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One of the main drivers of autism spectrum disorder is risk alleles within hundreds of genes, which may interact within shared but unknown protein complexes. Here we develop a scalable genome-editing-mediated approach to target 14 high-confidence autism risk genes within the mouse brain for proximity-based endogenous proteomics, achieving the identification of high-specificity spatial proteomes. The resulting native proximity proteomes are enriched for human genes dysregulated in the brain of autistic individuals, and reveal proximity interactions between proteins from high-confidence risk genes with those of lower-confidence that may provide new avenues to prioritize genetic risk. Importantly, the datasets are enriched for shared cellular functions and genetic interactions that may underlie the condition. We test this notion by spatial proteomics and CRISPR-based regulation of expression in two autism models, demonstrating functional interactions that modulate mechanisms of their dysregulation. Together, these results reveal native proteome networks in vivo relevant to autism, providing new inroads for understanding and manipulating the cellular drivers underpinning its etiology.
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Trastorno del Espectro Autista , Trastorno Autístico , Encéfalo , Modelos Animales de Enfermedad , Proteoma , Proteómica , Animales , Proteoma/metabolismo , Ratones , Humanos , Encéfalo/metabolismo , Proteómica/métodos , Trastorno Autístico/genética , Trastorno Autístico/metabolismo , Trastorno del Espectro Autista/metabolismo , Trastorno del Espectro Autista/genética , Fenotipo , Edición Génica , Masculino , Predisposición Genética a la Enfermedad , Ratones Endogámicos C57BL , Femenino , Sistemas CRISPR-CasRESUMEN
Prader-Willi Syndrome (PWS) is caused by loss of expression of paternally expressed genes in the human 15q11.2-q13 imprinting domain. A set of imprinted genes that are active on the paternal but silenced on the maternal chromosome are intricately regulated by a bipartite imprinting center (PWS-IC) located in the PWS imprinting domain. In past work, we discovered that euchromatic histone lysine N-methyltransferase-2 (EHMT2/G9a) inhibitors were capable of un-silencing PWS-associated genes by restoring their expression from the maternal chromosome. Here, in mice lacking the Ehmt2 gene, we document un-silencing of the imprinted Snrpn/Snhg14 gene on the maternal chromosome in the late embryonic and postnatal brain. Using PWS and Angelman syndrome patient derived cells with either paternal or maternal deletion of 15q11-q13, we have found that chromatin of maternal PWS-IC is closed and has compact 3D folding confirmation. We further show that a new and distinct noncoding RNA preferentially transcribed from upstream of the PWS-IC interacts with EHMT2 and forms a heterochromatin complex to silence gene expression of SNRPN in CIS on maternal chromosome. Taken together, these findings demonstrate that allele-specific recruitment of EHMT2 is required to maintain the maternal imprints. Our findings provide novel mechanistic insights and support a new model for imprinting maintenance of the PWS imprinted domain.
