The impact of age and number of mutations on the size of clonal hematopoiesis.

Clonal hematopoiesis (CH) represents the clonal expansion of hematopoietic stem cells and their progeny driven by somatic mutations. Accurate risk assessment of CH is critical for disease prevention and clinical decision-making. The size of CH has been showed to associate with higher disease risk, yet, factors influencing the size of CH are unknown. In addition, the characteristics of CH in long-lived individuals are not well documented. Here, we report an in-depth analysis of CH in longevous (≥90 y old) and common (60~89 y old) elderly groups. Utilizing targeted deep sequencing, we found that the development of CH is closely related to age and the expression of aging biomarkers. The longevous elderly group exhibited a significantly higher incidence of CH and significantly higher frequency of TET2 and ASXL1 mutations, suggesting that certain CH could be beneficial to prolong life. Intriguingly, the size of CH neither correlates significantly to age, in the range of 60 to 110 y old, nor to the expression of aging biomarkers. Instead, we identified a strong correlation between large CH size and the number of mutations per individual. These findings provide a risk assessment biomarker for CH and also suggest that the evolution of the CH is influenced by factor(s) in addition to age.

KCTD10 regulates brain development by destabilizing brain disorder-associated protein KCTD13.

KCTD10 belongs to the KCTD (potassiumchannel tetramerization domain) family, many members of which are associated with neuropsychiatric disorders. However, the biological function underlying the association with brain disorders remains to be explored. Here, we reveal that Kctd10 is highly expressed in neuronal progenitors and layer V neurons throughout brain development. Kctd10 deficiency triggers abnormal proliferation and differentiation of neuronal progenitors, reduced deep-layer (especially layer V) neurons, increased upper-layer neurons, and lowered brain size. Mechanistically, we screened and identified a unique KCTD10-interacting protein, KCTD13, associated with neurodevelopmental disorders. KCTD10 mediated the ubiquitination-dependent degradation of KCTD13 and KCTD10 ablation resulted in a considerable increase of KCTD13 expression in the developing cortex. KCTD13 overexpression in neuronal progenitors led to reduced proliferation and abnormal cell distribution, mirroring KCTD10 deficiency. Notably, mice with brain-specific Kctd10 knockout exhibited obvious motor deficits. This study uncovers the physiological function of KCTD10 and provides unique insights into the pathogenesis of neurodevelopmental disorders.

Quantification of tRNA m1A modification by templated-ligation qPCR.

N1-methyladenosine (m1A) is a widespread modification in all eukaryotic, many archaeal, and some bacterial tRNAs. m1A is generally located in the T loop of cytosolic tRNA and between the acceptor and D stems of mitochondrial tRNAs; it is involved in the tertiary interaction that stabilizes tRNA. Human tRNA m1A levels are dynamically regulated that fine-tune translation and can also serve as biomarkers for infectious disease. Although many methods have been used to measure m1A, a PCR method to assess m1A levels quantitatively in specific tRNAs has been lacking. Here we develop a templated-ligation followed by a qPCR method (TL-qPCR) that measures m1A levels in target tRNAs. Our method uses the SplintR ligase that efficiently ligates two tRNA complementary DNA oligonucleotides using tRNA as the template, followed by qPCR using the ligation product as the template. m1A interferes with the ligation in specific ways, allowing for the quantitative assessment of m1A levels using subnanogram amounts of total RNA. We identify the features of specificity and quantitation for m1A-modified model RNAs and apply these to total RNA samples from human cells. Our method enables easy access to study the dynamics and function of this pervasive tRNA modification.

Mucosal IFNλ1 mRNA-based immunomodulation effectively reduces SARS-CoV-2 induced mortality in mice.

