Variation in the spectrum of new mutations among inbred strains of mice.

The mouse serves as a mammalian model for understanding the nature of variation from new mutations, a question that has both evolutionary and medical significance. Previous studies suggest that the rate of single nucleotide mutations (SNMs) in mice is approximately 50% of that in humans. However, information largely comes from studies involving the C57BL/6 strain, and there is little information from other mouse strains. Here, we study the mutations that accumulated in 59 mouse lines derived from four inbred strains that are commonly used in genetics and clinical research (BALB/cAnNRj, C57BL/6JRj, C3H/HeNRj, and FVB/NRj), maintained for 8-9 generations by brother-sister mating. By analysing Illumina whole-genome sequencing data, we estimate that the average rate of new SNMs in mice is approximately µ = 6.7 × 10-9. However, there is substantial variation in the spectrum of SNMs among strains, so the burden from new mutations also varies among strains. For example, the FVB strain has a spectrum that is markedly skewed towards C→A transversions, and is likely to experience a higher deleterious load than other strains, due to an increased frequency of nonsense mutations in glutamic acid codons. Finally, we observe substantial variation in the rate of new SNMs among DNA sequence contexts, CpG sites and their adjacent nucleotides playing an important role.

Investigating the Bone Bruise Patterns in Pediatric Patients With Contact and Noncontact Acute Anterior Cruciate Ligament Tears: A Multicenter Study.

BACKGROUND: In adults with anterior cruciate ligament (ACL) tears, bone bruises on magnetic resonance imaging (MRI) scans provide insight into the underlying mechanism of injury. There is a paucity of literature that has investigated these relationships in children with ACL tears. PURPOSE: To examine and compare the number and location of bone bruises between contact and noncontact ACL tears in pediatric patients. STUDY DESIGN: Cohort study; Level of evidence, 3. METHODS: Boys ≤14 years and girls ≤12 years of age who underwent primary ACL reconstruction surgery between 2018 and 2022 were identified at 3 separate institutions. Eligibility criteria required detailed documentation of the mechanism of injury and MRI performed within 30 days of the initial ACL tear. Patients with congenital lower extremity abnormalities, concomitant fractures, injuries to the posterolateral corner and/or posterior cruciate ligament, previous ipsilateral knee injuries or surgeries, or closed physes evident on MRI scans were excluded. Patients were stratified into 2 groups based on a contact or noncontact mechanism of injury. Preoperative MRI scans were retrospectively reviewed for the presence of bone bruises in the coronal and sagittal planes using fat-suppressed T2-weighted images and a grid-based mapping technique of the tibiofemoral joint. RESULTS: A total of 109 patients were included, with 76 (69.7%) patients sustaining noncontact injuries and 33 (30.3%) patients sustaining contact injuries. There were no significant differences between the contact and noncontact groups in terms of age (11.8 ± 2.0 vs 12.4 ± 1.3 years; P = .12), male sex (90.9% vs 88.2%; P > .99), time from initial injury to MRI (10.3 ± 8.1 vs 10.4 ± 8.9 days; P = .84), the presence of a concomitant medial meniscus tear (18.2% vs 14.5%; P = .62) or lateral meniscus tear (69.7% vs 52.6%; P = .097), and sport-related injuries (82.9% vs 81.8%; P = .89). No significant differences were observed in the frequency of combined lateral tibiofemoral (lateral femoral condyle + lateral tibial plateau) bone bruises (87.9% contact vs 78.9% noncontact; P = .41) or combined medial tibiofemoral (medial femoral condyle [MFC] + medial tibial plateau) bone bruises (54.5% contact vs 35.5% noncontact; P = .064). Patients with contact ACL tears were significantly more likely to have centrally located MFC bruising (odds ratio, 4.3; 95% CI, 1.6-11; P = .0038) and less likely to have bruising on the anterior aspect of the lateral tibial plateau (odds ratio, 0.27; 95% CI, 0.097-0.76; P = .013). CONCLUSION: Children with contact ACL tears were 4 times more likely to present with centrally located MFC bone bruises on preoperative MRI scans compared with children who sustained noncontact ACL tears. Future studies should investigate the relationship between these bone bruise patterns and the potential risk of articular cartilage damage in pediatric patients with contact ACL tears.