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BACKGROUND: Autism Spectrum Disorder (ASD) is a group of neurodevelopmental disorders with higher incidence in males and is characterized by atypical verbal/nonverbal communication, restricted interests that can be accompanied by repetitive behavior, and disturbances in social behavior. This study investigated brain mechanisms that contribute to sociability deficits and sex differences in an ASD animal model. METHODS: Sociability was measured in C58/J and C57BL/6J mice using the 3-chamber social choice test. Bulk RNA-Seq and snRNA-Seq identified transcriptional changes in C58/J and C57BL/6J amygdala within which DMRseq was used to measure differentially methylated regions in amygdala. RESULTS: C58/J mice displayed divergent social strata in the 3-chamber test. Transcriptional and pathway signatures revealed immune-related biological processes differ between C58/J and C57BL/6J amygdala. Hypermethylated and hypomethylated genes were identified in C58/J versus C57BL/6J amygdala. snRNA-Seq data in C58/J amygdala identified differential transcriptional signatures within oligodendrocytes and microglia characterized by increased ASD risk gene expression and predicted impaired myelination that was dependent on sex and sociability. RNA velocity, gene regulatory network, and cell communication analysis showed diminished oligodendrocyte/microglia differentiation. Findings were verified using Bulk RNA-Seq and demonstrated oxytocin's beneficial effects on myelin gene expression. LIMITATIONS: Our findings are significant. However, limitations can be noted. The cellular mechanisms linking reduced oligodendrocyte differentiation and reduced myelination to an ASD phenotype in C58/J mice need further investigation. Additional snRNA-Seq and spatial studies would determine if effects in oligodendrocytes/microglia are unique to amygdala or if this occurs in other brain regions. Oxytocin's effects need further examination to understand its' potential as an ASD therapeutic. CONCLUSIONS: Our work demonstrates the C58/J mouse model's utility in evaluating the influence of sex and sociability on the transcriptome in concomitant brain regions involved in ASD. Our single-nucleus transcriptome analysis elucidates potential pathological roles of oligodendrocytes and microglia in ASD. This investigation provides details regarding regulatory features disrupted in these cell types, including transcriptional gene dysregulation, aberrant cell differentiation, altered gene regulatory networks, and changes to key pathways that promote microglia/oligodendrocyte differentiation. Our studies provide insight into interactions between genetic risk and epigenetic processes associated with divergent affiliative behavior and lack of positive sociability.
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Amígdala del Cerebelo , Trastorno del Espectro Autista , Ratones Endogámicos C57BL , Microglía , Oligodendroglía , Conducta Social , Animales , Masculino , Microglía/metabolismo , Ratones , Amígdala del Cerebelo/metabolismo , Femenino , Oligodendroglía/metabolismo , Trastorno del Espectro Autista/genética , Trastorno del Espectro Autista/patología , Perfilación de la Expresión Génica/métodos , Fenotipo , Caracteres Sexuales , Transcriptoma , Modelos Animales de Enfermedad , Oxitocina/genética , Oxitocina/metabolismoRESUMEN
Precision of transcription is critical because transcriptional dysregulation is disease causing. Traditional methods of transcriptional profiling are inadequate to elucidate the full spectrum of the transcriptome, particularly for longer and less abundant mRNAs. SHANK3 is one of the most common autism causative genes. Twenty-four Shank3-mutant animal lines have been developed for autism modeling. However, their preclinical validity has been questioned due to incomplete Shank3 transcript structure. We apply an integrative approach combining cDNA-capture and long-read sequencing to profile the SHANK3 transcriptome in humans and mice. We unexpectedly discover an extremely complex SHANK3 transcriptome. Specific SHANK3 transcripts are altered in Shank3-mutant mice and postmortem brain tissues from individuals with autism spectrum disorder. The enhanced SHANK3 transcriptome significantly improves the detection rate for potential deleterious variants from genomics studies of neuropsychiatric disorders. Our findings suggest that both deterministic and stochastic transcription of the genome is associated with SHANK family genes.
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Trastorno Autístico , Proteínas del Tejido Nervioso , Animales , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Humanos , Ratones , Trastorno Autístico/genética , Transcripción Genética , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Transcriptoma/genética , Trastorno del Espectro Autista/genética , Procesos Estocásticos , MasculinoRESUMEN
Prader-Willi syndrome (PWS) is the prototypic genomic disorder resulting from deficiency of paternally expressed genes in the human chromosome 15q11-q13 region. The unique molecular mechanism involving epigenetic modifications renders PWS as the most attractive candidate to explore a proof-of-concept of epigenetic therapy in humans. The premise is that epigenetic modulations could reactivate the repressed PWS candidate genes from the maternal chromosome and offer therapeutic benefit. Our prior study identifies an EHMT2/G9a inhibitor, UNC0642, that reactivates the expression of PWS genes via reduction of H3K9me2. However, low brain permeability and poor oral bioavailability of UNC0642 preclude its advancement into translational studies in humans. In this study, a newly developed inhibitor, MS152, modified from the structure of UNC0642, has better brain penetration and greater potency and selectivity against EHMT2/G9a. MS152 reactivated maternally silenced PWS genes in PWS patient fibroblasts and in brain and liver tissues of PWS mouse models. Importantly, the molecular efficacy of oral administration is comparable with the intraperitoneal route. MS152 treatment in newborns ameliorates the perinatal lethality and poor growth, maintaining reactivation in a PWS mouse model at postnatal 90 days. Our findings provide strong support for MS152 as a first-in-class inhibitor to advance the epigenetic therapy of PWS in humans.
