A Cortico-Cortical Pathway Targets Inhibitory Interneurons and Modulates Paw Movement during Locomotion in Mice.

The primary somatosensory cortex (S1) is important for the control of movement as it encodes sensory input from the body periphery and external environment during ongoing movement. Mouse S1 consists of several distinct sensorimotor subnetworks that receive topographically organized corticocortical inputs from distant sensorimotor areas, including the secondary somatosensory cortex (S2) and primary motor cortex (M1). The role of the vibrissal S1 area and associated cortical connections during active sensing is well documented, but whether (and if so, how) non-whisker S1 areas are involved in movement control remains relatively unexplored. Here, we demonstrate that unilateral silencing of the non-whisker S1 area in both male and female mice disrupts hind paw movement during locomotion on a rotarod and a runway. S2 and M1 provide major long-range inputs to this S1 area. Silencing S2→non-whisker S1 projections alters the hind paw orientation during locomotion, whereas manipulation of the M1 projection has little effect. Using patch-clamp recordings in brain slices from male and female mice, we show that S2 projection preferentially innervates inhibitory interneuron subtypes. We conclude that interneuron-mediated S2-S1 corticocortical interactions are critical for efficient locomotion.SIGNIFICANCE STATEMENT Somatosensory cortex participates in controlling rhythmic movements, such as whisking and walking, but the neural circuitry underlying movement control by somatosensory cortex remains relatively unexplored. We uncover a corticocortical circuit in primary somatosensory cortex that regulates paw orientation during locomotion in mice. We identify neuronal elements that comprise these cortical pathways using pharmacology, behavioral assays, and circuit-mapping methods.

Genome-wide characterization of circRNA expression profile in overexpression of RIP2 chicken macrophages associated with avian pathogenic E.coli infection.

Avian pathogenic E. coli (APEC) can cause localized and systemic diseases in poultry, threatening human health via meat or egg contamination and resulting in considerable economic losses to the poultry industry globally. Increasing evidence shows circRNAs were widely involved in various biological processes. However, the role of circRNAs in the host response against APEC infection, especially correlated with the regulation of RIP2, remains unclear. Herein, the RNAseq technology was used to identify the circRNA expression profiles in the overexpression of RIP2 macrophages with or without APEC infection. A total of 256 and 287 differentially expressed (DE) circRNAs were identified in the overexpression of RIP2 group (oeRIP2) vs. the wild-type group (WT) and oeRIP2 + APEC vs. APEC, respectively, whose parental genes were involved in MAPK signalling pathway, Wnt signalling pathway, focal adhesion, tight junction, and VEGF signalling pathways. Specifically, the key circRNAs, such as 5:814443-825127, 10:18922360-18928461, 2:8746306-8750639, and 2:124177751-124184063 might play a critical role in APEC infection and the regulation of RIP2. As a whole, these findings will facilitate understanding the molecular mechanism underlying circRNAs, especially related to the regulation of the RIP2 gene. Meanwhile, the study may offer new ideas to improve host immune and inflammatory response against APEC infection.

Identification and characterization of microRNAs, especially gga-miR-181b-5p, in chicken macrophages associated with avian pathogenic E. coli infection.

Avian pathogenic E. coli (APEC) is a common pathogen in the poultry industry, which can cause substantial economic losses. Recently, emerging evidence showed that miRNAs were involved in various viral and bacterial infections. To elucidate the role of miRNAs in chicken macrophages in response to APEC infection, we attempted to investigate the miRNAs expression pattern upon APEC infection via miRNA-seq, and to identify the molecular mechanism of the important miRNAs by using RT-qPCR, western blotting, dual-luciferase reporter assay, and CCK-8. The results showed that a total of 80 differentially expressed (DE) miRNAs were identified in comparison of APEC vs. wild-type group, which corresponded to 724 target genes. Moreover, the target genes of the identified DE miRNAs were mainly significantly enriched in the MAPK signalling pathway, autophagy-bird, mTOR signalling pathway, ErbB signalling pathway, Wnt signalling pathway, and TGF-beta signalling pathway. Remarkably, gga-miR-181b-5p is able to participate in host immune and inflammatory responses against APEC infection via targeting of TGFBR1 to modulate the activation of TGF-beta signalling pathway. Collectively, this study provides a perspective of miRNA expression patterns in chicken macrophages upon APEC infection. These findings provide insight into miRNAs against APEC infection, and gga-miR-181b-5p might be a potential target for treating APEC infection.