RNA vaccines elicit protective immunity against SARS-CoV-2, but the use of mRNA as an antiviral immunotherapeutic is unexplored. Here, we investigate the activity of lipidoid nanoparticle (LNP)-formulated mRNA encoding human IFNλ1 (ETH47), which is a critical driver of innate immunity at mucosal surfaces protecting from viral infections. IFNλ1 mRNA administration promotes dose-dependent protein translation, induction of interferon-stimulated genes without relevant signs of unspecific immune stimulation, and dose-dependent inhibition of SARS-CoV-2 replication in vitro. Pulmonary administration of IFNλ1 mRNA in mice results in a potent reduction of virus load, virus-induced body weight loss and significantly increased survival. These data support the development of inhaled administration of IFNλ1 mRNA as a potential prophylactic option for individuals exposed to SARS-CoV-2 or at risk suffering from COVID-19. Based on the broad antiviral activity of IFNλ1 regardless of virus or variant, this approach might also be utilized for other respiratory viral infections or pandemic preparedness.

A Functional Assessment of Fetal Liver and Monocyte-Derived Macrophages in the Lung Alveolar Environment.

It is becoming clear that every organ is seeded by a population of fetal liver-derived macrophages that are replaced at different rates by monocyte-derived macrophages. Using the Ms4a3tdTomato reporter mouse that reports on monocyte-derived alveolar macrophages (Mo-AMs) and our ability to examine AM function using our multichannel intravital microscopy, we examined the fetal-liver derived alveolar macrophage (FL-AM) and Mo-AM populations within the same mouse under various environmental conditions. The experiments unveiled that AMs migrated from alveolus to alveolus and phagocytosed bacteria identically regardless of ontogenic origin. Using 50 PFU of influenza A virus (IAV) determined using the Madin-Darby canine kidney (MDCK) cell line, we noted that both populations were susceptible to IAV-induced immunoparalysis, which also led to impaired phagocytosis of secondary bacterial infections. Both FL-AMs and Mo-AMs were trained by ß-glucan to resist IAV-induced paralysis. Over time (40 wk), Mo-AMs began to outperform FL-AMs, although both populations were still sensitive to IAV. Our data also show that clodronate depletion of AMs leads to replenishment, but by FL-AMs, and these macrophages do show some functional impairment for a limited time. Overall, the system is designed such that new macrophages rapidly assume the function of tissue-resident macrophages when both populations are examined in an identical environment. These data do differ from artificial depletion methods that compare Mo-AMs and FL-AMs.

CARM1 hypermethylates the NuRD chromatin remodeling complex to promote cell cycle gene expression and breast cancer development.

Protein arginine methyltransferase CARM1 has been shown to methylate a large number of non-histone proteins, and play important roles in gene transcriptional activation, cell cycle progress, and tumorigenesis. However, the critical substrates through which CARM1 exerts its functions remain to be fully characterized. Here, we reported that CARM1 directly interacts with the GATAD2A/2B subunit in the nucleosome remodeling and deacetylase (NuRD) complex, expanding the activities of NuRD to include protein arginine methylation. CARM1 and NuRD bind and activate a large cohort of genes with implications in cell cycle control to facilitate the G1 to S phase transition. This gene activation process requires CARM1 to hypermethylate GATAD2A/2B at a cluster of arginines, which is critical for the recruitment of the NuRD complex. The clinical significance of this gene activation mechanism is underscored by the high expression of CARM1 and NuRD in breast cancers, and the fact that knockdown CARM1 and NuRD inhibits cancer cell growth in vitro and tumorigenesis in vivo. Targeting CARM1-mediated GATAD2A/2B methylation with CARM1 specific inhibitors potently inhibit breast cancer cell growth in vitro and tumorigenesis in vivo. These findings reveal a gene activation program that requires arginine methylation established by CARM1 on a key chromatin remodeler, and targeting such methylation might represent a promising therapeutic avenue in the clinic.

Reversible Fluorescent Probes for Dynamic Imaging of Liver Ischemia-Reperfusion Injury.