Normative Running Kinematics in Healthy Adolescent Runners: A 2-Dimensional Video Analysis.

Background: The literature on the running kinematics of youth distance runners is limited. Purpose: We sought to describe 2-dimensional (2D) video analysis of running kinematics in healthy adolescent distance runners, which has not been previously described. Methods: We conducted an observational study of healthy, competitive runners between the ages of 14 and 18 years, prospectively recruited through local running clubs and our hospital's outreach between August 2019 and July 2023. Participants ran on a treadmill at a self-selected speed with markers attached to the thorax, pelvis, and lower extremities. A high-definition video camera recorded the runners in the sagittal and frontal planes. Kinematic measurements were completed using Dartfish software and reported as means and standard deviations. Results: Of the 53 participants (51% boys, mean age: 16.0 ± 1.4 years) included in the 2D running analysis, 91% ran with a rearfoot strike pattern, with a mean foot inclination angle of 10.2° ± 6.2°. Knee flexion angle at initial contact was 13.2° ± 3.8°, tibia inclination angle was 8.5° ± 3.2°, and peak knee flexion was 44.5° ± 3.6°. Cadence was 168.7° ± 8.6°. Contralateral pelvic drop was 6.0° ± 2.2° and peak rearfoot eversion was 11.8° ± 3.6°. Conclusions: This study is the first to describe running kinematics as captured by 2D video in healthy adolescent runners and to identify kinematic variables that may differ from those of adult runners. Further research is required to determine if adult recommendations are applicable to adolescent populations.

Performance and clinical utility of an artificial intelligence-enabled tool for pulmonary embolism detection.

PURPOSE: Diagnosing pulmonary embolism (PE) is still challenging due to other conditions that can mimic its appearance, leading to incomplete or delayed management and several inter-observer variabilities. This study evaluated the performance and clinical utility of an artificial intelligence (AI)-based application designed to assist clinicians in the detection of PE on CT pulmonary angiography (CTPA). PATIENTS AND METHODS: CTPAs from 230 US cities acquired on 57 scanner models from 6 different vendors were retrospectively collected. Three US board certified expert radiologists defined the ground truth by majority agreement. The same cases were analyzed by CINA-PE, an AI-driven algorithm capable of detecting and highlighting suspected PE locations. The algorithm's performance at a per-case and per-finding level was evaluated. Furthermore, cases with PE not mentioned in the clinical report but correctly detected by the algorithm were analyzed. RESULTS: A total of 1204 CTPAs (mean age 62.1 years ± 16.6[SD], 44.4 % female, 14.9 % positive) were included in the study. Per-case sensitivity and specificity were 93.9 % (95%CI: 89.3 %-96.9 %) and 94.8 % (95%CI: 93.3 %-96.1 %), respectively. Per-finding positive predictive value was 89.5 % (95%CI: 86.7 %-91.9 %). Among the 196 positive cases, 29 (15.6 %) were not mentioned in the clinical report. The algorithm detected 22/29 (76 %) of these cases, leading to a reduction in the miss rate from 15.6 % to 3.8 % (7/186). CONCLUSIONS: The AI-based application may improve diagnostic accuracy in detecting PE and enhance patient outcomes through timely intervention. Integrating AI tools in clinical workflows can reduce missed or delayed diagnoses, and positively impact healthcare delivery and patient care.

Evaluation of Tibial Slope on Radiographs in Pediatric Patients With Tibial Spine Fractures: An Age- and Sex-Matched Study.

