe losses with Pareto-calibrated Lévy-stable severities.

For catastrophe losses, the conventional risk finance paradigm of enterprise risk management identifies transfer, as opposed to pooling or avoidance, as the preferred solution. However, this analysis does not necessarily account for differences between light- and heavy-tailed characteristics of loss portfolios. Of particular concern are the decreasing benefits of diversification (through pooling) as the tails of severity distributions become heavier. In the present article, we study a loss portfolio characterized by nonstochastic frequency and a class of Lévy-stable severity distributions calibrated to match the parameters of the Pareto II distribution. We then propose a conservative risk finance paradigm that can be used to prepare the firm for worst-case scenarios with regard to both (1) the firm's intrinsic sensitivity to risk and (2) the heaviness of the severity's tail.

Rotator Cuff Repair Using Coracoacromial Ligament Autograft for Supraspinatus Footprint Augmentation.

Failure of rotator cuff repair can be a disastrous clinical outcome. Although failure is a multifactorial issue, recent interest has piqued in understanding the biology of the insertional components of the supraspinatus and infraspinatus at the footprint. When the torn tendon is of poor quality, especially if it is diminutive or thin, rotator cuff repair augmentation should be considered to aid in long-term healing. Various allograft options have been described in the past, and more recently, xenografts and synthetics have become more commonly used. The use of autografts in the treatment of insertional footprint deficiency has great potential; however, few grafts have been described. This study describes the surgical technique for footprint augmentation in arthroscopic supraspinatus repair using harvested autologous coracoacromial ligament tissue.

Arsenic Metabolism, Toxicity and Accumulation in the White Button Mushroom Agaricus bisporus.

Mushrooms have unique properties in arsenic metabolism. In many commercial and wild-grown mushrooms, arsenobetaine (AsB), a non-toxic arsenical, was found as the dominant arsenic species. The AsB biosynthesis remains unknown, so we designed experiments to study conditions for AsB formation in the white button mushroom, Agaricus bisporus. The mushrooms were treated with various arsenic species including arsenite (As(III)), arsenate (As(V)), methylarsenate (MAs(V)), dimethylarsenate (DMAs(V)) and trimethylarsine oxide (TMAsO), and their accumulation and metabolism were determined using inductively coupled mass spectrometer (ICP-MS) and high-pressure liquid chromatography coupled with ICP-MS (HPLC-ICP-MS), respectively. Our results showed that mycelia have a higher accumulation for inorganic arsenicals while fruiting bodies showed higher accumulation for methylated arsenic species. Two major arsenic metabolites were produced in fruiting bodies: DMAs(V) and AsB. Among tested arsenicals, only MAs(V) was methylated to DMAs(V). Surprisingly, AsB was only detected as the major arsenic product when TMAsO was supplied. Additionally, AsB was only detected in the fruiting body, but not mycelium, suggesting that methylated products were transported to the fruiting body for arsenobetaine formation. Overall, our results support that methylation and AsB formation are two connected pathways where trimethylated arsenic is the optimal precursor for AsB formation.

Type I signal peptidase and protein secretion in Staphylococcus epidermidis.

Bacterial protein secretion is a highly orchestrated process that is essential for infection and virulence. Despite extensive efforts to predict or experimentally detect proteins that are secreted, the characterization of the bacterial secretome has remained challenging. A central event in protein secretion is the type I signal peptidase (SPase)-mediated cleavage of the N-terminal signal peptide that targets a protein for secretion via the general secretory pathway, and the arylomycins are a class of natural products that inhibit SPase, suggesting that they may be useful chemical biology tools for characterizing the secretome. Here, using an arylomycin derivative, along with two-dimensional gel electrophoresis and liquid chromatography-tandem mass spectrometry (LC-MS/MS), we identify 11 proteins whose secretion from stationary-phase Staphylococcus epidermidis is dependent on SPase activity, 9 of which are predicted to be translated with canonical N-terminal signal peptides. In addition, we find that the presence of extracellular domains of lipoteichoic acid synthase (LtaS) and the ß-lactam response sensor BlaR1 in the medium is dependent on SPase activity, suggesting that they are cleaved at noncanonical sites within the protein. In all, the data define the proteins whose stationary-phase secretion depends on SPase and also suggest that the arylomycins should be valuable chemical biology tools for the study of protein secretion in a wide variety of different bacteria.