Publications search

Global population immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is accumulating through heterogeneous combinations of infection and vaccination. Vaccine distribution in low- and middle-income countries has been variable and reliant on diverse vaccine platforms. We studied B-cell immunity in Mexico, a middle-income country where five different vaccines have been deployed to populations with high SARS-CoV-2 incidences. Levels of antibodies that bound a stabilized prefusion spike trimer, neutralizing antibody titers, and memory B-cell expansion correlated with each other across vaccine platforms. Nevertheless, the vaccines elicited variable levels of B-cell immunity, and the majority of recipients had undetectable neutralizing activity against the recently emergent omicron variant. SARS-CoV-2 infection, experienced before or after vaccination, potentiated B-cell immune responses and enabled the generation of neutralizing activity against omicron and SARS-CoV for all vaccines in nearly all individuals. These findings suggest that broad population immunity to SARS-CoV-2 will eventually be achieved but by heterogeneous paths. IMPORTANCE The majority of studies on SARS-CoV-2 vaccine-elicited immunity and immune evasion have focused on single vaccines corresponding to those distributed in high-income countries. However, in low- and middle-income countries, vaccine deployment has been far less uniform. It is therefore important to determine the levels of immunity elicited by vaccines that have been deployed globally. Such data should help inform policy. Thus, this paper is very much a "real-world" study that focuses on a middle-income country, Mexico, in which five different vaccines based on mRNA, adenovirus, and inactivated-virus platforms have been extensively deployed, while (as documented in our study) SARS-CoV-2 variants with increasing degrees of immune evasiveness have propagated in the Mexican population, culminating in the recent emergence of B.1.1.529 (omicron). The majority of studies on SARS-CoV-2 vaccine-elicited immunity and immune evasion have focused on single vaccines corresponding to those distributed in high-income countries. However, in low- and middle-income countries, vaccine deployment has been far less uniform.

Proteogenomic identification of translated small open reading frames has revealed thousands of previously unannotated, largely uncharacterized microproteins, or polypeptides of less than 100 amino acids, and alternative proteins (alt-proteins) that are co-encoded with canonical proteins and are often larger. The subcellular localizations of microproteins and alt-proteins are generally unknown but can have significant implications for their functions. Proximity biotinylation is an attractive approach to define the protein composition of subcellular compartments in cells and in animals. Here, we developed a high-throughput technology to map unannotated microproteins and alt-proteins to subcellular localizations by proximity biotinylation with TurboID (MicroID). More than 150 microproteins and alt-proteins are associated with subnuclear organelles. One alt-protein, alt-LAMA3, localizes to the nucleolus and functions in pre-rRNA transcription. We applied MicroID in a mouse model, validating expression of a conserved nuclear microprotein, and establishing MicroID for discovery of microproteins and alt-proteins in vivo.

HK022 put is an RNA element that inhibits transcription termination without aids from protein factors. Here, authors solved cryo-EM structures of put-associated RNA polymerase and showed the structure of putRNA and its binding to the RNA polymerase. Transcription, in which RNA polymerases (RNAPs) produce RNA from DNA, is the first step of gene expression. As such, it is highly regulated either by trans-elements like protein factors and/or by cis-elements like specific sequences on the DNA. Lambdoid phage HK022 contains a cis-element, put, which suppresses pausing and termination during transcription of the early phage genes. The putRNA transcript solely performs the anti-pausing/termination activities by interacting directly with the E.coli RNAP elongation complex (EC) by an unknown structural mechanism. In this study, we reconstituted putRNA-associated ECs and determined the structures using cryo-electron microscopy. The determined structures of putRNA-associated EC, putRNA-absent EC, and sigma(70)-bound EC suggest that the putRNA interaction with the EC counteracts swiveling, a conformational change previously identified to promote pausing and sigma(70) might modulate putRNA folding via sigma(70)-dependent pausing during elongation.