Background: A recent study has reported that the radiographic measurement of posterior tibial slope (PTS) is larger in male pediatric patients with tibial spine fractures (TSF) than in controls. However, they found no difference in PTS between female patients and controls. Purpose: (1) To identify whether PTS is larger in female pediatric patients with TSF than in female controls and (2) to validate the relationship between PTS and pediatric TSF in male patients. Study Design: Cross-sectional study; Level of evidence, 3. Methods: After an a priori power analysis, 84 pediatric patients with TSF (50 female patients and 34 male patients) and 84 age- and sex-matched controls were enrolled in this study. Demographic information, including sex, age, and race, was recorded. Skeletal maturity was determined based on the stage of epiphyseal union on knee radiographs. PTS was defined as the angle between a line perpendicular to the longitudinal axis of the tibia and the posterior inclination of the medial tibial plateau on standard knee lateral radiographs. Results: The mean age when the TSF occurred was 11.2 ± 2.7 years for female patients and 12.9 ± 2.5 years for male patients. There was no significant difference in skeletal maturity between female patients and female controls or between male patients and male controls. The mean PTS was not significantly different between female patients (8.8°± 2.8°) and female controls (8.3°± 3.1°) (P = .366) or between male patients (9.0°± 2.8°) and male controls (9.3°± 2.6°) (P = .675). Those with a PTS >1 SD (2.9°) above the mean (8.8°) had no greater odds (1.0 [95% CI, 0.4-2.5]; P≥ .999) of having a TSF than others. Conclusion: PTS was not found to be a risk factor for pediatric TSF in female or male patients in this study.

Assessment of bronchodilator response in preschoolers: A systematic review.

BACKGROUND: Several techniques can be used to assess bronchodilator response (BDR) in preschool-aged children, including spirometry, respiratory oscillometry, the interrupter technique, and specific airway resistance. However, there has not been a systematic comparison of BDR thresholds across studies yet. METHODS: A systematic review was performed on all studies up to May 2023 measuring a bronchodilator effect in children 2-6 years old using one of these techniques (PROSPERO CRD42021264659). Studies were identified using MEDLINE, Cochrane, EMBASE, CINAHL via EBSCO, Web of Science databases, and reference lists of relevant manuscripts. RESULTS: Of 1224 screened studies, 43 were included. Over 85% were from predominantly European ancestry populations, and only 22 studies (51.2%) calculated a BDR cutoff based on a healthy control group. Five studies included triplicate testing with a placebo to account for the within-subject intrasession repeatability. A relative BDR was most consistently reported by the included studies (95%) but varied widely across all techniques. Various statistical methods were used to define a BDR, with six studies using receiver operating characteristic analyses to measure the discriminative power to distinguish healthy from wheezy and asthmatic children. CONCLUSION: A BDR in 2- to 6-year-olds cannot be universally defined based on the reviewed literature due to inconsistent methodology and cutoff calculations. Further studies incorporating robust methods using either distribution-based or clinical anchor-based approaches to define BDR are required.

Structural architecture of TolQ-TolR inner membrane protein complex from opportunistic pathogen Acinetobacter baumannii.

Gram-negative bacteria harness the proton motive force (PMF) within their inner membrane (IM) to uphold the integrity of their cell envelope, an indispensable aspect for both division and survival. The IM TolQ-TolR complex is the essential part of the Tol-Pal system, serving as a conduit for PMF energy transfer to the outer membrane. Here we present cryo-EM reconstructions of Acinetobacter baumannii TolQ in apo and TolR- bound forms at atomic resolution. The apo TolQ configuration manifests as a symmetric pentameric pore, featuring a trans-membrane funnel leading towards a cytoplasmic chamber. In contrast, the TolQ-TolR complex assumes a proton non-permeable stance, characterized by the TolQ pentamer's flexure to accommodate the TolR dimer, where two protomers undergo a translation-based relationship. Our structure-guided analysis and simulations support the rotor-stator mechanism of action, wherein the rotation of the TolQ pentamer harmonizes with the TolR protomers' interplay. These findings broaden our mechanistic comprehension of molecular stator units empowering critical functions within the Gram-negative bacterial cell envelope. Teaser: Apo TolQ and TolQ-TolR structures depict structural rearrangements required for cell envelope organization in bacterial cell division.

