Tracking antigen-specific TCR clonotypes in SARS-CoV-2 infection reveals distinct severity trajectories.

Despite the importance of antigen-specific T cells in infectious disease, characterizing and tracking clonally amplified T cells during the progression of a patient's symptoms remain unclear. Here, we performed a longitudinal, in-depth single-cell multiomics analysis of samples from asymptomatic, mild, usual severe, and delayed severe patients of SARS-CoV-2 infection. Our in-depth analysis revealed that hyperactive or improper T-cell responses were more aggressive in delayed severe patients. Interestingly, tracking of antigen-specific T-cell receptor (TCR) clonotypes along the developmental trajectory indicated an attenuation in functional T cells upon severity. In addition, increased glycolysis and interleukin-6 signaling in the cytotoxic T cells were markedly distinct in delayed severe patients compared to usual severe patients, particularly in the middle and late stages of infection. Tracking B-cell receptor clonotypes also revealed distinct transitions and somatic hypermutations within B cells across different levels of disease severity. Our results suggest that single-cell TCR clonotype tracking can distinguish the severity of patients through immunological hallmarks, leading to a better understanding of the severity differences in and improving the management of infectious diseases by analyzing the dynamics of immune responses over time.

Single-Cell Molecular Barcoding to Decode Multimodal Information Defining Cell States.

Single-cell research has provided a breakthrough in biology to understand heterogeneous cell groups, such as tissues and organs, in development and disease. Molecular barcoding and subsequent sequencing technology insert a singlecell barcode into isolated single cells, allowing separation cell by cell. Given that multimodal information from a cell defines precise cellular states, recent technical advances in methods focus on simultaneously extracting multimodal data recorded in different biological materials (DNA, RNA, protein, etc.). This review summarizes recently developed singlecell multiomics approaches regarding genome, epigenome, and protein profiles with the transcriptome. In particular, we focus on how to anchor or tag molecules from a cell, improve throughputs with sample multiplexing, and record lineages, and we further discuss the future developments of the technology.

Comparable humoral and cellular immunity against Omicron variant BA.4/5 of once-boosted BA.1/2 convalescents and twice-boosted COVID-19-naïve individuals.

The fourth vaccination dose confers additional protective immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in individuals with no prior coronavirus disease-19 (COVID-19). However, its immunological benefit against currently circulating BA.4/5 is unclear in individuals who have received a booster shot and been infected with Omicron variant BA.1/2. We analyzed immune responses in whom had been boosted once and did not have COVID-19 (n = 16), boosted once and had COVID-19 when BA.1/2 was dominant in Korea (Hybrid-6M group, n = 27), and boosted twice and did not have COVID-19 (Vx4 group, n = 15). Antibody binding activities against RBDo BA.1 and RBDo BA.4/5 , antigen-specific memory CD4+ and CD8+ T-cell responses against BA.4/5, and B-cell responses against SARS-CoV-2 wild-type did not differ statistically between the Hybrid-6M and Vx4 groups. The humoral and cellular immune responses of the Hybrid-6M group against BA.4/5 were comparable to those of the Vx4 group. Individuals who had been boosted and had an Omicron infection in early 2022 may not have high priority for an additional vaccination.

Nanocomplexes of doxorubicin and DNA fragments for efficient and safe cancer chemotherapy.