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Modelos Animales de Enfermedad , Epigénesis Genética , Síndrome de Prader-Willi , Humanos , Animales , Síndrome de Prader-Willi/tratamiento farmacológico , Síndrome de Prader-Willi/genética , Ratones , Epigénesis Genética/efectos de los fármacos , Administración Oral , Antígenos de Histocompatibilidad/genética , Antígenos de Histocompatibilidad/metabolismo , Encéfalo/metabolismo , Encéfalo/efectos de los fármacos , N-Metiltransferasa de Histona-LisinaRESUMEN
Numerous large scale genomic studies have uncovered rare but recurrent pathogenetic variants in a significant number of genes encoding epigenetic machinery in cases with neurodevelopmental disorders (NDD) especially autism spectrum disorder (ASD). These findings provide strong support for the functional importance of epigenetic regulators in neurodevelopment. After the clinical genomics evaluation of the patients using exome sequencing, we have identified, three novel protein-truncating variants (PTVs) in the MSL2 gene (OMIM: 614802) which encodes a chromatin modifying enzyme. MSL2 modifies chromatin through both mono-ubiquitination of histone 2B on lysine 34 (K34) and acetylation of histone H4 on lysine 16 (K16). We reported first time the detailed clinical features associated with 3 MSL2 PTVs. There are 15 PTVs (13 de novo) reported from the large genomics studies (12 cases) or ClinVar (3 cases) of NDD, ASD, and developmental disorders (DD) but the specific clinical features for these cases are not described. Taken together, our descriptions of dysmorphic face and other features support the causal role of MSL2 in a likely syndromic neurodevelopmental disorder and add MSL2 to a growing list of epigenetic genes implicated in ASD.
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Trastorno del Espectro Autista , Niño , Preescolar , Femenino , Humanos , Masculino , Trastorno del Espectro Autista/genética , Cromatina/genética , Cromatina/metabolismo , Mutación , Trastornos del Neurodesarrollo/genética , Trastornos del Neurodesarrollo/patología , SíndromeRESUMEN
Precision of transcription is critical because transcriptional dysregulation is disease causing. Traditional methods of transcriptional profiling are inadequate to elucidate the full spectrum of the transcriptome, particularly for longer and less abundant mRNAs. SHANK3 is one of the most common autism causative genes. Twenty-four Shank3 mutant animal lines have been developed for autism modeling. However, their preclinical validity has been questioned due to incomplete Shank3 transcript structure. We applied an integrative approach combining cDNA-capture and long-read sequencing to profile the SHANK3 transcriptome in human and mice. We unexpectedly discovered an extremely complex SHANK3 transcriptome. Specific SHANK3 transcripts were altered in Shank3 mutant mice and postmortem brains tissues from individuals with ASD. The enhanced SHANK3 transcriptome significantly improved the detection rate for potential deleterious variants from genomics studies of neuropsychiatric disorders. Our findings suggest the stochastic transcription of genome associated with SHANK family genes.