ConspectusHepatic ischemia-reperfusion injury (HIRI) is an inevitable complication of clinical surgeries such as liver resection or transplantation, often resulting in postoperative liver dysfunction, hepatic failure in up to 13% of postresection patients, and early graft failure in 11-18% of liver transplantation patients. HIRI involves a series of biochemical events triggered by abnormal alterations in multiple biomarkers, characterized by short lifespans, dynamic changes, subcellular regional distribution, and multicollaborative regulation. However, traditional diagnosis, including serology, imaging, and liver puncture biopsy, suffers from low sensitivity, poor resolution, and hysteresis, which hinder effective monitoring of HIRI markers. Thus, to address the unique properties of HIRI markers, there is a pressing demand for developing novel detection strategies that are highly selective, transiently responsive, dynamically reversible, subcellular organelle-targeted, and capable of simultaneous multicomponent analysis.Optical probe-based fluorescence imaging is a powerful tool for real-time monitoring of biomarkers with the advantages of high sensitivity, noninvasiveness, rapid analysis, and high-fidelity acquisition of spatiotemporal information on signaling molecules compared with conventional methods. Moreover, with the growing demand for continuous monitoring of biomarkers, probes with reversible detection features are receiving more and more attention. Importantly, reversible probes can not only monitor fluctuations in marker concentrations but also distinguish between transient bursts of markers during physiological events and long-term sustained increases in pathological marker levels. This can effectively avoid false-positive test results, and in addition, reversible probes can be reutilized with green and economical features. Therefore, our team has employed various effective methods to design reversible optical probes for HIRI. We proposed reversible recognition strategies based on specific reactions or interactions to detect dynamic changes in markers. Given the biomarkers' unique signaling in subcellular organelles and the synergistic regulatory properties of multiple markers for HIRI, bifunctional reversible detection strategies are exploited, including organelle-targeted reversible and multicomponent simultaneous detection. With these strategies, we have tailored a variety of high-fidelity fluorescent probes for a series of HIRI markers, including reactive oxygen/nitrogen species (O2•- and ONOO-), ATP, protein (Keap1), mitochondrial DNA, etc. Utilizing the probes, the in situ dynamic imaging detection of the HIRI markers was successfully achieved. While performing the precise examination of the earlier occurrence of HIRI disease and visualizing the real-time monitoring of the disease process, we have also further elucidated the HIRI-associated signaling pathways. It is envisioned that our summarized work will inspire the design of future reversible fluorescent probes and help to improve the clinical diagnosis and therapeutic efficiency of these diseases.

Mutations in CLCN6 as a Novel Genetic Cause of Neuronal Ceroid Lipofuscinosis in a Murine Model.

OBJECTIVE: The aim of this study was to explore the pathogenesis of CLCN6-related disease and to assess whether its Cl-/H+-exchange activity is crucial for the biological role of ClC-6. METHODS: We performed whole-exome sequencing on a girl with development delay, intractable epilepsy, behavioral abnormities, retinal dysfunction, progressive brain atrophy, suggestive of neuronal ceroid lipofuscinoses (NCLs). We generated and analyzed the first knock-in mouse model of a patient variant (p.E200A) and compared it with a Clcn6-/- mouse model. Additional functional tests were performed with heterologous expression of mutant ClC-6. RESULTS: We identified a de novo heterozygous p.E200A variant in the proband. Expression of disease-causing ClC-6E200A or ClC-6Y553C mutants blocked autophagic flux and activated transcription factors EB (TFEB) and E3 (TFE3), leading to autophagic vesicle and cholesterol accumulation. Such alterations were absent with a transport-deficient ClC-6E267A mutant. Clcn6E200A/+ mice developed severe neurodegeneration with typical features of NCLs. Mutant ClC-6E200A, but not loss of ClC-6 in Clcn6-/- mice, increased lysosomal biogenesis by suppressing mTORC1-TFEB signaling, blocked autophagic flux through impairing lysosomal function, and increased apoptosis. Carbohydrate and lipid deposits accumulated in Clcn6E200A/+ brain, while only lipid storage was found in Clcn6-/- brain. Lysosome dysfunction, autophagy defects, and gliosis were early pathogenic events preceding neuron loss. INTERPRETATION: CLCN6 is a novel genetic cause of NCLs, highlighting the importance of considering CLCN6 mutations in the diagnostic workup for molecularly undefined forms of NCLs. Uncoupling of Cl- transport from H+ countertransport in the E200A mutant has a dominant effect on the autophagic/lysosomal pathway. ANN NEUROL 2024;96:608-624.