Cancer-targeted therapy by a chemotherapeutic agent formulated in a nanoscale platform has been challenged by complex and inefficient manufacturing, low drug loading, difficult characterization, and marginally improved therapeutic efficacy. This study investigated facile-to-produce nanocomplexes of doxorubicin (DOX), a widely used cancer drug, and clinically approved DNA fragments that are extracted from a natural source. DOX was found to self-assemble DNA fragments into relatively monodispersed nanocomplexes with a diameter of ∼70 nm at 14.3% (w/w) drug loading by simple and scalable mixing. The resulting DOX/DNA nanocomplexes showed sustained DOX release, unlike overly stable Doxil®, cellular uptake via multiple endocytosis pathways, and high hematological and immunological compatibility. DOX/DNA nanocomplexes eradicated EL4 T lymphoma cells in a time-dependent manner, eventually surpassing free DOX. Extended circulation of DOX/DNA nanocomplexes, while avoiding off-target accumulation in the lung and being cleared from the liver, resulted in rapid accumulation in tumor and lowered cardio toxicity. Finally, tumor growth of EL4-challenged C57BL/6 mice (syngeneic model) and OPM2-challenged NSG mice (human xenograft model) were efficiently inhibited by DOX/DNA nanocomplexes with enhanced overall survival, in comparison with free DOX and Doxil®, especially upon repeated administrations. DOX/DNA nanocomplexes are a promising chemotherapeutics delivery platform for their ease of manufacturing, high biocompatibility, desired drug release and accumulation, efficient tumor eradication with improved safety, and further engineering versatility for extended therapeutic applications.

Amorphous Carbon Films for Electronic Applications.

While various crystalline carbon allotropes, including graphene, have been actively investigated, amorphous carbon (a-C) thin films have received relatively little attention. The a-C is a disordered form of carbon bonding with a broad range of the CC bond length and bond angle. Although accurate structural analysis and theoretical approaches are still insufficient, reproducible structure-property relationships have been accumulated. As the a-C thin film is now adapted as a hardmask in the semiconductor industry and new properties are reported continuously, expectations are growing that it can be practically used as active materials beyond as a simple sacrificial layer. In this perspective review article, after a brief introduction to the synthesis and properties of the a-C thin films, their potential practical applications are proposed, including hardmasks, extreme ultraviolet (EUV) pellicles, diffusion barriers, deformable electrodes and interconnects, sensors, active layers, electrodes for energy, micro-supercapacitors, batteries, nanogenerators, electromagnetic interference (EMI) shielding, and nanomembranes. The article ends with a discussion on the technological challenges in a-C thin films.

DNA Barcoding Technology for Lineage Recording and Tracing to Resolve Cell Fate Determination.

In various biological contexts, cells receive signals and stimuli that prompt them to change their current state, leading to transitions into a future state. This change underlies the processes of development, tissue maintenance, immune response, and the pathogenesis of various diseases. Following the path of cells from their initial identity to their current state reveals how cells adapt to their surroundings and undergo transformations to attain adjusted cellular states. DNA-based molecular barcoding technology enables the documentation of a phylogenetic tree and the deterministic events of cell lineages, providing the mechanisms and timing of cell lineage commitment that can either promote homeostasis or lead to cellular dysregulation. This review comprehensively presents recently emerging molecular recording technologies that utilize CRISPR/Cas systems, base editing, recombination, and innate variable sequences in the genome. Detailing their underlying principles, applications, and constraints paves the way for the lineage tracing of every cell within complex biological systems, encompassing the hidden steps and intermediate states of organism development and disease progression.

NaCl Ionization-Based Moisture Sensor Prepared by Aerosol Deposition for Monitoring Respiratory Patterns.

A highly polarizable moisture sensor with multimodal sensing capabilities has great advantages for healthcare applications such as human respiration monitoring. We introduce an ionically polarizable moisture sensor based on NaCl/BaTiO3 composite films fabricated using a facile aerosol deposition (AD) process. The proposed sensing model operates based on an enormous NaCl ionization effect in addition to natural moisture polarization, whereas all previous sensors are based only on the latter. We obtained an optimal sensing performance in a 0.5 µm-thick layer containing NaCl-37.5 wt% by manipulating the sensing layer thickness and weight fraction of NaCl. The NaCl/BaTiO3 sensing layer exhibits outstanding sensitivity over a wide humidity range and a fast response/recovery time of 2/2 s; these results were obtained by performing the one-step AD process at room temperature without using any auxiliary methods. Further, we present a human respiration monitoring system using a sensing device that provides favorable and stable electrical signals under diverse respiratory scenarios.

Corrigendum: Distinct Immune Response at 1 Year Post-COVID-19 According to Disease Severity.

[This corrects the article DOI: 10.3389/fimmu.2022.830433.].