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Background A retrospective study was performed to evaluate the patterns of cytogenomic findings detected from a case series of products of conception (POC) in recurrent pregnancy loss (RPL) over a 16-year period from 2007 to 2023. Results This case series of RPL was divided into a single analysis (SA) group of 266 women and a consecutive analysis (CA) group of 225 women with two to three miscarriages analyzed. Of the 269 POC from the SA group and the 469 POC from the CA group, a spectrum of cytogenomic abnormalities of simple aneuploidies, compound aneuploidies, polyploidies, and structural rearrangements/pathogenic copy number variants (pCNVs) were detected in 109 (41%) and 160 cases (34%), five (2%) and 11 cases (2%), 35 (13%) and 36 cases (8%), and 10 (4%) and 19 cases (4%), respectively. Patterns with recurrent normal karyotypes, alternating normal and abnormal karyotypes, and recurrent abnormal karyotypes were detected in 74 (33%), 71 (32%), and 80 (35%) of consecutive miscarriages, respectively. Repeat aneuploidies of monosomy X and trisomy 16, triploidy, and tetraploidy were detected in nine women. Conclusions A comparable spectrum of cytogenomic abnormalities was noted in the SA and CA groups of RPL. A skewed likelihood of 2/3 for recurrent normal and abnormal karyotypes and 1/3 for alternating normal and abnormal karyotypes in consecutive miscarriages was observed. Routine cytogenetic analysis should be performed for consecutive miscarriages. Further genomic sequencing to search for detrimental and embryonic lethal variants causing miscarriages and pathogenic variants inducing aneuploidies and polyploidies should be considered for RPL with recurrent normal and abnormal karyotypes.
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Intrathecal injection is a commonly employed procedure in both pediatric and adult clinics, serving as an effective means to administer medications and treatments. By directly delivering medications and treatments into the cerebrospinal fluid of the central nervous system, this method achieves higher localized drug concentrations while reducing systemic side-effects compared to other routes such as intravenous, subcutaneous, or intramuscular injections. Its importance extends beyond clinical settings, as intrathecal injection plays a vital role in preclinical studies focused on treating neurogenetic disorders in rodents and other large animals, including non-human primates. However, despite its widespread application, intrathecal injection in young, particularly neonatal pups, poses significant technical challenges due to their small size and fragile nature. Successful and reliable administration of intrathecal injections in newborn mice requires meticulous attention to detail and careful consideration of various factors. Thus, there is a crucial need for a standardized protocol that not only provides instructions but also highlights key technical considerations and good laboratory practices to ensure procedural consistency, as well as the safety and welfare of the animals. To address this unmet need, we present a detailed and comprehensive protocol for performing intrathecal injections specifically in newborn pups on postnatal day 1 (P1). By following the step-by-step instructions, researchers can confidently perform intrathecal injections in neonatal pups, enabling the accurate delivery of drugs, antisense oligos, and viruses for gene replacement or genome editing-based treatments. Furthermore, the importance of adhering to good laboratory practices is emphasized to maintain the well-being of animals and ensure reliable experimental outcomes. This protocol aims to address the technical challenges associated with intrathecal injections in neonatal mice, ultimately facilitating advances in the field of neurogenetic research that aims to develop potential therapeutic interventions.
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Sistemas de Liberación de Medicamentos , Edición Génica , Adulto , Animales , Ratones , Humanos , Niño , Animales Recién Nacidos , Inyecciones Espinales/métodos , Sistema Nervioso Central , Preparaciones FarmacéuticasRESUMEN
