S6B-2 Matrine Inhibits Itching by Lowering the Activity of Calcium Channel

Sophorae Flavescentis Radix(SFR)is a medicinal herb with many functions that are involved in anti-inflammation,antinociception,and anticancer.SFR is also used to treat a variety of itching diseases.Matrine(MT)is one of the main constituents in SFR and also has the effect of relieving itching,but the antipruritic mechanism is still unclear.Here,we investigated the effect of MT on antipruritus.In acute and chronic itch models,MT significantly inhibited the scratching behavior not only in acute itching induced by histamine(His),chloroquine(CQ)and compound 48/80 with a dose-depended manner,but also in the chronic pruritus models of Atopic dermatitis(AD)and Acetone-ether-water(AEW)in mice.Furthermore,MT can be detected in the blood after intraperitoneal injection(i.p.)and subcutaneous injection(s.c.).Finally,electrophysiological and calcium image results show that MT inhibits the excitatory synaptic transmission from dorsal root ganglion(DRG)to the dorsal horn of the spinal cord by suppressing presynaptic N-type calcium channels.Taken together,we believe that MT is a novel drug candidate in treating pruritus diseases,especially for histamine-independent and chronic pruritus,which might be attributed to inhibition of presynaptic N-type calcium channels.

Deficiency of transmembrane AMPA receptor regulatory protein γ-8 leads to attention-deficit hyperactivity disorder-like behavior in mice

Attention-deficit hyperactivity disorder(ADHD) is a neurodevelopmental disorder prevalent in schoolage children. At present, however, its etiologies and risk factors are unknown. Transmembrane α-amino-3-hydroxy-5-methyl-4-isoxazolepropionicacid(AMPA) receptor regulatory protein γ-8(TARP γ-8,also known as calcium voltage-gated channel auxiliary subunit gamma 8(CACNG8)) is an auxiliary AMPA receptor(AMPAR) subunit. Here, we report an association between TARP γ-8 and ADHD,whereby adolescent TARP γ-8 knockout(KO) mice exhibitedADHD-likebehaviors,including hyperactivity, impulsivity, anxiety, impaired cognition,and memory deficits. Human single-nucleotide polymorphism(SNP) analysis also revealed strong associations between intronic alleles in CACNG8genes and ADHD susceptibility. In addition,synaptosomal proteomic analysis revealed dysfunction of the AMPA glutamate receptor complex in the hippocampi of TARP γ-8 KO mice.Proteomic analysis also revealed dysregulation of dopaminergic and glutamatergic transmissions in the prefrontal cortices of TARP γ-8 KO mice.Methylphenidate(MPH), which is commonly used to treat ADHD, significantly rescued the major behavioral deficits and abnormal synaptosomal proteins in TARP γ-8 KO mice. Notably, MPH significantly reversed the up-regulation of Grik2 and Slc6a3 in the prefrontal cortex. MPH also significantly improved synaptic AMPAR complex function by up-regulating other AMPAR auxiliary proteins in hippocampal synaptosomes. Taken together, our results suggest that TARP γ-8 is involved in the development of ADHD in humans.This study provides a useful alternative animal model with ADHD-like phenotypes related to TARP γ-8deficiency, which has great potential for the development of new therapies.

肝胆肿瘤中肿瘤特异性CircRNA衍生抗原肽的鉴定

基于肿瘤抗原的免疫治疗的应用受到验证免疫原性肽稀缺性的阻碍。本研究旨在研究环状RNA(circRNA)在肝胆肿瘤类器官中作为肿瘤抗原肽新来源的潜力。使用RNA测序(RNA-seq)和基于算法的评分工具,预测3950个翻译的肿瘤特异性环状RNA在27个类器官中产生18971个抗原肽。从抗原格局来看,11个氨基酸长度(mer)肽和人白细胞抗原(HLA)-A结合肽具有最高的免疫原性相关评分。在分析的3/5类器官中,有13个预测抗原肽通过质谱(MS)免疫肽组学被直接确认为HLA-A、HLA-B和HLA-C(HLA-ABC)结合肽。在流式细胞术和酶联免疫吸附试验(ELISA)中,由HLA-ABC分子呈递的circRNA衍生的肿瘤特异性肽刺激CD8(CD8)T细胞,显示CD107a干扰素γ(IFNγ)共表达和IFNγ分泌增加。免疫原性环状RNA衍生肽诱导的靶向类器官的细胞毒性T细胞活性在杀伤实验中得到验证。值得注意的是,来自circTBC1D15的抗原肽YGFNEILKK不仅被认为是类器官的HLA-ABC呈递肽,而且还显著降低了肿瘤类器官的存活率。本研究的发现强调了产生肿瘤抗原的一个关键亚群,这对靶向肿瘤特异性circRNA具有重要意义。