Three-Dimensional-Printed Patient-Specific Total Cuboid Replacement for Treatment of Post-traumatic Arthritis: A Case Report.

Cuboid injuries, including fractures, are rare and infrequently occur in isolation. Often, cuboid injuries can be treated nonoperatively. However, when surgery is indicated, appropriate management is necessary for maintaining the associated biomechanics of the midfoot. Current procedures for surgical management of the cuboid include open reduction and internal fixation, application of external fixation, or primary arthrodesis of the calcaneocuboid joint. Secondary procedures for symptomatic or poor outcomes of nonoperative and operative cuboid injuries consist of corrective osteotomy, bone resection, and interpositional arthroplasty. We present a novel surgical technique using a patient-specific three-dimensional-printed total cuboid replacement. This is an alternative treatment for post-traumatic arthritis of the cuboid along with a shortened lateral column. A single case example is given as well as details and discussion of the surgical technique.

Substrate-induced strain in molybdenum disulfide grown by aerosol-assisted chemical vapor deposition.

Transition metal dichalcogenides have been extensively studied in recent years because of their fascinating optical, electrical, and catalytic properties. However, low-cost, scalable production remains a challenge. Aerosol-assisted chemical vapor deposition (AACVD) provides a new method for scalable thin film growth. In this study, we demonstrate the growth of molybdenum disulfide (MoS2) thin films using AACVD method. This method proves its suitability for low-temperature growth of MoS2thin films on various substrates, such as glass, silicon dioxide, quartz, silicon, hexagonal boron nitride, and highly ordered pyrolytic graphite. The as-grown MoS2shows evidence of substrate-induced strain. The type of strain and the morphology of the as-grown MoS2highly depend on the growth substrate's surface roughness, crystallinity, and chemical reactivity. Moreover, the as-grown MoS2shows the presence of both direct and indirect band gaps, suitable for exploitation in future electronics and optoelectronics.

High-performance silicon photonic single-sideband modulators for cold-atom interferometry.

The laser system is the most complex component of a light-pulse atom interferometer (LPAI), controlling frequencies and intensities of multiple laser beams to configure quantum gravity and inertial sensors. Its main functions include cold-atom generation, state preparation, state-selective detection, and generating a coherent two-photon process for the light-pulse sequence. To achieve substantial miniaturization and ruggedization, we integrate key laser system functions onto a photonic integrated circuit. Our study focuses on a high-performance silicon photonic suppressed-carrier single-sideband (SC-SSB) modulator at 1560 nanometers, capable of dynamic frequency shifting within the LPAI. By independently controlling radio frequency (RF) channels, we achieve 30-decibel carrier suppression and unprecedented 47.8-decibel sideband suppression at peak conversion efficiency of -6.846 decibels (20.7%). We investigate imbalances in both amplitudes and phases between the RF signals. Using this modulator, we demonstrate cold-atom generation, state-selective detection, and atom interferometer fringes to estimate gravitational acceleration, g ≈ 9.77 ± 0.01 meters per second squared, in a rubidium (87Rb) atom system.

Proteomic biomarkers related to obesity in heart failure with reduced ejection fraction and their associations with outcome.