Background: Autism Spectrum Disorder (ASD) is a group of neurodevelopmental disorders with higher incidence in males and is characterized by atypical verbal/nonverbal communication, restricted interests that can be accompanied by repetitive behavior, and disturbances in social behavior. This study investigated brain mechanisms that contribute to sociability deficits and sex differences in an ASD animal model. Methods: Sociability was measured in C58/J and C57BL/6J mice using the 3-chamber social choice test. Bulk RNA-Seq and snRNA-Seq identified transcriptional changes in C58/J and C57BL/6J amygdala within which DMRseq was used to measure differentially methylated regions in amygdala. Results: C58/J mice displayed divergent social strata in the 3-chamber test. Transcriptional and pathway signatures revealed immune-related biological processes differ between C58/J and C57BL/6J amygdala. Hypermethylated and hypomethylated genes were identified in C58/J versus C57BL/6J amygdala. snRNA-Seq data in C58/J amygdala identified differential transcriptional signatures within oligodendrocytes and microglia characterized by increased ASD risk gene expression and predicted impaired myelination that was dependent on sex and sociability. RNA velocity, gene regulatory network, and cell communication analysis showed diminished oligodendrocyte/microglia differentiation. Findings were verified using bulk RNA-Seq and demonstrated oxytocin's beneficial effects on myelin gene expression. Limitations: Our findings are significant. However, limitations can be noted. The cellular mechanisms linking reduced oligodendrocyte differentiation and reduced myelination to an ASD phenotype in C58/J mice need further investigation. Additional snRNA-Seq and spatial studies would determine if effects in oligodendrocytes/microglia are unique to amygdala or if this occurs in other brain regions. Oxytocin's effects need further examination to understand its potential as an ASD therapeutic. Conclusions: Our work demonstrates the C58/J mouse model's utility in evaluating the influence of sex and sociability on the transcriptome in concomitant brain regions involved in ASD. Our single-nucleus transcriptome analysis elucidates potential pathological roles of oligodendrocytes and microglia in ASD. This investigation provides details regarding regulatory features disrupted in these cell types, including transcriptional gene dysregulation, aberrant cell differentiation, altered gene regulatory networks, and changes to key pathways that promote microglia/oligodendrocyte differentiation. Our studies provide insight into interactions between genetic risk and epigenetic processes associated with divergent affiliative behavior and lack of positive sociability.
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BACKGROUND: SHANK3 gene is a highly replicated causative gene for autism spectrum disorder and has been well characterized in multiple Shank3 mutant rodent models. When compared to rodents, domestic dogs are excellent animal models in which to study social cognition as they closely interact with humans and exhibit similar social behaviors. Using CRISPR/Cas9 editing, we recently generated a dog model carrying Shank3 mutations, which displayed a spectrum of autism-like behaviors, such as social impairment and heightened anxiety. However, the neural mechanism underlying these abnormal behaviors remains to be identified. METHODS: We used Shank3 mutant dog models to examine possible relationships between Shank3 mutations and neuronal dysfunction. We studied electrophysiological properties and the synaptic transmission of pyramidal neurons from acute brain slices of the prefrontal cortex (PFC). We also examined dendrite elaboration and dendritic spine morphology in the PFC using biocytin staining and Golgi staining. We analyzed the postsynaptic density using electron microscopy. RESULTS: We established a protocol for the electrophysiological recording of canine brain slices and revealed that excitatory synaptic transmission onto PFC layer 2/3 pyramidal neurons in Shank3 heterozygote dogs was impaired, and this was accompanied by reduced dendrite complexity and spine density when compared to wild-type dogs. Postsynaptic density structures were also impaired in Shank3 mutants; however, pyramidal neurons exhibited hyperexcitability. LIMITATIONS: Causal links between impaired PFC pyramidal neuron function and behavioral alterations remain unclear. Further experiments such as manipulating PFC neuronal activity or restoring synaptic transmission in Shank3 mutant dogs are required to assess PFC roles in altered social behaviors. CONCLUSIONS: Our study demonstrated the feasibility of using canine brain slices as a model system to study neuronal circuitry and disease. Shank3 haploinsufficiency causes morphological and functional abnormalities in PFC pyramidal neurons, supporting the notion that Shank3 mutant dogs are new and valid animal models for autism research.