A bacterial artificial chromosome transgenic mouse model for visualization of neurite growth

Class Ⅲ β-tubulin （Tubb3） is a component of the microtubules in neurons and contributes to microtubule dynamics that are required for axon outgrowth and guidance during neuronal development. We here report a novel bacterial artificial chromosome （BAC） transgenic mouse line that expresses Class Ⅲ β-tubulin fused to mCherry, an improved monomeric red fluorescent protein, for the visualization of microtubules during neuronal development. A BAC containing Tubb3 gene was modified by insertion of mCherry complementary DNA downstream of Tubb3 coding sequence via homologous recombination, mCherry fusion protein was expressed in the nervous system and testis of the transgenic animal, and the fluorescent signal was observed in the neurons that located in the olfactory bulb, cerebral cortex, hippocampal formation, cerebellum, as well as the retina. Besides, Tubb3-mCherry fusion protein mainly distributed in neurites and colocalized with endogenous Class Ⅲ β-tubulin The fusion protein labels Purkinje cell dendrites during cerebellar circuit formation. Therefore, this transgenic line might be a novel tool for scientific community to study neuronal development both in vitro and in vivo.

TBC1D1 is an energy-responsive polarization regulator of macrophages via governing ROS production in obesity

Energy status is linked to the production of reactive oxygen species(ROS)in macrophages,which is elevated in obesity.However,it is unclear how ROS production is upregulated in macrophages in response to energy overload for mediating the development of obesity.Here,we show that the Rab-GTPase activating protein(Rab GAP)TBC1D1,a substrate of the energy sensor AMP-activated protein kinase(AMPK),is a critical regulator of macrophage ROS production and consequent adipose inflammation for obesity development.TBC1D1 deletion decreases,whereas an energy overload-mimetic non-phosphorylatable TBC1D1^(S231A)Amutation increases,ROS production and M1-like polarization in macrophages.Mechanistically,TBC1D1 and its downstream target Rab8a form an energy-responsive complex with NOX2 for ROS generation.Transplantation of TBC1D1^(S231A)bone marrow aggravates diet-induced obesity whereas treatment with an ultra-stable Tt SOD for removal of ROS selectively in macrophages alleviates both TBC1D1~(S231A)mutation-and diet-induced obesity.Our findings therefore have implications for drug discovery to combat obesity.

Depletion of Gsdma1/2/3 alleviates PMA-induced epidermal hyperplasia by inhibiting the EGFR–Stat3/Akt pathway

Homeostasis of the skin barrier is essential for maintaining normal skin function.Gasdermin A(GSDMA)is highly expressed in the skin and associated with many skin diseases,such as melanoma and psoriasis.In mice,GSDMA is encoded by three gene homologues,namely Gsdma1,Gsdma2,and Gsdma3.Although Gsdma3 gain-of-function mutations cause hair loss and skin inflammation,Gsdma3-deficient mice do not show any visible phenotypes in skin and hair structures.To explore the physiological function of GSDMA,we generated conventional Gsdma1/2/3 knockout(KO)mice.These mice showed significantly alleviated epidermal hyperplasia and inflammation induced by phorbol 12-myristate 13-acetate(PMA).Furthermore,the alleviation of epidermal hyperplasia depended on the expression of Gsdma1/2/3 specifically in keratinocytes.Mechanistically,Gsdma1/2/3 depletion downregulated epidermal growth factor receptor(EGFR)ligands,leading to the decreased EGFR–Stat3/Akt signalling.These results demonstrate that depletion of Gsdma1/2/3 alleviates PMA-induced epidermal hyperplasia partially by inhibiting the EGFR–Stat3/Akt pathway.

TMEM63B regulates postnatal development of cochlear sensory epithelia via thyroid hormone signaling

Sound transmission occurs in the cochlea,a complex and ingenious subdivision in the inner ear.The structure of the cochlea develops structurally and functionally by the time before two postnatal weeks(the time of hearing onset)in mice(Geal-Dor et al.,1993).Greater epithelial ridge(GER,also known as Kolliker's organ)is a transient cochlear structure containing a group of columnar epithelial supporting cells surrounding the inner hair cells(IHCs).