OBJECTIVE: Heart failure (HF) pathophysiology in patients with obesity may be distinct. To study these features, we identified obesity-related biomarkers from 4210 circulating proteins in patients with HF with reduced ejection fraction (HFrEF) and examined associations of these proteins with HF prognosis and biological mechanisms. METHODS: In 373 patients with trimonthly blood sampling during a median follow-up of 2.1 (25th-75th percentile: 1.1-2.6) years, we applied an aptamer-based multiplex approach measuring 4210 proteins in baseline samples and the last two samples before study end. Associations between obesity (BMI > 30 kg/m2) and baseline protein levels were analyzed. Subsequently, associations of serially measured obesity-related proteins with biological mechanisms and the primary endpoint (PEP; composite of cardiovascular mortality, HF hospitalization, left ventricular assist device implantation, and heart transplantation) were examined. RESULTS: Obesity was identified in 26% (96/373) of patients. A total of 30% (112/373) experienced a PEP (with obesity: 26% [25/96] vs. without obesity: 31% [87/277]). A total of 141/4210 proteins were linked to obesity, reflecting mechanisms of neuron projection development, cell adhesion, and muscle cell migration. A total of 50/141 proteins were associated with the PEP, of which 12 proteins related to atherosclerosis or hypertrophy provided prognostic information beyond clinical characteristics, N-terminal pro-B-type natriuretic peptide, and high-sensitivity troponin T. CONCLUSIONS: Patients with HFrEF and obesity show distinct proteomic profiles compared to patients with HFrEF without obesity. Obesity-related proteins are independently associated with HF outcome. These proteins carry potential to improve management of obesity-related HF and could be leads for future research.

"Expected to happen": perspectives on post-release overdose from recently incarcerated people with opioid use disorder.

BACKGROUND: Opioid-related overdose is the leading cause of death for people recently released from incarceration, however treatment with medications for opioid use disorder (MOUD) during incarceration can reduce the mortality risk. This study seeks to qualitatively analyze perceptions of post-release overdose risk from the perspectives of people who received MOUD while incarcerated in one of eight Massachusetts jails during 2021-2022 using the Risk Environment Framework to guide analyses. METHODS: N = 38 participants with lived experience of MOUD treatment during incarceration who are now living in the community were interviewed on factors that may contribute to or protect against post-release overdose risk. Themes were identified inductively and deductively using the Risk Environment Framework and its domains, which organizes themes along physical, social, economic, and policy environments on both the micro- and macro- scales. RESULTS: The physical risk environment included loss of opioid tolerance during incarceration, polysubstance use, and the toxicity of the regional drug supply as key producers of increased risk for post-release overdose. Social drivers of risk included peer group risk norms-including peer-driven harm reduction practices and interpersonal relationships between drug sellers and buyers-as well as macro-level social determinants of health such as housing insecurity and availability of mental health services. Economic drivers of post-release overdose risk included lack of income generation during incarceration and employment challenges. Participants discussed several aspects of policy that contribute to post-release overdose risk, including availability of harm reduction supplies, public health services, and broader policy around MOUD. CONCLUSIONS: The perspectives of people with lived experience are vital to understanding the disproportionate risks of overdose for those recently released from incarceration. Our results highlight the intersectional factors that produce and reproduce the post-release overdose risk environment, providing support for interventions across each domain of the Risk Environment Framework. By capturing perspectives from people with lived experience of OUD and incarceration during this critical period of risk, we can better identify interventions that target and mitigate overdose-related harm in this population.

An evaluation of ChatGPT and Bard (Gemini) in the context of biological knowledge retrieval.

ChatGPT and Bard (now called Gemini), two conversational AI models developed by OpenAI and Google AI, respectively, have garnered considerable attention for their ability to engage in natural language conversations and perform various language-related tasks. While the versatility of these chatbots in generating text and simulating human-like conversations is undeniable, we wanted to evaluate their effectiveness in retrieving biological knowledge for curation and research purposes. To do so we asked each chatbot a series of questions and scored their answers based on their quality. Out of a maximal score of 24, ChatGPT scored 5 and Bard scored 13. The encountered issues included missing information, incorrect answers, and instances where responses combine accurate and inaccurate details. Notably, both tools tend to fabricate references to scientific papers, undermining their usability. In light of these findings, we recommend that biologists continue to rely on traditional sources while periodically assessing the reliability of ChatGPT and Bard. As ChatGPT aptly suggested, for specific and up-to-date scientific information, established scientific journals, databases, and subject-matter experts remain the preferred avenues for trustworthy data.

The Genome Explorer genome browser.