LRRC8 family proteins within lysosomes regulate cellular osmoregulation and enhance cell survival to multiple physiological stresses.

LRRC8 family proteins on the plasma membrane play a critical role in cellular osmoregulation by forming volume-regulated anion channels (VRACs) necessary to prevent necrotic cell death. We demonstrate that intracellular LRRC8 proteins acting within lysosomes also play an essential role in cellular osmoregulation. LRRC8 proteins on lysosome membranes generate large lysosomal volume-regulated anion channel (Lyso-VRAC) currents in response to low cytoplasmic ionic strength conditions. When a double-leucine L706L707 motif at the C terminus of LRRC8A was mutated to alanines, normal plasma membrane VRAC currents were still observed, but Lyso-VRAC currents were absent. We used this targeting mutant, as well as pharmacological tools, to demonstrate that Lyso-VRAC currents are necessary for the formation of large lysosome-derived vacuoles, which store and then expel excess water to maintain cytosolic water homeostasis. Thus, Lyso-VRACs allow lysosomes of mammalian cells to act as the cell`s "bladder." When Lyso-VRAC current was selectively eliminated, the extent of necrotic cell death to sustained stress was greatly increased, not only in response to hypoosmotic stress, but also to hypoxic and hypothermic stresses. Thus Lyso-VRACs play an essential role in enabling cells to mount successful homeostatic responses to multiple stressors.

Intrageneric Relationship of Datnioides (Lobotiformes) Inferred from the Complete Nuclear Ribosomal DNA Operon.

Tiger fish (genus Datnioides) are critical ornamental and economic fish and are valuable freshwater fish worldwide, belonging to the order Lobotiformes. Currently, there are five extant species (Datnioides campbelli, D. microlepis, D. polota, D. pulcher, and D. undecimradiatus) of Datnioides in the world, usually inhabiting in south and southeast Asia. Due to the decline of wild population sizes of tiger fish and the lack of molecular research on them, in the present study, we sequenced, assembled, and characterized the complete nuclear ribosomal DNA (nrDNA) operon of all five extant tiger fish species, in order to elucidate the phylogenetic relationship among the genus Datnioides. The nrDNA sequences of five tiger fish species were 8548-9182 bp in length, encompassing complete 18S rDNA, ITS1, 5.8S rDNA, ITS2, 28S rDNA, and IGS regions. Numerous repetitive sequences were detected, substantially influencing the sequence length of different regions in each species. We employed maximum-likelihood (ML) method and Bayesian inference (BI) method to construct phylogenetic trees for Datnioides. Phylogenetic analyses indicated that each region in nrDNA operon is not sufficiently phylogenetically informative to delineate the species in Datnioides; nevertheless, the whole operon is able to delineate five tiger fish species much better, three of five species were successfully partitioned. Particularly, regardless of employed markers, it was strongly supported that D. campbelli was considerably partitioned from the other four species, possibly due to the geographical separation. In spite of the fact that discrimination of Datnioides species requires further investigation, our study provides reference genome resources for the Lobotiformes, as well as insights into the phylogenetic position of Lobotiformes and further biological conservation.

Gene expression profiling of RIP2-knockdown in HD11 macrophages - elucidation of potential pathways (gene network) when challenged with avian pathogenic E.coli (APEC).