Metabolic plasticity is the ability of a biological system to adapt its metabolic phenotype to different environmental stressors. We used a whole-body and tissue-specific phenotypic, functional, proteomic, metabolomic and transcriptomic approach to systematically assess metabolic plasticity in diet-induced obese mice after a combined nutritional and exercise intervention. Although most obesity and overnutrition-related pathological features were successfully reverted, we observed a high degree of metabolic dysfunction in visceral white adipose tissue, characterized by abnormal mitochondrial morphology and functionality. Despite two sequential therapeutic interventions and an apparent global healthy phenotype, obesity triggered a cascade of events in visceral adipose tissue progressing from mitochondrial metabolic and proteostatic alterations to widespread cellular stress, which compromises its biosynthetic and recycling capacity. In humans, weight loss after bariatric surgery showed a transcriptional signature in visceral adipose tissue similar to our mouse model of obesity reversion. Overall, our data indicate that obesity prompts a lasting metabolic fingerprint that leads to a progressive breakdown of metabolic plasticity in visceral adipose tissue.

Motor circuits develop in sequence from those governing fast movements to those governing slow. Here we examine whether upstream sensory circuits are organized by similar principles. Using serial-section electron microscopy in larval zebrafish, we generated a complete map of the gravity-sensing (utricular) system spanning from the inner ear to the brainstem. We find that both sensory tuning and developmental sequence are organizing principles of vestibular topography. Patterned rostrocaudal innervation from hair cells to afferents creates an anatomically inferred directional tuning map in the utricular ganglion, forming segregated pathways for rostral and caudal tilt. Furthermore, the mediolateral axis of the ganglion is linked to both developmental sequence and neuronal temporal dynamics. Early-born pathways carrying phasic information preferentially excite fast escape circuits, whereas later-born pathways carrying tonic signals excite slower postural and oculomotor circuits. These results demonstrate that vestibular circuits are organized by tuning direction and dynamics, aligning them with downstream motor circuits and behaviors. How sensory systems are organized during development remains unclear. Here, the authors used electron microscopy to examine the gravity-sensing system in zebrafish, finding that directional tuning and developmental age are organizing principles of the transformation from vestibular sensation to motor control.

Iron is the most abundant transition metal essential for numerous cellular processes. Although most mammalian cells acquire iron through transferrin receptors, molecular players of iron utilization under iron restriction are incompletely understood. To address this, we performed metabolism-focused CRISPRa gain-of-function screens, which revealed metabolic limitations under stress conditions. Iron restriction screens identified not only expected members of iron utilization pathways but also SLCO2B1, a poorly characterized membrane carrier. SLCO2B1 expression is sufficient to increase intracellular iron, bypass the essentiality of the transferrin receptor, and enable proliferation under iron restriction. Mechanistically, SLCO2B1 mediates heme analog import in cellular assays. Heme uptake by SLCO2B1 provides sufficient iron for proliferation through heme oxygenases. Notably, SLCO2B1 is predominantly expressed in microglia in the brain, and primary Slco2b1(-/-) mouse microglia exhibit strong defects in heme analog import. Altogether, our work identifies SLCO2B1 as a microglia-enriched plasma membrane heme importer and provides a genetic platform to identify metabolic limitations under stress conditions.

Estrogens affect dopamine-dependent diseases/behavior and have rapid effects on dopamine release and receptor availability in the nucleus accumbens (NAc). Low levels of nuclear estrogen receptor (ER) alpha and ER beta are seen in the NAc, which cannot account for the rapid effects of estrogens in this region. G-protein coupled ER 1 (GPER1) is observed at low levels in the NAc shell, which also likely does not account for the array of estrogens' effects in this region. Prior studies demonstrated membrane-associated ERs in the dorsal striatum; these experiments extend those findings to the NAc core and shell. Single- and dual-immunolabeling electron microscopy determined whether ER alpha, ER beta, and GPER1 are at extranuclear sites in the NAc core and shell and whether ER alpha and GPER1 were localized to catecholaminergic or gamma-aminobutyric acid-ergic (GABAergic) neurons. All three ERs are observed, almost exclusively, at extranuclear sites in the NAc, and similarly distributed in the core and shell. ER alpha, ER beta, and GPER1 are primarily in axons and axon terminals suggesting that estrogens affect transmission in the NAc via presynaptic mechanisms. About 10% of these receptors are found on glia. A small proportion of ER alpha and GPER1 are localized to catecholaminergic terminals, suggesting that binding at these ERs alters release of catecholamines, including dopamine. A larger proportion of ER alpha and GPER1 are localized to GABAergic dendrites and terminals, suggesting that estrogens alter GABAergic transmission to indirectly affect dopamine transmission in the NAc. Thus, the localization of ERs could account for the rapid effects of estrogen in the NAc.