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Trastorno del Espectro Autista , Trastorno Autístico , Humanos , Perros , Animales , Trastorno Autístico/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Células Piramidales/metabolismo , Transmisión Sináptica/genética , Corteza Prefrontal , Ansiedad , Modelos Animales de EnfermedadRESUMEN
Few studies have examined the association between greenness exposure and birth outcomes. This study aims to identify critical exposure time windows during preconception and pregnancy for the association between greenness exposure and birth weight. A cohort of 13 890 pregnant women and newborns in Shanghai, China from 2016-2019 were included in the study. We assessed greenness exposure using Normalized Difference Vegetation Index (NDVI) during the preconception and gestational periods, and evaluated the association with term birthweight, birthweight z-score, small-for-gestational age, and large-for-gestational age using linear and logistic regressions adjusting for key maternal and newborn covariates. Ambient temperature, relative humidity, ambient levels of fine particles (PM2.5) and nitrogen dioxide (NO2) assessed during the same period were adjusted for as sensitivity analyses. Furthermore, we explored the potential different effects by urbanicity and park accessibility through stratified analysis. We found that higher greenness exposure at the second trimester of pregnancy and averaged exposure during the entire pregnancy were associated with higher birthweight and birthweight Z-score. Specifically, a 0.1 unit increase in second trimester averaged NDVI value was associated with an increase in birthweight of 10.2 g (95% CI: 1.8-18.5 g) and in birthweight Z-score of 0.024 (0.003-0.045). A 0.1 unit increase in an averaged NDVI during the entire pregnancy was associated with 10.1 g (95% CI: 1.0-19.2 g) increase in birthweight and 0.025 (0.001-0.048) increase in birthweight Z-score. Moreover, the associations were larger in effect size among urban residents than suburban residents and among residents without park accessibility within 500 m compared to those with park accessibility within 500 m. Our findings suggest that increased greenness exposure, particularly during the second trimester, may be beneficial to birth weight in a metropolitan area.
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BACKGROUND: Few studies have assessed air pollution exposure association with birthweight during both preconception and gestational periods. METHODS: Leveraging a preconception cohort consisting of 14220 pregnant women and newborn children in Shanghai, China during 2016-2018, we aim to assess associations of NO2 and PM2.5 exposure, derived from high-resolution spatial-temporal models, during preconception and gestational periods with outcomes including term birthweight, birthweight Z-score, small-for-gestational age (SGA) and large-for-gestational age (LGA). Linear and logistic regressions were used to estimate 3-month preconception and trimester-averaged air pollution exposure associations; and distributed lag models (DLM) were used to identify critical exposure windows at the weekly resolution from preconception to delivery. Two-pollutant models and children's sex-specific associations were explored. RESULTS: After controlling for covariates, one standard deviation (SD) (11.5 µg/m3, equivalent to 6.1 ppb) increase in NO2 exposure during the second and the third trimester was associated with 13% (95% confidence interval: 2 - 26%) and 14% (95% CI: 1 - 29%) increase in SGA, respectively; and one SD (9.6 µg/m3) increase in PM2.5 exposure during the third trimester was associated with 15% (95% CI: 1 - 31%) increase in SGA. No association have been found for outcomes of birthweight, birthweight Z-score and LGA. DLM found that gestational weeks 22-32 were a critical window, when NO2 exposure had strongest associations with SGA. The associations of air pollution exposure tended to be stronger in female newborns than in male newborns. However, no significant associations of air pollution exposure during preconception period on birthweight outcomes were found. CONCLUSION: Consistent with previous studies, we found that air pollution exposure during mid-to-late pregnancy was associated with adverse birthweight outcomes.
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Contaminantes Atmosféricos , Contaminación del Aire , Femenino , Recién Nacido , Embarazo , Masculino , Humanos , Peso al Nacer , Contaminantes Atmosféricos/efectos adversos , Contaminantes Atmosféricos/análisis , Estudios Prospectivos , Dióxido de Nitrógeno/análisis , Exposición Materna/efectos adversos , China/epidemiología , Contaminación del Aire/análisis , Retardo del Crecimiento Fetal/inducido químicamente , Material Particulado/análisisRESUMEN
Despite intensive studies in modeling neuropsychiatric disorders especially autism spectrum disorder (ASD) in animals, many challenges remain. Genetic mutant mice have contributed substantially to the current understanding of the molecular and neural circuit mechanisms underlying ASD. However, the translational value of ASD mouse models in preclinical studies is limited to certain aspects of the disease due to the apparent differences in brain and behavior between rodents and humans. Non-human primates have been used to model ASD in recent years. However, a low reproduction rate due to a long reproductive cycle and a single birth per pregnancy, and an extremely high cost prohibit a wide use of them in preclinical studies. Canine model is an appealing alternative because of its complex and effective dog-human social interactions. In contrast to non-human primates, dog has comparable drug metabolism as humans and a high reproduction rate. In this study, we aimed to model ASD in experimental dogs by manipulating the Shank3 gene as SHANK3 mutations are one of most replicated genetic defects identified from ASD patients. Using CRISPR/Cas9 gene editing, we successfully generated and characterized multiple lines of Beagle Shank3 (bShank3) mutants that have been propagated for a few generations. We developed and validated a battery of behavioral assays that can be used in controlled experimental setting for mutant dogs. bShank3 mutants exhibited distinct and robust social behavior deficits including social withdrawal and reduced social interactions with humans, and heightened anxiety in different experimental settings (n = 27 for wild-type controls and n = 44 for mutants). We demonstrate the feasibility of producing a large number of mutant animals in a reasonable time frame. The robust and unique behavioral findings support the validity and value of a canine model to investigate the pathophysiology and develop treatments for ASD and potentially other psychiatric disorders.