BACKGROUND: Receptor interacting serine/threonine kinase 2 (RIP2), ubiquitous in many tissue/cell types, is the key regulator of immune and inflammatory responses for many diseases, including avian pathogenic E. coli (APEC), which causes a wide variety of localized or systemic infections. However, the molecular mechanisms by which RIP2 drives its transcriptional program to affect immune and inflammatory response upon APEC infection remains poorly understood. RESULTS: In this study, RNA-seq and bioinformatics analyses were used to detect gene expression and new direct/indirect RIP2 targets in the treatments of wild type HD11 cells (WT), RIP2 knockdown cells (shRIP2), APEC stimulation cells (APEC), and RIP2 knockdown cells combined with APEC infection (shRIP2 + APEC). The results revealed that a total of 4691 and 2605 differentially expressed genes (DEGs) were screened in shRIP2 + APEC vs. APEC and shRIP2 vs. WT, respectively. Functional annotation analysis showed that apoptosis, MAPK, p53, Toll-like receptor, and Nod-like receptor signaling pathways were involved in APEC-induced RIP2 knockdown HD11 cells. By analyzing the enriched pathway and gene networks, we identified that several DEGs, including HSP90AB1, BID, and CASP9 were targeted by RIP2 upon APEC infection. CONCLUSION: As a whole, this study can not only provide data support for constructing gene networks of RIP2 knockdown with APEC challenge but also provide new ideas for improving the immune and inflammatory response.

Exotic fishes that are phylogenetically close but functionally distant to native fishes are more likely to establish.

Since Darwin's time, degree of ecological similarity between exotic and native species has been assumed to affect the establishment success or failure of exotic species. However, a direct test of the effect of exotic-native similarity on establishment of exotics is scarce because of the difficulty in recognizing failures of species to establish in the field. Here, using a database on the establishment success and failure of exotic fish species introduced into 673 freshwater lakes, we evaluate the effect of similarity on the establishment of exotic fishes by combining phylogenetic and functional information. We illustrate that, relative to other biotic and abiotic factors, exotic-native phylogenetic and functional similarities were the most important correlates of exotic fish establishment. While phylogenetic similarity between exotic and resident fish species promoted successful establishment, functional similarity led to failure of exotics to become established. Those exotic species phylogenetically close to, but functionally distant from, native fishes were most likely to establish successfully. Our findings provide a perspective to reconcile Darwin's naturalization conundrum and suggest that, while phylogenetic relatedness allows exotic fish species to pre-adapt better to novel environments, they need to possess distinct functional traits to reduce competition with resident native fish species.

Impact of Chronic HIV Infection on SARS-CoV-2 Infection, COVID-19 Disease and Vaccines.

PURPOSE OF REVIEW: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has developed into a global pandemic that affect the health of hundreds of millions worldwide. In particular, SARS-CoV-2 infection in people with chronic human immune deficiency virus (HIV) infection is of concern, due to their already immunocompromised status. Yet, whether and how the immunological changes brought about by HIV will affect the immune responses against SARS-CoV-2 acute infection and impact the effectiveness of vaccines remain unclear. We discuss the intersection of COVID-19 in HIV-infected individuals. RECENT FINDINGS: People living with HIV (PLWH) may be at increased risk of severe SARS-CoV-2 mediated disease complication due to functional impairment of the immune system and persistent inflammation, which can be ameliorated by antiretroviral therapy. Importantly, limited data suggest that current approved vaccines may be safe and efficacious in PLWH. To address remaining questions and supplement limited experimental evidence, more studies examining the interplay between HIV and SARS-CoV-2 through their impact on the host immune system are required.

Analysis of miRNA Expression Profiling of RIP2 Knockdown in Chicken HD11 Cells When Infected with Avian Pathogenic E. coli (APEC).

Colibacillosis is an acute and chronic avian disease caused by avian pathogenic E. coli (APEC). Previous studies have demonstrated that RIP2 plays a significant role in APEC infection. Moreover, increasing evidence indicates that microRNAs (miRNAs) are involved in host-pathogen interactions and the immune response. However, the role of miRNAs in the host against APEC infection remains unclear. Herein, we attempted to reveal new miRNAs potentially involved in the regulation of the immune and inflammatory response against APEC infection, with a particular focus on those possibly correlated with RIP2 expression, via miRNA-seq, RT-qPCR, Western blotting, dual-luciferase reporter assay, and CCK-8. The results showed that a total of 93 and 148 differentially expressed (DE) miRNAs were identified in the knockdown of RIP2 cells following APEC infection (shRIP2+APEC) vs. knockdown of RIP2 cells (shRIP2) and shRIP2 vs. wild-type cells (WT), respectively. Among those identified DE miRNAs, the biological function of gga-miR-455-5p was investigated. It was found that gga-miR-455-5p regulated by RIP2 was involved in the immune and inflammatory response against APEC infection via targeting of IRF2 to modulate the expression of type I interferons. Additionally, RIP2 could directly regulate the production of the type I interferons. Altogether, these findings highlighted the crucial role of miRNAs, especially gga-miR-455-5p, in host defense against APEC infection.