A disinhibitory microcircuit of the orbitofrontal cortex mediates cocaine preference in mice.

Both clinical and animal studies showed that the impaired functions of the orbitofrontal cortex (OFC) underlie the compulsive drug-seeking behavior of drug addiction. However, the functional changes of the microcircuit in the OFC and the underlying molecular mechanisms in drug addiction remain elusive, and little is known for whether microcircuits in the OFC contributed to drug addiction-related behaviors. Utilizing the cocaine-induced conditioned-place preference model, we found that the malfunction of the microcircuit led to disinhibition in the OFC after cocaine withdrawal. We further showed that enhanced Somatostatin-Parvalbumin (SST-PV) inhibitory synapse strength changed microcircuit function, and SST and PV inhibitory neurons showed opposite contributions to the drug addiction-related behavior of mice. Brevican of the perineuronal nets of PV neurons regulated SST-PV synapse strength, and the knockdown of Brevican alleviated cocaine preference. These results reveal a novel molecular mechanism of the regulation of microcircuit function and a novel circuit mechanism of the OFC in gating cocaine preference.

Memory consolidation drives the enhancement of remote cocaine memory via prefrontal circuit.

Remote memory usually decreases over time, whereas remote drug-cue associated memory exhibits enhancement, increasing the risk of relapse during abstinence. Memory system consolidation is a prerequisite for remote memory formation, but neurobiological underpinnings of the role of consolidation in the enhancement of remote drug memory are unclear. Here, we found that remote cocaine-cue associated memory was enhanced in rats that underwent self-administration training, together with a progressive increase in the response of prelimbic cortex (PrL) CaMKII neurons to cues. System consolidation was required for the enhancement of remote cocaine memory through PrL CaMKII neurons during the early period post-training. Furthermore, dendritic spine maturation in the PrL relied on the basolateral amygdala (BLA) input during the early period of consolidation, contributing to remote memory enhancement. These findings indicate that memory consolidation drives the enhancement of remote cocaine memory through a time-dependent increase in activity and maturation of PrL CaMKII neurons receiving a sustained BLA input.

Engineered mischarged transfer RNAs for correcting pathogenic missense mutations.

Missense mutations account for approximately 50% of pathogenic mutations in human genetic diseases, and most lack effective treatments. Gene therapies, gene editing, and RNA therapies, including transfer RNA (tRNA) modalities, are common strategies for genetic disease treatments. However, reported tRNA therapies are for nonsense mutations only. It has not been explored how tRNAs can be engineered to correct missense mutations. Here, we describe missense-correcting tRNAs (mc-tRNAs) as a potential therapeutic for correcting pathogenic missense mutations. Mc-tRNAs are engineered tRNAs charged with one amino acid, but read codons of another in translation. We first developed a series of fluorescent protein-based reporters that indicate the successful correction of missense mutations via restoration of fluorescence. We engineered mc-tRNAs that effectively corrected serine and arginine missense mutations in the reporters and confirmed the amino acid substitution by mass spectrometry and mc-tRNA expression by sequencing. We examined the transcriptome response to mc-tRNA expression and found some mc-tRNAs induced minimum transcriptomic changes. Furthermore, we applied an mc-tRNA to rescue a pathogenic CAPN3 Arg-to-Gln mutant involved in LGMD2A. These results establish a versatile pipeline for mc-tRNA engineering and demonstrate the potential of mc-tRNA as an alternative therapeutic platform for the treatment of genetic disorders.

Chiral bisphosphine Ph-BPE ligand: a rising star in asymmetric synthesis.