Background Hidradenitis suppurativa (HS) is an inflammatory skin disease with dysregulation of the interleukin (IL)-17 axis. Recently, we reported the clinical benefit of brodalumab, a human anti-IL-17 receptor A (IL-17RA) monoclonal antibody, in moderate-to-severe HS. Objectives To characterize the molecular response to brodalumab in HS skin and serum, and to identify biomarkers of treatment response. Methods Ten participants, who received brodalumab 210 mg /1 center dot 5 mL subcutaneously at weeks 0, 1, 2, 4 and every 2 weeks thereafter, were included in this molecular profiling study (NCT03960268). RNA sequencing and immunohistochemistry of nonlesional, perilesional and lesional HS skin biopsies, and Olink high-throughput proteomics of serum at baseline, weeks 4 and 12 were assessed. Results At week 12, brodalumab led to a decrease of overall inflammation, and improvement of psoriasis-, keratinocyte- and neutrophil-related pathways. Despite perilesional and lesional skin exhibiting no differentially expressed genes at baseline, treatment response was best assessed in perilesional skin. In serum, brodalumab treatment decreased pathways involved in neutrophil inflammation. Patients with higher baseline expression of neutrophil-associated lipocalin-2 (LCN2) in the skin or IL-17A in the serum demonstrated greater decreases of HS-related inflammatory cytokines as measured in skin biopsies at week 12. Conclusions IL-17RA inhibition by brodalumab decreases several pathogenic inflammatory axes in HS. Perilesional skin provides a valid and robust assessment of treatment response. Expression of LCN2 in skin or IL-17A in serum may be used as biomarkers to stratify patients that may have a superior molecular response to brodalumab. What is already known about this topic? Hidradenitis suppurativa (HS) is a chronic inflammatory disease with upregulation of several pro-inflammatory and keratinocyte proliferation-related pathways. Interleukin (IL)-17 is a known modulator of these pathways and has been implicated in the pathogenesis of HS. We have recently reported a clinical improvement in HS with brodalumab, an IL-17RA antagonist. What does this study add? This study details the molecular response to brodalumab using RNA sequencing, high-throughput proteomics and immunohistochemistry. This research identifies lipocalin-2 (LCN2) in skin and IL-17A in serum as potential predictive biomarkers for treatment response. These data may help guide physicians in choosing the appropriate therapy for patients with moderate-to-severe HS. What is the translational message? IL-17RA blockade with brodalumab is an effective treatment for patients with moderate-to-severe HS. Assessing pathways that respond to treatment, rather than individual biomarkers, may be a superior approach for molecular characterization of the treatment response in HS therapeutic studies. Brodalumab modulates neutrophil-related pathways in the skin and blood, and known psoriasis pathways in the skin. We demonstrate that patients with high baseline expression of IL-17-regulated neutrophil-created LCN2 in the skin or IL-17A in the serum have a superior molecular response to treatment, allowing the possibility for physicians to select therapeutic treatment based on baseline molecular profiles.

In yeast and animals, cyclin B binds and activates the cyclin-dependent kinase ('CDK') CDK1 to drive entry into mitosis. We show that CYCB1, the sole cyclin B in Chlamydomonas, activates the plant-specific CDKB1 rather than the CDK1 ortholog CDKA1, confirming and extending previous results. Time-lapse microscopy shows that CYCB1 is synthesized before each division in the multiple fission cycle, then is rapidly degraded 3-5 minutes before division occurs. CYCB1 degradation is dependent on the anaphase-promoting complex (APC). Like CYCB1, CDKB1 is not synthesized until late G1; however, CDKB1 is not degraded with each division within the multiple fission cycle, but is degraded after all divisions have ceased. The microtubule plus-end-binding protein EB1 labeled with mNeon-Green allowed detection of mitotic events in live cells. The earliest detectable step in mitosis, splitting of polar EB1 signal into two foci, likely associated with future spindle poles, was dependent on CYCB1. CYCB1-GFP localized close to these foci immediately before spindle formation. Spindle breakdown, cleavage furrow formation and accumulation of EB1 in the furrow were dependent on the APC. In interphase, rapidly growing microtubules are marked by 'comets' of EB1; comets are absent in the absence of APC function. Thus CYCB1/CDKB1 and the APC modulate microtubule function and assembly while regulating mitotic progression. Genetic results suggest an independent additional role for the APC in regulating sister chromatid cohesion; this role is likely conserved across eukaryotes.