Tissue-resident memory CD8+ T cells in cancer immunology and immunotherapy.

Memory T cells can be generated and remain long-term in different tissues following infection or immunization. Tissue-resident memory T (TRM) cells are a unique group of memory T cells that form and persist mainly in peripheral non-lymphoid organs. Unlike effector or central memory T (TEM or TCM) cells, TRM cells do not circulate to the blood but can provide a rapid and robust local response to re-infection. Recently, a large body of clinical studies has shown that CD103+ CD8+ TRM-like cells also exist intratumorally and strongly correlate with favorable prognosis in cancer patients. Cancer vaccine-induced CD103+ CD8+ TRM cells have been reported to suppress tumor growth in mouse models. This suggests that CD8+ TRM-like cells play a crucial role in cancer immunosurveillance and immunotherapy. In this review, we focus on the features and cytotoxic mechanisms of CD8+ TRM-like cells in multiple solid tumors and discuss their potential implications for cancer immunotherapy. We believe a better understanding of the generation, function, and longevity of CD8+ TRM-like cells in the tumor microenvironment will provide new insights for cancer immunotherapies.

LRRC14 attenuates Toll-like receptor-mediated NF-κB signaling through disruption of IKK complex.

Activation of NF-κB signaling plays pivotal roles in innate immune responses against pathogens. It requires strict control to avert inflammatory diseases. However, the mechanisms underlying this tight regulation are not completely understood. Here, we identified LRRC14, a novel member of LRR (leucine-rich repeat) protein family, as a negative regulator in TLR signaling. Expression of LRRC14 resulted in decreased activation of NF-κB, whereas knockdown of LRRC14 enhanced NF-κB activation as well as the production of inflammatory cytokines. Mechanistically, LRRC14 bound to HLH domain of IKKß to block its interaction with NEMO and thereby inhibiting the phosphorylation of IKKß and NF-κB activation. In addition, our data showed that TLR signaling led to lower expression of LRRC14. Together, LRRC14 may function as a checkpoint to prevent overzealous inflammation.

CoSFISH: a comprehensive reference database of COI and 18S rRNA barcodes for fish.

Fish, being a crucial component of aquatic ecosystems, holds significant importance from both economic and ecological perspectives. However, the identification of fish at the species level remains challenging, and there is a lack of a taxonomically complete and comprehensive reference sequence database for fish. Therefore, we developed CoSFISH, an online fish database. Currently, the database contains 21 535 cytochrome oxidase I sequences and 1074 18S rRNA sequences of 21 589 species, belonging to 8 classes and 90 orders. We additionally incorporate online analysis tools to aid users in comparing, aligning and analyzing sequences, as well as designing primers. Users can upload their own data for analysis, in addition to using the data stored in the database directly. CoSFISH offers an extensive fish database and incorporates online analysis tools, making it a valuable resource for the study of fish diversity, phylogenetics and biological evolution. Database URL:  http://210.22.121.250:8888/CoSFISH/home/indexPage.

Gap-free chromosome-level genomes of male and female spotted longbarbel catfish Hemibagrus guttatus.