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Trastorno del Espectro Autista , Animales , Perros , Humanos , Trastorno del Espectro Autista/genética , Sistemas CRISPR-Cas/genética , Modelos Animales de Enfermedad , Edición Génica , Proteínas de Microfilamentos/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismoRESUMEN
Aspergillus flavus not only reduces kiwifruit production but also synthesizes carcinogenic aflatoxins, resulting in a relevant threat to human health. p-Hydroxybenzoic acid (pHBA) is one of the most abundant phenolics in kiwifruit. In this study, pHBA was found to reduce A. flavus mycelial growth by blocking the fungal mitotic exit network (MEN) and cytokinesis and to inhibit the biosynthesis of aflatoxins B1 and B2. The application of pHBA promoted the accumulation of endogenous pHBA and induced oxidative stress in A. flavus-infected kiwifruit, resulting in an increase in H2O2 content and catalase (CAT) and superoxide dismutase (SOD) activities. Preventive and curative treatments with 5 mM pHBA reduced A. flavus advancement by 46.1% and 68.0%, respectively. Collectively, the antifungal and elicitor properties of pHBA were examined for the first time, revealing new insights into the role of pHBA in the defense response of kiwifruit against A. flavus infection.
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Aflatoxinas , Aspergillus flavus , Humanos , Antifúngicos/farmacología , Peróxido de HidrógenoRESUMEN
Genomic imprinting disorders are caused by the disruption of genomic imprinting processes leading to a deficit or increase of an active allele. Their unique molecular mechanisms underlying imprinted genes offer an opportunity to investigate epigenetic-based therapy for reactivation of an inactive allele or reduction of an active allele. Current treatments are based on managing symptoms, not targeting the molecular mechanisms underlying imprinting disorders. Here, we highlight molecular approaches of therapeutic candidates in preclinical and clinical studies for individual imprinting disorders. These include the significant progress of discovery and testing of small molecules, antisense oligonucleotides, and CRISPR mediated genome editing approaches as new therapeutic strategies. We discuss the significant challenges of translating these promising therapies from the preclinical stage to the clinic, especially for genome editing based approaches.
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Edición Génica , Impresión Genómica , Impresión Genómica/genética , Metilación de ADNRESUMEN
BACKGROUND: p-Aminobenzoic acid (pABA) is an environmentally friendly bioactive metabolite synthesized by Lysobacter antibioticus. This compound showed an unusual antifungal mode of action based on cytokinesis inhibition. However, the potential antibacterial properties of pABA remain unexplored. RESULTS: In this study, pABA showed antibacterial activity against Gram-negative bacteria. This metabolite inhibited growth (EC50 = 4.02 mM), and reduced swimming motility, extracellular protease activity, and biofilm formation in the soybean pathogen Xanthomonas axonopodis pv. glycines (Xag). Although pABA was previously reported to inhibit fungal cell division, no apparent effect was observed on Xag cell division genes. Instead, pABA reduced the expression of various membrane integrity-related genes, such as cirA, czcA, czcB, emrE, and tolC. Consistently, scanning electron microscopy observations revealed that pABA caused major alternations in Xag morphology and blocked the formation of bacterial consortiums. In addition, pABA reduced the content and profile of outer membrane proteins and lipopolysaccharides in Xag, which may explain the observed effects. Preventive and curative applications of 10 mM pABA reduced Xag symptoms in soybean plants by 52.1% and 75.2%, respectively. CONCLUSIONS: The antibacterial properties of pABA were studied for the first time, revealing new insights into its potential application for the management of bacterial pathogens. Although pABA was previously reported to show an antifungal mode of action based on cytokinesis inhibition, this compound inhibited Xag growth by altering the outer membrane's integrity. © 2023 Society of Chemical Industry.