Model animals and their applications

Looking back a decade ago when I decided to use ＂model animal＂ to name our newly established institute, my outrageous boldness can only be described by the old Chinese slang, ＂newborn calf could never sense the danger of tiger＂. Happily, my courageous belief, along with the hard work of my colleagues at Model Animal Research Center of Nanjing University, paid off eventually. We have witnessed the great progress in this research field in China in the past 10 years. This issue of Science China Life Sciences samplings some of these accomplishments.

Towards a better understanding of mouse and human diseases—International Mouse Phenotyping Consortium

With the completion of the human genome sequence, the last several decades have seen the pursuit of gene function in vivo. The mouse is generally considered the best model animal for examining gene function because of its high productivity, pure genetic background, and degree of similarity to the human genome. With the development of several gene manipulation methodologies, mice are universally used to study genetics and human diseases. There are more than 20,000 protein-coding genes in the mouse genome. The International Knockout Mouse Consortium （IKMC） project was started to generate a knockout of every protein-coding gene in embryonic stem （ES） cells using standard knockout targeting strategies. Following completion of this project, the International Mouse Phenotyping Consortium （IMPC） was established, with the aim of turning these ES cells into live mice, and exploring their potential defective phenotypes using a standard phenotyping pipeline. Globally, about 20 mouse clinics from Europe, the USA, Canada, Australia, and Asia participated in this 10-year project, in which Model Animal Research Center of Nanjing University is the only one from China's Mainland. The combined data from these centers will represent a new data platform providing a gene function encyclopedia, and more importantly, a comprehensive picture of the whole genome.

Corrigendum to“Actin polymerization induces mitochondrial distribution during collective cell migration”[Journal of Genetics and Genomics(2023)50 46-49]

This erratum clarifies information in the Letter to the Editor“Actin polymerization induces mitochondrial distribution during collective cell migration”by Qu et al.(2023).In the section for the list of author names,“Chen Qu,Yating Kan,Hui Zuo,Mengqi Wu,Zhixiang Dong,Xinyi Wang,Qing Zhang,Heng Wang,Dou Wang,Jiong Chen”should be“Chen Qu,Yating Kan,Xinyi Wang,Hui Zuo,Mengqi Wu,Zhixiang Dong,Qing Zhang,Heng Wang,Dou Wang,Jiong Chen”.

Deficiency of Autophagy-Related Gene 5 in Keratinocytes Leads to Aggravation of Epidermal Damage in 2,4-Dinitrochlorobenzene-Induced Allergic Contact Dermatitis

Objective:The interrelationship between apoptosis and autophagy plays an important role in many pathophysiological processes,however,whether their interplay is involved in allergic contact dermatitis(ACD)has not yet been elucidated.So,we conducted this study to determine whether keratinocyte-specific autophagy-related gene 5(ATG5)deficiency can regulate apoptosis to inhibit skin damage in mice with 2,4-dinitrochlorobenzene(DNCB)-induced ACD.Methods:This study involved keratinocyte-specificAtg5 conditional knockout(cKO)mice(Krt14cre/+-Atg5flox/flox)and control mice(Krt14+/+-Atg5flox/flox).We painted DNCB on the right ear of each mouse to induce ACD.Dermatitis scoring and measurements of ear weight and thickness were performed to evaluate inflammation levels.An immunohistochemical assay was performed to analyze immune cell infiltration.Histological study and TUNEL staining were performed to compare the differences in skin lesions betweenAtg5 cKO mice and control mice.Immunofluorescence and western blotting were used to examine the levels of ATG5 and apoptosis-related protein.The results were statistically analyzed byt test.Results:After DNCB stimulation of mice ears,we observed a more severe phenotype inAtg5 cKO mice than in control mice(dermatitis score:7.500±2.588vs.3.250±0.822,P=0.003).Further analysis of ATG5 protein confirmed keratinocyte-specific ablation ofAtg5 in cKO mice and showed that DNCB did not influence ATG5 expression.Immunohistochemistry assay revealed that the infiltrated immune cells were not involved in aggravation of the phenotype of DNCB-stimulatedAtg5 cKO mice.However,the histological study(P=0.024),TUNEL staining(P=0.024),immunofluorescence(P=0.036),and western blotting showed that the increase in keratinocyte death,especially apoptosis,contributed to aggravation of the phenotype of DNCB-stimulatedAtg5 cKO mice.Conclusion:Deficiency ofAtg5 in keratinocytes increases apoptosis,aggravating skin damage in DNCB-induced ACD mice.This has no relationship with the involvement of immune cells.