Chiral 1,2-bis(2,5-diphenylphospholano)ethane (Ph-BPE) is a class of optimal organic bisphosphine ligands with C2-symmetry. Ph-BPE with its excellent catalytic performance in asymmetric synthesis has attracted much attention of chemists with increasing popularity and is growing into one of the most commonly used organophosphorus ligands, especially in asymmetric catalysis. Over two hundred examples have been reported since 2012. This review presents how Ph-BPE is utilized in asymmetric synthesis and how powerful it is as a chiral ligand or even a catalyst in a wide range of reactions including applications in the total synthesis of bioactive molecules.

What can molecular assembly learn from catalysed assembly in living organisms?

Molecular assembly is the process of organizing individual molecules into larger structures and complex systems. The self-assembly approach is predominantly utilized in creating artificial molecular assemblies, and was believed to be the primary mode of molecular assembly in living organisms as well. However, it has been shown that the assembly of many biological complexes is "catalysed" by other molecules, rather than relying solely on self-assembly. In this review, we summarize these catalysed-assembly (catassembly) phenomena in living organisms and systematically analyse their mechanisms. We then expand on these phenomena and discuss related concepts, including catalysed-disassembly and catalysed-reassembly. Catassembly proves to be an efficient and highly selective strategy for synergistically controlling and manipulating various noncovalent interactions, especially in hierarchical molecular assemblies. Overreliance on self-assembly may, to some extent, hinder the advancement of artificial molecular assembly with powerful features. Furthermore, inspired by the biological catassembly phenomena, we propose guidelines for designing artificial catassembly systems and developing characterization and theoretical methods, and review pioneering works along this new direction. Overall, this approach may broaden and deepen our understanding of molecular assembly, enabling the construction and control of intelligent assembly systems with advanced functionality.

Optimising first-line subtyping-based therapy in triple-negative breast cancer (FUTURE-SUPER): a multi-cohort, randomised, phase 2 trial.

BACKGROUND: Triple-negative breast cancers display heterogeneity in molecular drivers and immune traits. We previously classified triple-negative breast cancers into four subtypes: luminal androgen receptor (LAR), immunomodulatory, basal-like immune-suppressed (BLIS), and mesenchymal-like (MES). Here, we aimed to evaluate the efficacy and safety of subtyping-based therapy in the first-line treatment of triple-negative breast cancer. METHODS: FUTURE-SUPER is an ongoing, open-label, randomised, controlled phase 2 trial being conducted at Fudan University Shanghai Cancer Center (FUSCC), Shanghai, China. Eligible participants were females aged 18-70 years, with an Eastern Cooperative Oncology Group performance status of 0-1, and histologically confirmed, untreated metastatic or recurrent triple-negative breast cancer. After categorising participants into five cohorts according to molecular subtype and genomic biomarkers, participants were randomly assigned (1:1) with a block size of 4, stratified by subtype, to receive, in 28-day cycles, nab-paclitaxel (100 mg/m2, intravenously on days 1, 8, and 15) alone (control group) or with a subtyping-based regimen (subtyping-based group): pyrotinib (400 mg orally daily) for the LAR-HER2mut subtype, everolimus (10 mg orally daily) for the LAR-PI3K/AKTmut and MES-PI3K/AKTmut subtypes, camrelizumab (200 mg intravenously on days 1 and 15) and famitinib (20 mg orally daily) for the immunomodulatory subtype, and bevacizumab (10 mg/kg intravenously on days 1 and 15) for the BLIS/MES-PI3K/AKTWT subtype. The primary endpoint was investigator-assessed progression-free survival for the pooled subtyping-based group versus the control group in the intention-to-treat population (all randomly assigned participants). Safety was analysed in all patients with safety records who received at least one dose of study drug. This study is registered with ClinicalTrials.gov (NCT04395989). FINDINGS: Between July 28, 2020, and Oct 16, 2022, 139 female participants were enrolled and randomly assigned to the subtyping-based group (n=69) or control group (n=70). At the data cutoff (May 31, 2023), the median follow-up was 22·5 months (IQR 15·2-29·0). Median progression-free survival was significantly longer in the pooled subtyping-based group (11·3 months [95% CI 8·6-15·2]) than in the control group (5·8 months [4·0-6·7]; hazard ratio 0·44 [95% CI 0·30-0·65]; p<0·0001). The most common grade 3-4 treatment-related adverse events were neutropenia (21 [30%] of 69 in the pooled subtyping-based group vs 16 [23%] of 70 in the control group), anaemia (five [7%] vs none), and increased alanine aminotransferase (four [6%] vs one [1%]). Treatment-related serious adverse events were reported for seven (10%) of 69 patients in the subtyping-based group and none in the control group. No treatment-related deaths were reported in either group. INTERPRETATION: These findings highlight the potential clinical benefits of using molecular subtype-based treatment optimisation in patients with triple-negative breast cancer, suggesting a path for further clinical investigation. Phase 3 randomised clinical trials assessing the efficacy of subtyping-based regimens are now underway. FUNDING: National Natural Science Foundation of China, Natural Science Foundation of Shanghai, Shanghai Hospital Development Center, and Jiangsu Hengrui Pharmaceuticals. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.