Hemibagrus guttatus, also named as spotted longbarbel catfish, is an economical fish in China. However, their gender cannot be easily distinguished from their appearance, which largely impedes their artificial breeding. Therefore, we provided two gap-free chromosome-level genomes of male and female spotted longbarbel catfish by combining wtdbg2, LR_Gapcloser and TGS-GapCloser assembly approaches with Hi-C data and accurate Pacbio HiFi long-reads. We assembled 30 chromosomes without any gap. Their genome sizes are approximately 749.1 Mb and 747.8 Mb of male and female individuals. The completeness results of BUSCO evaluation show about 94.2% and 95.0%, representing a high-level of completeness of both genomes. We also obtained 35,277 and 34,571 protein-coding gene sets from male and female individuals. Both available gap-free chromosome-level genomes of H. guttatus will provide excellent references for resequencing of male and female individuals to identify accurate markers for distinguishing gender of this fish.

Long noncoding RNAs expression profile of RIP2 knockdown in chicken HD11 macrophages associated with avian pathogenic E. coli (APEC) infection.

Avian pathogenic E. coli (APEC) has been detected to cause many acute and chronic diseases, resulting in huge economic losses to the poultry industry. Previous experiments have identified the effect of receptor interacting serine/threonine kinase 2 (RIP2) gene in APEC infection. Moreover, increasing evidence indicates that long noncoding RNAs (lncRNAs) play important roles in the anti-bacteria responses. However, little is known about the functions of lncRNAs, especially related to RIP2, in response to APEC. Therefore, we tried to reveal lncRNAs potentially involved in the immune and inflammatory response against APEC infection, with a particular focus on those possibly correlated with RIP2. A total of 1856 and 1373 differentially expressed (DE) lncRNAs were identified in knockdown of RIP2 cells following APEC infection (shRIP2+APEC) vs. APEC and shRIP2 vs. wild type cells (WT), respectively, which were mainly enriched in lysosome, phagosome, NOD-like receptor signaling pathway, TGF-beta signaling pathway. Significantly, TCONS_00009695 regulated by RIP2 could directly alter the expression of target BIRC3 to modulate cytokines and to participate in immune and inflammatory response against APEC infection. Our findings aid to a better understanding of host responses to APEC infection and provide new directions for understanding the potential association between lncRNAs and APEC pathogenesis.

Nucleolar Protein 56 Deficiency in Zebrafish Leads to Developmental Abnormalities and Anemia via p53 and JAK2-STAT3 Signaling.

Ribosomes are the vital molecular machine for protein translation in a cell. Defects in several nucleolar proteins have been observed in human ribosomopathies. In zebrafish, a deficiency in these ribosomal proteins often results in an anemic phenotype. It remains to be determined whether any other ribosome proteins are involved in regulating erythropoiesis. Here, we generated a nucleolar protein 56 (nop56)-/- zebrafish model and investigated its function. A nop56 deficiency induced severe morphological abnormalities and anemia. WISH analysis showed that the specification of the erythroid lineage in definitive hematopoiesis and the maturation of erythroid cells were impaired in the nop56 mutants. Additionally, transcriptome analysis revealed that the p53 signaling pathway was abnormally activated, and the injection of a p53 morpholino partially rescued the malformation, but not the anemia. Moreover, qPCR analysis showed that the JAK2-STAT3 signaling pathway was activated in the mutants, and the inhibition of JAK2 partially rescued the anemic phenotype. This study suggests that nop56 is a potential target for investigation in erythropoietic disorders, particularly those that may be associated with JAK-STAT activation.

Mechanical Properties and Deformation Mechanisms of Nanocrystalline U-10Mo Alloys by Molecular Dynamics Simulation.

U-Mo alloys were considered to be the most promising candidates for high-density nuclear fuel. The uniaxial tensile behavior of nanocrystalline U-10Mo alloys with average grain sizes of 8-23 nm was systematically studied by molecular dynamics (MD) simulation, mainly focusing on the influence of average grain size on the mechanical properties and deformation mechanisms. The results show that Young's modulus, yield strength and ultimate tensile strength follow as average grain size increases. During the deformation process, localized phase transitions were observed in samples. Grain boundary sliding and grain rotation, as well as twinning, dominated the deformation in the smaller and larger grain sizes samples, respectively. Increased grain size led to greater localized shear deformation, resulting in greater stress drop. Additionally, we elucidated the effects of temperature and strain rate on tensile behavior and found that lower temperatures and higher strain rates not only facilitated the twinning tendency but also favored the occurrence of phase transitions in samples. Results from this research could provide guidance for the design and optimization of U-10Mo alloys materials.