Are two adjacent genes in the same operon? What are the order and spacing between several transcription factor binding sites? Genome browsers are software data visualization and exploration tools that enable biologists to answer questions such as these. In this paper, we report on a major update to our browser, Genome Explorer, that provides nearly instantaneous scaling and traversing of a genome, enabling users to quickly and easily zoom into an area of interest. The user can rapidly move between scales that depict the entire genome, individual genes, and the sequence; Genome Explorer presents the most relevant detail and context for each scale. By downloading the data for the entire genome to the user's web browser and dynamically generating visualizations locally, we enable fine control of zoom and pan functions and real-time redrawing of the visualization, resulting in smoother and more intuitive exploration of a genome than is possible with other browsers. Further, genome features are presented together, in-line, using familiar graphical depictions. In contrast, many other browsers depict genome features using data tracks, which have low information density and can visually obscure the relative positions of features. Genome Explorer diagrams have a high information density that provides larger amounts of genome context and sequence information to be presented in a given-sized monitor than for tracks-based browsers. Genome Explorer provides optional data tracks for the analysis of large-scale data sets and a unique comparative mode that aligns genomes at orthologous genes with synchronized zooming. IMPORTANCE: Genome browsers provide graphical depictions of genome information to speed the uptake of complex genome data by scientists. They provide search operations to help scientists find information and zoom operations to enable scientists to view genome features at different resolutions. We introduce the Genome Explorer browser, which provides extremely fast zooming and panning of genome visualizations and displays with high information density.

Multidimensional Analysis of the Adult Human Heart in Health and Disease Using Hierarchical Phase-Contrast Tomography.

Background Current clinical imaging modalities such as CT and MRI provide resolution adequate to diagnose cardiovascular diseases but cannot depict detailed structural features in the heart across length scales. Hierarchical phase-contrast tomography (HiP-CT) uses fourth-generation synchrotron sources with improved x-ray brilliance and high energies to provide micron-resolution imaging of intact adult organs with unprecedented detail. Purpose To evaluate the capability of HiP-CT to depict the macro- to microanatomy of structurally normal and abnormal adult human hearts ex vivo. Materials and Methods Between February 2021 and September 2023, two adult human donor hearts were obtained, fixed in formalin, and prepared using a mixture of crushed agar in a 70% ethanol solution. One heart was from a 63-year-old White male without known cardiac disease, and the other was from an 87-year-old White female with a history of multiple known cardiovascular pathologies including ischemic heart disease, hypertension, and atrial fibrillation. Nondestructive ex vivo imaging of these hearts without exogenous contrast agent was performed using HiP-CT at the European Synchrotron Radiation Facility. Results HiP-CT demonstrated the capacity for high-spatial-resolution, multiscale cardiac imaging ex vivo, revealing histologic-level detail of the myocardium, valves, coronary arteries, and cardiac conduction system across length scales. Virtual sectioning of the cardiac conduction system provided information on fatty infiltration, vascular supply, and pathways between the cardiac nodes and adjacent structures. HiP-CT achieved resolutions ranging from gross (isotropic voxels of approximately 20 µm) to microscopic (approximately 6.4-µm voxel size) to cellular (approximately 2.3-µm voxel size) in scale. The potential for quantitative assessment of features in health and disease was demonstrated. Conclusion HiP-CT provided high-spatial-resolution, three-dimensional images of structurally normal and diseased ex vivo adult human hearts. Whole-heart image volumes were obtained with isotropic voxels of approximately 20 µm, and local regions of interest were obtained with resolution down to 2.3-6.4 µm without the need for sectioning, destructive techniques, or exogenous contrast agents. Published under a CC BY 4.0 license Supplemental material is available for this article. See also the editorial by Bluemke and Pourmorteza in this issue.

Performance of pan-enteroscopy in children with intestinal failure due to short bowel syndrome: A single-center retrospective study.