Rapid turnover and short-term blooms of Escherichia coli in the human gut.

Escherichia coli (E. coli) is a common microorganism that is widely present in the environment and closely related to human health. The extent of E. coli presence in the human gut has been a subject of ongoing debate. Through whole-genome shotgun metagenomic sequencing, our study revealed that E. coli exists in the human body at a low abundance (average abundance 1.21%), with occasional short-term bursts leading to temporary increases in abundance, with the highest recorded at 50.91%. Further investigations into the factors contributing to these short-term blooms of E. coli showed significant variations in strain types and genomes within fecal samples collected from the same individuals at different time points. Evolutionary tree analysis indicated that samples from different individuals crossed, suggesting a change in the dominant E. coli strains within the human gut. Therefore, it can be inferred that E. coli in the human body are more likely to be transient bacteria rather than permanent residents in the gut. The rapid rate of turnover among months (87.5% within a month) and short-term blooms of E. coli in the human body can establish "latent infections" of nonpathogenic strains in healthy individuals while also posing a potential risk of introducing pathogenic strains, thereby impacting human health. In summary, our study revealed the variation in E. coli abundance and strains within the human gut, influenced by geographic area and temporal factors. These findings contribute to a better understanding of the relationship between E. coli, the gut microbiota, and human health. IMPORTANCE Escherichia coli (E. coli) is a microorganism closely linked to human health, and its presence in the human gut has been a topic of debate. Our study, using whole-genome shotgun metagenomic sequencing, revealed that E. coli exists at a low abundance in the human body, with occasional short-term bursts leading to temporary increases. Strain and genome variations were observed within fecal samples from the same individuals at different time points, suggesting transient rather than permanent residence of E. coli in the gut. The rapid turnover rate and short-term blooms of E. coli can establish latent infections while also posing a risk of introducing pathogenic strains. These findings enhance our understanding of the relationship between E. coli, the gut microbiota, and human health.

SEPHS1 attenuates intervertebral disc degeneration by delaying nucleus pulposus cell senescence through the Hippo-Yap/Taz pathway.

Nucleus pulposus cell (NPC) senescence is a major cause of intervertebral disc degeneration (IVDD). Oxidative stress and reactive oxygen species (ROS) play critical roles in regulating cell senescence. Selenophosphate synthetase 1 (SEPHS1) was reported to play an important role in mitigating oxidative stress in an osteoarthritis (OA) model by reducing the production of ROS, thereby, delaying the occurrence and development of osteoarthritis. In this study, we explored the, hitherto unknown, role of SEPHS1 in IVDD in vitro and in vivo using an interleukin-1ß (IL-1ß)-induced NPC senescence model and a rat needle puncture IVDD model, respectively. SEPHS1 delayed NPC senescence in vitro by reducing ROS production. Age-related dysfunction was also ameliorated by the overexpression of SEPHS1 and inhibition of the Hippo-Yap/Taz signaling pathway. In vivo experiments revealed that the overexpression of SEPHS1 and inhibition of Hippo-Yap/Taz alleviated IVDD in rats. Moreover, a selenium (Se)-deficient diet and lack of SEPHS1 synergistically aggravated IVDD progression. Taken together, our results demonstrate that SEPHS1 plays a significant role in NPC senescence. Overexpression of SEPHS1 and inhibition of Hippo-Yap/Taz can delay NPC senescence, restore the balance of extracellular matrix metabolism, and attenuate IVDD. SEPHS1 could be a promising therapeutic target for IVDD.NEW & NOTEWORTHY Selenophosphate synthetase 1 (SEPHS1) deficiency leads to an increase in reactive oxygen species levels and in the subsequent activation of the Hippo-Yap/Taz signaling pathway. In the rat model of intervertebral disc degeneration (IVDD), overexpression of SEPHS1 and inhibition of Hippo-YAP/Taz mitigated the progression of disc degeneration indicating the involvement of SEPHS1 in IVDD. SEPHS1 is a promising therapeutic target for IVDD.