Distinct Immune Response at 1 Year Post-COVID-19 According to Disease Severity.

Background: Despite the fact of ongoing worldwide vaccination programs for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), understanding longevity, breadth, and type of immune response to coronavirus disease-19 (COVID-19) is still important to optimize the vaccination strategy and estimate the risk of reinfection. Therefore, we performed thorough immunological assessments 1 year post-COVID-19 with different severity. Methods: We analyzed peripheral blood mononuclear cells and plasma samples at 1 year post-COVID-19 in patients who experienced asymptomatic, mild, and severe illness to assess titers of various isotypes of antibodies (Abs) against SARS-CoV-2 antigens, phagocytic capability, and memory B- and T-cell responses. Findings: A total of 24 patients (7, 9, and 8 asymptomatic, mild, and severe patients, respectively) and eight healthy volunteers were included in this study. We firstly showed that disease severity is correlated with parameters of immune responses at 1 year post-COVID-19 that play an important role in protecting against reinfection with SARS-CoV-2, namely, the phagocytic capacity of Abs and memory B-cell responses. Interpretation: Various immune responses at 1 year post-COVID-19, particularly the phagocytic capacity and memory B-cell responses, were dependent on the severity of the prior COVID-19. Our data could provide a clue for a tailored vaccination strategy after natural infection according to the severity of COVID-19.

Morphological, Ecological, and Molecular Divergence of Conogethes pinicolalis from C. punctiferalis (Lepidoptera: Crambidae).

Conogethes pinicolalis has long been considered as a Pinaceae-feeding type of the yellow peach moth, C. punctiferalis, in Korea. In this study, the divergence of C. pinicolalis from the fruit-feeding moth C. punctiferalis was analyzed in terms of morphology, ecology, and genetics. C. pinicolalis differs from C. punctiferalis in several morphological features. Through field observation, we confirmed that pine trees are the host plants for the first generation of C. pinicolalis larvae, in contrast to fruit-feeding C. punctiferalis larvae. We successfully reared C. pinicolalis larvae to adults by providing them pine needles as a diet. From a genetic perspective, the sequences of mitochondrial COI of these two species substantially diverged by an average of 5.46%; moreover, phylogenetic analysis clearly assigned each species to an independent clade. On the other hand, nuclear EF1α showed a lower sequence divergence (2.10%) than COI. Overall, EF1α-based phylogenetic analysis confirmed each species as an independent clade, but a few haplotypes of EF1α indicated incomplete lineage sorting between these two species. In conclusion, our results demonstrate that C. pinicolalis is an independent species according to general taxonomic criteria; however, analysis of the EF1α sequence revealed a short divergence time.

Longitudinal Analysis of Human Memory T-Cell Response According to the Severity of Illness up to 8 Months After Severe Acute Respiratory Syndrome Coronavirus 2 Infection.

BACKGROUND: Understanding the memory T-cell response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is crucial for assessing the longevity of protective immunity after SARS-CoV-2 infection or coronavirus disease 2019 (COVID-19) vaccination. However, the longitudinal memory T-cell response up to 8 months post-symptom onset (PSO) according to the severity of illness is unknown. METHODS: We analyzed peripheral blood mononuclear cells (PBMCs) from healthy volunteers or patients with COVID-19 who experienced asymptomatic, mild, or severe illness at 2, 5, and 8 months PSO. SARS-CoV-2 spike, nucleocapsid, and membrane protein-stimulated PBMCs were subjected to flow cytometry analysis. RESULTS: A total of 24 patients (7 asymptomatic, 9 with mild disease, and 8 with severe disease) and 6 healthy volunteers were analyzed. SARS-CoV-2-specific OX40+CD137+CD4+ T cells and CD69+CD137+CD8+ T cells persisted at 8 months PSO. Also, antigen-specific cytokine-producing or polyfunctional CD4+ T cells were maintained for up to 8 months PSO. Memory CD4+ T-cell responses tended to be greater in patients who had severe illness than in those with mild or asymptomatic disease. CONCLUSIONS: Memory response to SARS-CoV-2, based on the frequency and functionality, persists for 8 months PSO. Further investigations involving its longevity and protective effect from reinfection are warranted.