Characterization and functional analysis of pax3 in body color transition of polychromatic Midas cichlids (Amphilophus citrinellus).

As the representative genetic and economic trait of ornamental fish, skin color has a strong impact on speciation and adaptation. However, the genetic basis of skin color pigmentation, differentiation and change is still not understood. The Midas cichlid fish with three typical body color transition stages of "black-gray­gold" is an ideal model system for investigating the formation and change of fish body color. In this study, to investigate the regulatory role of the pair box 3 (pax3) gene in the early body color fading process of Midas cichlids, the complete cDNA sequence (3513 bp) of pax3 was successfully isolated from Midas cichlids (Amphilophus Citrinellus), and found to encode polypeptides of 491 amino acids. Expression patterns of the pax3 gene in tissues of Midas cichlids during different periods, including embryonic development and body color fading stages were detected by quantitative real-time PCR. The qRT-PCR analysis showed that pax3 was expressed in all tissues of adult fish, with a higher expression level in muscle and skin. The highest expression level in muscle tissue was significantly higher than that in other tissues (P < 0.05). During embryonic development, the expression tendency of pax3 was first increased and then decreased. In the three typical stages of early skin color fading from black to gold, pax3 expression in skin, caudal fin and scales all showed a downward trend. The expression level in the black stage was significantly higher than that in other stages (P < 0.05). Positive signal of pax3 protein was detected in the three typical skin color conversion stages, and the highest positive signal intensity was detected in the black stage, which was consistent with qRT-PCR results. After pax3 RNA interference, pax3 and the downstream genes mitf and tyr all decreased, while dct mRNA expression increased in the skin of fish. Western blotting also showed a decrease in pax3 protein concentration. Those results suggest that pax3 plays an important role in skin color formation, distribution and change in Midas cichlids through the melanogenesis pathway.

SpG and SpRY variants expand the CRISPR toolbox for genome editing in zebrafish.

Precise genetic modifications in model organisms are essential for biomedical research. The recent development of PAM-less base editors makes it possible to assess the functional impact and pathogenicity of nucleotide mutations in animals. Here we first optimize SpG and SpRY systems in zebrafish by purifying protein combined with synthetically modified gRNA. SpG shows high editing efficiency at NGN PAM sites, whereas SpRY efficiently edit PAM-less sites in the zebrafish genome. Then, we generate the SpRY-mediated cytosine base editor SpRY-CBE4max and SpRY-mediated adenine base editor zSpRY-ABE8e. Both target relaxed PAM with up to 96% editing efficiency and high product purity. With these tools, some previously inaccessible disease-relevant genetic variants are generated in zebrafish, supporting the utility of high-resolution targeting across genome-editing applications. Our study significantly improves CRISPR-Cas targeting in the genomic landscape of zebrafish, promoting the application of this model organism in revealing gene function, physiological mechanisms, and disease pathogenesis.

Upconversion fluorescence-based paper disc for multiplex point-of-care testing in water quality monitoring.

Water quality monitoring is of great significance for human health, which involves a large number of distinct targets detection, such as pathogens, heavy metal ions, and toxins. Traditional detection methods usually carried out in well-equipped central labs are subjected to time, labor, and expense consumption, thus not suitable for implementing water quality monitoring at point of care, where the detection implemented on a multiplexed, miniaturized, and simplified device with rapid and sensitive answer out is desired. To this end, we developed a paper disc relying on upconversion fluorescence signal and aptamer recognition probes for multiplex detection of water contaminants with high sensitivity and specificity. Finally, several different typical kinds of water contaminants have been successfully detected on our paper disc with limit of detection of 115 cfu/mL for Salmonella, 3 ng/mL for Ochratoxin A and Microcystin-LR, 20 nM for Hg2+ and 4 nM for Pb2+, respectively. Additionally, we designed a compact smartphone-based reading device to enable water quality monitoring at point-of-care. Furthermore, this paper disc-based detection platform can be extended to apply to multiplex detection for disease diagnosis and food safety monitoring.