Activation of the alternative complement pathway modulates inflammation in thoracic aortic aneurysm/dissection.

Thoracic aortic aneurysm/dissection (TAAD) is a lethal vascular disease, and several pathological factors participate in aortic medial degeneration. We previously discovered that the complement C3a-C3aR axis in smooth muscle cells promotes the development of thoracic aortic dissection (TAD) through regulation of matrix metalloproteinase 2. However, discerning the specific complement pathway that is activated and elucidating how inflammation of the aortic wall is initiated remain unknown. We ascertained that the plasma levels of C3a and C5a were significantly elevated in patients with TAD and that the levels of C3a, C4a, and C5a were higher in acute TAD than in chronic TAD. We also confirmed the activation of the complement in a TAD mouse model. Subsequently, knocking out Cfb (Cfb) or C4 in mice with TAD revealed that the alternative pathway and Cfb played a significant role in the TAD process. Activation of the alternative pathway led to generation of the anaphylatoxins C3a and C5a, and knocking out their receptors reduced the recruitment of inflammatory cells to the aortic wall. Moreover, we used serum from wild-type mice or recombinant mice Cfb as an exogenous source of Cfb to treat Cfb KO mice and observed that it exacerbated the onset and rupture of TAD. Finally, we knocked out Cfb in the FBN1C1041G/+ Marfan-syndrome mice and showed that the occurrence of TAA was reduced. In summary, the alternative complement pathway promoted the development of TAAD by recruiting infiltrating inflammatory cells. Targeting the alternative pathway may thus constitute a strategy for preventing the development of TAAD.NEW & NOTEWORTHY The alternative complement pathway promoted the development of TAAD by recruiting infiltrating inflammatory cells. Targeting the alternative pathway may thus constitute a strategy for preventing the development of TAAD.

Zinc finger BED-type containing 3 promotes hepatic steatosis by interacting with polypyrimidine tract-binding protein 1.

AIMS/HYPOTHESIS: The relationship between metabolic dysfunction-associated steatotic liver disease (MASLD) and type 2 diabetes mellitus, insulin resistance and the metabolic syndrome is well established. While zinc finger BED-type containing 3 (ZBED3) has been linked to type 2 diabetes mellitus and the metabolic syndrome, its role in MASLD remains unclear. In this study, we aimed to investigate the function of ZBED3 in the context of MASLD. METHODS: Expression levels of ZBED3 were assessed in individuals with MASLD, as well as in cellular and animal models of MASLD. In vitro and in vivo analyses were conducted using a cellular model of MASLD induced by NEFA and an animal model of MASLD induced by a high-fat diet (HFD), respectively, to investigate the role of ZBED3 in MASLD. ZBED3 expression was increased by lentiviral infection or tail-vein injection of adeno-associated virus. RNA-seq and bioinformatics analysis were employed to examine the pathways through which ZBED3 modulates lipid accumulation. Findings from these next-generation transcriptome sequencing studies indicated that ZBED3 controls SREBP1c (also known as SREBF1; a gene involved in fatty acid de novo synthesis); thus, co-immunoprecipitation and LC-MS/MS were utilised to investigate the molecular mechanisms by which ZBED3 regulates the sterol regulatory element binding protein 1c (SREBP1c). RESULTS: In this study, we found that ZBED3 was significantly upregulated in the liver of individuals with MASLD and in MASLD animal models. ZBED3 overexpression promoted NEFA-induced triglyceride accumulation in hepatocytes in vitro. Furthermore, the hepatocyte-specific overexpression of Zbed3 promoted hepatic steatosis. Conversely, the hepatocyte-specific knockout of Zbed3 resulted in resistance of HFD-induced hepatic steatosis. Mechanistically, ZBED3 interacts directly with polypyrimidine tract-binding protein 1 (PTBP1) and affects its binding to the SREBP1c mRNA precursor to regulate SREBP1c mRNA stability and alternative splicing. CONCLUSIONS/INTERPRETATION: This study indicates that ZBED3 promotes hepatic steatosis and serves as a critical regulator of the progression of MASLD. DATA AVAILABILITY: RNA-seq data have been deposited in the NCBI Gene Expression Omnibus ( www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE231875 ). MS proteomics data have been deposited to the ProteomeXchange Consortium via the iProX partner repository ( https://proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD041743 ).