OBJECTIVES: In pediatric patients with intestinal failure (IF) due to short bowel syndrome, we hypothesized that young children, those with shorter residual small bowel and those with congenital malrotation of the bowel would be more likely to undergo pan-enteroscopy. We aimed to determine the feasibility and diagnostic yield of pan-enteroscopy in this cohort. METHODS: We performed a single-center, retrospective study of pediatric patients with IF due to short bowel syndrome (SBS) who had undergone at least one GI endoscopic evaluation between January 1, 2018 and January 1, 2023. RESULTS: A pan-enteroscopy might have been possible in 381 of the 431 procedures (206 patients) reviewed. 44 (21%) patients underwent 54 pan-enteroscopies. Children with a residual bowel length <35 cm had higher odds of undergoing pan-enteroscopy (odds ratio [OR] 3.72, 95% confidence interval [CI] [1.32, 10.48], p = 0.01), as did patients with periprocedural glucagon-like peptide 2 (GLP-2) analog use (OR 4.30, 95% CI [1.24, 14.95], p = 0.02). Patients with diagnoses other than necrotizing enterocolitis (NEC) tended to be more likely to achieve a pan-enteroscopy (OR 2.73, 95% CI [0.95,7.88], p = 0.06). Evidence of gross and histopathologic abnormalities were found in 77.8% and 78% of the procedures, respectively. No complications were identified. CONCLUSION: In a large cohort of children with SBS, pan-enteroscopy was successfully performed in 14.2% of the procedures and microscopic abnormalities were common. Shorter residual bowel length, underlying diagnoses of non-NEC, and GLP-2 analog use were generally associated with successful pan-enteroscopy, independent of age and several other factors. These data suggest that pan-enteroscopy is feasible and of high-yield in a subset of patients with SBS.

Small molecule FICD inhibitors suppress endogenous and pathologic FICD-mediated protein AMPylation.

The AMP transferase, FICD, is an emerging drug target finetuning stress signaling in the endoplasmic reticulum (ER). FICD is a bi-functional enzyme, catalyzing both AMP addition (AMPylation) and removal (deAMPylation) from the ER resident chaperone BiP/GRP78. Despite increasing evidence linking excessive BiP/GRP78 AMPylation to human diseases, small molecules to inhibit pathogenic FICD variants are lacking. Using an in-vitro high-throughput screen, we identify two small-molecule FICD inhibitors, C22 and C73. Both molecules significantly inhibit FICD-mediated BiP/GRP78 AMPylation in intact cells while only weakly inhibiting BiP/GRP78 deAMPylation. C22 and C73 also efficiently inhibit pathogenic FICD variants and improve proinsulin processing in ß cells. Our study identifies and validates FICD inhibitors, highlighting a novel therapeutic avenue against pathologic protein AMPylation.

HIV-1-envelope trimer transitions from prefusion-closed to CD4-bound-open conformations through an occluded-intermediate state.

HIV-1 infection is initiated by the interaction between the gp120 subunit in the envelope (Env) trimer and the cellular receptor CD4 on host cells. This interaction induces substantial structural rearrangement of the Env trimer. Currently, static structural information for prefusion-closed trimers, CD4-bound prefusion-open trimers, and various antibody-bound trimers is available. However, dynamic features between these static states (e.g., transition structures) are not well understood. Here, we investigate the full transition pathway of a site specifically glycosylated Env trimer between prefusion-closed and CD4-bound-open conformations by collective molecular dynamics and single-molecule Förster resonance energy transfer (smFRET). Our investigations reveal and confirm important features of the transition pathway, including movement of variable loops to generate a glycan hole at the trimer apex and formation or rearrangements of α-helices and ß-strands. Notably, by comparing the transition pathway to known Env-structures, we uncover evidence for a transition intermediate, with four antibodies, Ab1303, Ab1573, b12, and DH851.3, recognizing this intermediate. Each of these four antibodies induce population shifts of Env to occupy a newly observed smFRET state: the "occluded-intermediate" state. We propose this occluded-intermediate state to be both a prevalent state of Env and an on-path conformation between prefusion-closed and CD4-bound-open states, previously overlooked in smFRET analyses.