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Fabaceae , Xanthomonas axonopodis , Xanthomonas , Glycine max/microbiología , Xanthomonas axonopodis/genética , Xanthomonas axonopodis/metabolismo , Ácido 4-Aminobenzoico/farmacología , Ácido 4-Aminobenzoico/química , Ácido 4-Aminobenzoico/metabolismo , Antifúngicos/farmacología , Antifúngicos/metabolismo , Glicina/metabolismo , Antibacterianos/farmacología , Enfermedades de las Plantas/microbiología , Xanthomonas/metabolismoRESUMEN
Anxiety disorders are the most prevalent co-morbidity factor associated with the core domains of autism spectrum disorders (ASD). Investigations on potential common neuronal mechanisms that may explain the co-occurrence of ASD and anxiety disorders are still poorly explored. One of the key questions that remained unsolved is the role of Shank3 protein in anxiety behaviours. Firstly, we characterize the developmental trajectories of locomotor, social behaviour and anxiety traits in a mouse model of ASD. We highlight that the anxiety phenotype is a late-onset emerging phenotype in mice with a Shank3Δe4-22 mutation. Consequently, we used an shRNA strategy to model Shank3 insufficiency in the bed nucleus of the stria terminalis (BNST), a brain region exerting a powerful control on anxiety level. We found that Shank3 downregulation in the anteromedial BNST (amBNST) induced anxiogenic effects and enhanced social avoidance after aversive social defeat. Associated with these behavioural defects, we showed alteration of glutamatergic synaptic functions in the amBNST induced by Shank3 insufficiency during adolescence. Our data strongly support the role of Shank3 in the maturation of amBNST, and its key role in anxiety control. Our results may further help to pave the road on a better understanding of the neuronal mechanisms underlying anxiety disorders implicated in ASDs.
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Núcleos Septales , Ratones , Animales , Núcleos Septales/metabolismo , Conducta Social , Ansiedad/metabolismo , Trastornos de Ansiedad/metabolismo , Fenotipo , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismoRESUMEN
BACKGROUND: Kiwifruit is highly susceptible to fungal pathogens, such as Botrytis cinerea, which reduce crop production and quality. In this study, dipicolinic acid (DPA), which is one of the main components of Bacillus spores, was evaluated as a new elicitor to enhance kiwifruit resistance to B. cinerea. RESULTS: DPA enhances antioxidant capacity and induces the accumulation of phenolics in B. cinerea-infected 'Xuxiang' kiwifruit. The contents of the main antifungal phenolics in kiwifruit, including caffeic acid, chlorogenic acid and isoferulic acid, increased after DPA treatment. DPA enhanced H2 O2 levels after 0 and 1 days, which promoted catalase (CAT) and superoxide dismutase (SOD) activities, reducing long-term H2 O2 levels. DPA promoted the up-regulation of several kiwifruit defense genes, including CERK1, MPK3, PR1-1, PR1-2, PR5-1 and PR5-2. Furthermore, DPA at 5 mM inhibited B. cinerea symptoms in kiwifruit (95.1% lesion length inhibition) more effectively than the commercial fungicides carbendazim, difenoconazole, prochloraz and thiram. CONCLUSIONS: The antioxidant properties of DPA and the main antifungal phenolics of kiwifruit were examined for the first time. This study uncovers new insights regarding the potential mechanisms used by Bacillus species to induce disease resistance. © 2023 Society of Chemical Industry.