Unique Noncontact Monitoring of Human Respiration and Sweat Evaporation Using a CsPb2Br5-Based Sensor.

Respiration monitoring and human sweat sensing have promising application prospects in personal healthcare data collection, disease diagnostics, and the effective prevention of human-to-human transmission of fatal viruses. Here, we have introduced a unique respiration monitoring and touchless sensing system based on a CsPb2Br5/BaTiO3 humidity-sensing layer operated by water-induced interfacial polarization and prepared using a facile aerosol deposition process. Based on the relationship between sensing ability and layer thickness, the sensing device with a 1.0 µm thick layer was found to exhibit optimal sensing performance, a result of its ideal microstructure. This sensor also exhibits the highest electrical signal variation at 0.5 kHz due to a substantial polarizability difference between high and low humidity. As a result, the CsPb2Br5/BaTiO3 sensing device shows the best signal variation of all types of breath-monitoring devices reported to date when used to monitor sudden changes in respiratory rates in diverse situations. Furthermore, the sensor can effectively detect sweat evaporation when placed 1 cm from the skin, including subtle changes in capacitance caused by finger area and motion, skin moisture, and contact time. This ultrasensitive sensor, with its fast response, provides a potential new sensing platform for the long-term daily monitoring of respiration and sweat evaporation.

Pontin arginine methylation by CARM1 is crucial for epigenetic regulation of autophagy.

Autophagy is a catabolic process through which cytoplasmic components are degraded and recycled in response to various stresses including starvation. Recently, transcriptional and epigenetic regulations of autophagy have emerged as essential mechanisms for maintaining homeostasis. Here, we identify that coactivator-associated arginine methyltransferase 1 (CARM1) methylates Pontin chromatin-remodeling factor under glucose starvation, and methylated Pontin binds Forkhead Box O 3a (FOXO3a). Genome-wide analyses and biochemical studies reveal that methylated Pontin functions as a platform for recruiting Tip60 histone acetyltransferase with increased H4 acetylation and subsequent activation of autophagy genes regulated by FOXO3a. Surprisingly, CARM1-Pontin-FOXO3a signaling axis can work in the distal regions and activate autophagy genes through enhancer activation. Together, our findings provide a signaling axis of CARM1-Pontin-FOXO3a and further expand the role of CARM1 in nuclear regulation of autophagy.

Parallel Single-Cell RNA-Seq and Genetic Recording Reveals Lineage Decisions in Developing Embryoid Bodies.

Early developmental specification can be modeled by differentiating embryonic stem cells (ESCs) to embryoid bodies (EBs), a heterogeneous mixture of three germ layers. Here, we combine single-cell transcriptomics and genetic recording to characterize EB differentiation. We map transcriptional states along a time course and model cell fate trajectories and branchpoints as cells progress to distinct germ layers. To validate this inferential model, we propose an innovative inducible genetic recording technique that leverages recombination to generate cell-specific, timestamp barcodes in a narrow temporal window. We validate trajectory architecture and key branchpoints, including early specification of a primordial germ cell (PGC)-like lineage from preimplantation epiblast-like cells. We further identify a temporally defined role of DNA methylation in this PGC-epiblast decision. Our study provides a high-resolution lineage map for an organoid model of embryogenesis, insights into epigenetic determinants of fate specification, and a strategy for lineage mapping of rapid differentiation processes.

RORα Regulates Cholesterol Metabolism of CD8+ T Cells for Anticancer Immunity.