LonP1 Links Mitochondria–ER Interaction to Regulate Heart Function

Interorganelle contacts and communications are increasingly recognized to play a vital role in cellular function and homeostasis.In particular,the mitochondria–endoplasmic reticulum(ER)membrane contact site(MAM)is known to regulate ion and lipid transfer,as well as signaling and organelle dynamics.However,the regulatory mechanisms of MAM formation and their function are still elusive.Here,we identify mitochondrial Lon protease(LonP1),a highly conserved mitochondrial matrix protease,as a new MAM tethering protein.The removal of LonP1 substantially reduces MAM formation and causes mitochondrial fragmentation.Furthermore,deletion of LonP1 in the cardiomyocytes of mouse heart impairs MAM integrity and mitochondrial fusion and activates the unfolded protein response within the ER(UPR^(ER)).Consequently,cardiac-specific LonP1 deficiency causes aberrant metabolic reprogramming and pathological heart remodeling.These findings demonstrate that LonP1 is a novel MAM-localized protein orchestrating MAM integrity,mitochondrial dynamics,and UPR^(ER),offering exciting new insights into the potential therapeutic strategy for heart failure.

TLR5 expression marks brain boarder associated macrophages and protects neonatal mice from bacterial meningitis

Meningitis is one of the major threats to prenatal and newborn babies,causing more than 300,000 infant deaths worldwide annually.Survivors often suffer from long-term neuron-developmental disorders caused by the infection[1–3].Bacterial meningitis,a prevalent form of this condition,remains a major global public health challenge,despite significant progress in combating specific pathogens[4].In infants,where systemic adaptive immunity is not fully developed,brain macrophage plays a crucial role in defensing pathogen infection in this vital organ[5,6].However,our understanding of the precise markers that reflect the distinct contributions of various macrophage populations in the neonatal brain remains incomplete.

Molecular mechanisms of dietary restriction in aging—insights from Caenorhabditis elegans research

Dietary restriction(DR) is one of the most robust environmental manipulations that not only extend life span but also delay the onset of age-related diseases in almost every species examined. Caenorhabditis elegans plays an important role in aging studies due to its simple life cycle, easy genetic manipulations and highly conserved genome. Recent studies have demonstrated that the beneficial effects of DR are mediated by the highly conserved transcription factors and signaling pathways in C. elegans. Here we review recent progress in the methodology and molecular mechanisms of DR using C. elegans as a model, as well as prospects for future research.

Germ layer formation during Xenopus embryogenesis: the balance between pluripotency and differentiation

The African clawed frog, Xenopus laevis, has long been a model animal for the studies in the fields of animal cloning, developmental biology, biochemistry, cell biology, and physiology. With the aid of Xenopus, major molecular mechanisms that are involved in embryonic development have been understood. Germ layer formation is the first event of embryonic cellular differentiation, which is induced by a few key maternal factors and subsequently by zygotic signals. Meanwhile, another type of signals, the pluripotency factors in ES cells, which maintain the undifferentiated state, are also present during early embryonic cells. In this review, the functions of the pluripotency factors during Xenopus germ layer formation and the regulatory relationship between the signals that promote differentiation and pluripotency factors are discussed.

PTPROt maintains T cell immunity in the microenvironment of hepatocellular carcinoma

Intratumoral T cells play a central role in anti-tumor immunity,and the balance between T effector cells(Teff)and regulatory T cells(Treg)affects the prognosis of cancer patients.However,educated by tumor microenvironment,T cells frequently fail in their responsibility.In this study,we aimed to investigate the role of truncated isoform of protein tyrosine phosphatase receptor-typeO(PTPROt)in T cell-mediated anti-tumor immunity.We recruited 70 hepatocellular carcinoma(HCC)patients and 30 healthy volunteers for clinical investigation,and analyzed cellular tumor immunity by using ptpro^(-/-) C57BL/6 mice and NOD/SCID mice.PTPROt expression was significantly downregulated in human HCC-infiltrating T cells due to the hypoxia microenvironment;PTPROt expression highly correlated with the intratumoral Teff/Treg ratio and clinicopathologic characteristics.Moreover,PTPROt deficiency attenuated T cell-mediated anti-tumor immunity and remarkably promoted mouse HCC growth.Mechanistically,deletion of PTPROt decreased Teff quantity and quality through phosphorylation of lymphocyte-specific tyrosine kinase,but increased Treg differentiation through phosphorylation of signal transducer and activator of transcription 5.In support of the Teff/Treg homeostasis,PTPROt serves as an important tumor suppressor in HCC microenvironment.