Pituitary transcriptome profile from laying period to incubation period of Changshun green-shell laying hens.

BACKGROUND: Incubation behaviour, an instinct for natural breeding in poultry, is strictly controlled by the central nervous system and multiple neuroendocrine hormones and neurotransmitters, and is closely associated with the cessation of egg laying. Therefore, it is essential for the commercial poultry industry to clarify the molecular regulation mechanism of incubation behaviour. Here, we used high-throughput sequencing technology to examine the pituitary transcriptome of Changshun green-shell laying hen, a local breed from Guizhou province, China, with strong broodiness, in two reproductive stages, including egg-laying phase (LP) and incubation phase (BP). We also analyze the differences in gene expression during the transition from egg-laying to incubation, and identify critical pathways and candidate genes involved in controlling the incubation behaviour in the pituitary. RESULTS: In this study, we demonstrated that a total of 2089 differently expressed genes (DEGs) were identified in the pituitary, including 842 up-regulated and 1247 down-regulated genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that steroid biosynthesis pathway and neuroactive ligand-receptor interaction were significantly enriched based on DEGs commonly identified in pituitary. Further analysis revealed that SRC, ITGB4, ITGB3, PIK3R3 and DRD2 may play crucial roles in the regulation of incubation behaviour. CONCLUSIONS: We identified 2089 DEGs and the key signaling pathways which may be closely correlated with incubation in Changshun green-shell laying hens, and clarified the molecular regulation mechanism of incubation behaviour. Our results indicate the complexity and variety of differences in reproductive behaviour of different chicken breeds.

Discovery of novel RNA viruses through analysis of fungi-associated next-generation sequencing data.

BACKGROUND: Like all other species, fungi are susceptible to infection by viruses. The diversity of fungal viruses has been rapidly expanding in recent years due to the availability of advanced sequencing technologies. However, compared to other virome studies, the research on fungi-associated viruses remains limited. RESULTS: In this study, we downloaded and analyzed over 200 public datasets from approximately 40 different Bioprojects to explore potential fungal-associated viral dark matter. A total of 12 novel viral sequences were identified, all of which are RNA viruses, with lengths ranging from 1,769 to 9,516 nucleotides. The amino acid sequence identity of all these viruses with any known virus is below 70%. Through phylogenetic analysis, these RNA viruses were classified into different orders or families, such as Mitoviridae, Benyviridae, Botourmiaviridae, Deltaflexiviridae, Mymonaviridae, Bunyavirales, and Partitiviridae. It is possible that these sequences represent new taxa at the level of family, genus, or species. Furthermore, a co-evolution analysis indicated that the evolutionary history of these viruses within their groups is largely driven by cross-species transmission events. CONCLUSIONS: These findings are of significant importance for understanding the diversity, evolution, and relationships between genome structure and function of fungal viruses. However, further investigation is needed to study their interactions.