Retinoic acid-related orphan receptor α (RORα) functions as a transcription factor for various biological processes, including circadian rhythm, inflammation, cancer, and lipid metabolism. Here, we demonstrate that RORα is crucial for maintaining cholesterol homeostasis in CD8+ T cells by attenuating NF-kB transcriptional activity. Cholesterol sulfate, the established natural agonist of RORα, exhibits cellular cytotoxicity on, and increased effector responses in, CD8+ T cells. Transcript analysis reveals that the suppression of RORα leads to the upregulation of NF-kB target genes in T cells. Chromatin immunoprecipitation analysis was used to determine the corecruitment of RORα and histone deacetylase (HDAC) on NF-kB target promoters and the subsequent dismissal of coactivators for transcriptional repression. We demonstrate that RORα/HDAC-mediated attenuation of NF-kB signaling controls the balance of cholesterol metabolism in CD8+ T cells, and that therapeutic strategies targeting this epigenetic regulation could be beneficial to the treatment of solid tumors including colon cancers.

Inducible histone K-to-M mutations are dynamic tools to probe the physiological role of site-specific histone methylation in vitro and in vivo.

Development and differentiation are associated with profound changes to histone modifications, yet their in vivo function remains incompletely understood. Here, we generated mouse models expressing inducible histone H3 lysine-to-methionine (K-to-M) mutants, which globally inhibit methylation at specific sites. Mice expressing H3K36M developed severe anaemia with arrested erythropoiesis, a marked haematopoietic stem cell defect, and rapid lethality. By contrast, mice expressing H3K9M survived up to a year and showed expansion of multipotent progenitors, aberrant lymphopoiesis and thrombocytosis. Additionally, some H3K9M mice succumbed to aggressive T cell leukaemia/lymphoma, while H3K36M mice exhibited differentiation defects in testis and intestine. Mechanistically, induction of either mutant reduced corresponding histone trimethylation patterns genome-wide and altered chromatin accessibility as well as gene expression landscapes. Strikingly, discontinuation of transgene expression largely restored differentiation programmes. Our work shows that individual chromatin modifications are required at several specific stages of differentiation and introduces powerful tools to interrogate their roles in vivo.

RORα is crucial for attenuated inflammatory response to maintain intestinal homeostasis.

Retinoic acid-related orphan receptor α (RORα) functions as a transcription factor for various biological processes, including circadian rhythm, cancer, and metabolism. Here, we generate intestinal epithelial cell (IEC)-specific RORα-deficient (RORαΔIEC) mice and find that RORα is crucial for maintaining intestinal homeostasis by attenuating nuclear factor κB (NF-κB) transcriptional activity. RORαΔIEC mice exhibit excessive intestinal inflammation and highly activated inflammatory responses in the dextran sulfate sodium (DSS) mouse colitis model. Transcriptome analysis reveals that deletion of RORα leads to up-regulation of NF-κB target genes in IECs. Chromatin immunoprecipitation analysis reveals corecruitment of RORα and histone deacetylase 3 (HDAC3) on NF-κB target promoters and subsequent dismissal of CREB binding protein (CBP) and bromodomain-containing protein 4 (BRD4) for transcriptional repression. Together, we demonstrate that RORα/HDAC3-mediated attenuation of NF-κB signaling controls the balance of inflammatory responses, and therapeutic strategies targeting this epigenetic regulation could be beneficial to the treatment of chronic inflammatory diseases, including inflammatory bowel disease (IBD).

Mitosis-specific phosphorylation of Mis18α by Aurora B kinase enhances kinetochore recruitment of polo-like kinase 1.

Mis18α, a component of Mis18 complex comprising of Mis18α, Mis18ß, and M18BP1, is known to localize at the centromere from late telophase to early G1 phase and plays a priming role in CENP-A deposition. Although its role in CENP-A deposition is well established, the other function of Mis18α remains unknown. Here, we elucidate a new function of Mis18α that is critical for the proper progression of cell cycle independent of its role in CENP-A deposition. We find that Aurora B kinase phosphorylates Mis18α during mitosis not affecting neither centromere localization of Mis18 complex nor centromere loading of CENP-A. However, the replacement of endogenous Mis18α by phosphorylation-defective mutant causes mitotic defects including micronuclei formation, chromosome misalignment, and chromosomal bridges. Together, our data demonstrate that Aurora B kinase-mediated mitotic phosphorylation of Mis18α is a crucial event for faithful cell cycle progression through the enhanced recruitment of polo-like kinase 1 to the kinetochore.