In vivo RNAi screen identifies candidate signaling genes required for collective cell migration in Drosophila ovary

Collective migration of loosely or closely associated cell groups is prevalent in animal development, physiological events, and cancer metastasis. However, our understanding of the mechanisms of collective cell migration is incomplete. Drosophila border cells provide a powerful in vivo genetic model to study collective migration and identify essential genes for this process. Using border cell-specific RNAi-silencing in Drosophila, we knocked down 360 conserved signaling transduction genes in adult flies to identify essential pathways and genes for border cell migration. We uncovered a plethora of signaling genes, a large proportion of which had not been reported for border cells, including Rack1 （Receptor of activated C kinase） and brk （brinker）, mad （mother against dpp）, and sax （saxophone）, which encode three components of TGF-β signaling. The RNAi knock down phenotype was validated by clonal analysis of Rack1 mutants. Our data suggest that inhibition of Src activity by Rackl may be important for border cell migration and cluster cohesion maintenance. Lastly, results from our screen not only would shed light on signaling pathways involved in collective migration during embryogenesis and organogenesis in general, but also could help our understanding for the functions of conserved human genes involved in cancer metastasis.

Glucose metabolic abnormality is associated with defective mineral homeostasis in skeletal disorder mouse model

Bone was reported as a crucial organ for regulating glucose homeostasis. In this study, we found that Phex mutant mice(PUG), a model of human X-linked hypophosphatemic rickets(XLH), displayed metabolic abnormality in addition to abnormal phosphate homeostasis, skeletal deformity and growth retardation. Glucose tolerance was elevated with enhanced insulin sensitivity in PUG, though circulating insulin level decreased. Interestingly, bone mineral density defects and glucose metabolic abnormality were both rescued by adding phosphorus- and calcium-enriched supplements in daily diet. Serum insulin level, glucose tolerance and insulin sensitivity showed no differences between PUG and wild-type mice with rescued osteocalcin(OCN) following treatment. Our study suggested that OCN is a potential mediator between mineral homeostasis and glucose metabolism. This investigation brings a new perspective on glucose metabolism regulation through skeleton triggered mineral homeostasis and provides new clues in clinical therapeutics of potential metabolic disorders in XLH patients.

Master microRNA-222 regulates cardiac microRNA maturation and triggers Tetralogy of Fallot

Dear Editor,Tetralogy of Fallot(TOF)is the most common complex congenital heart disease.Besides gene mutations and copy number variants,altered protein function induced by posttranscriptional or translational regulation also contributes to the onset of TOF.1 MiRNAs are short noncoding RNAs that bind to the 3’-UTR of target mRNAs to repress protein production.However,the causal link between miRNAs and TOF and the underlying mechanism has not been established.

The role of immune cells in obesogenic memory

Obesity is a serious chronic disease and worldwide public health problem.According to data from the World Health Organization(WHO),more than 650 million adults in the world were obese and three times more individuals were overweight in 2016.Obesity is associated with type II diabetes,hepatic steatosis,cardiovascular diseases,systemic chronic inflammation,and at least 13 types of cancer.Unfortunately,few effective remedies are available for obesity.The major problem is that most patients suffer from weight regain 2–3 years after obesity treatments,such as obesity drugs,lifestyle interventions(including exercise,diet restrictions,and diet modification),and bariatric surgery.Along with weight regain,most of the initial beneficial changes in metabolomic and transcriptomic profiles dissipate,including insulin sensitivity,the level of adipokines and blood cholesterol.This phenomenon has been defined as“obesogenic memory”.1,2 Obesogenic memory causes repeated weight loss and gain along with obesity therapy,which is called“weight cycling”,leading to a worse healthy status than would result from simple obesity.3,4.