RESUMEN
Presynaptic syntaxin binding protein 1 (STXBP1) is essential for neurotransmitter release. Heterozygous mutations in this protein cause STXBP1 encephalopathy (STXBP1-E), which is characterized by intellectual disabilities and epilepsies. Since nonhuman primates closely resemble humans, monkey models may advance studies on the pathogenesis and therapeutic treatments of STXBP1-E. We generated cynomolgus monkeys carrying STXBP1 (R292H) mutation through base editing of in vitro fertilized embryos to mimic a clinical condition. The newborn STXBP1-edited monkeys exhibited focal epilepsy, and the animal that survived beyond the first week postpartum presented typical EEG phenotypes. Biochemical analysis of brain biopsy samples showed reduced levels of STXBP1 (MUNC18-1) and SNARE complex proteins. Single-cell sequencing identified one specific cell cluster that may contribute to encephalopathy. Thus, our case report shows that base-edited STXBP1 mutant monkeys are a good animal model for STXBP1-E, and that a base-editing approach is useful for generating primate models of human genetic disorders.
Asunto(s)
Encefalopatías , Epilepsia , Animales , Encéfalo/metabolismo , Epilepsia/tratamiento farmacológico , Epilepsia/genética , Femenino , Macaca fascicularis/metabolismo , Proteínas Munc18/genética , Proteínas Munc18/metabolismo , MutaciónRESUMEN
INTRODUCTION: Extrauterine endometrial stromal sarcoma (EESS) arising from Deep pelvic endometriosis (DPE) has a poor life quality and is difficult to diagnose pre-operatively. However, the patient's quality of life can be improved when it is diagnosed precisely and managed successfully. CASE REPORT: A 35-year-old woman presented to our hospital with a 3-month history of hematochezia and anal pain. Initially, she was misdiagnosed as a rectal stromal tumor and then precisely diagnosed as having EESS from DPE following several multidisciplinary team (MDT) meetings. The lesion was shrunk by gonadotrophin-releasing hormone agonist (GnRH-α) treatment and then resected with minimal trauma. CONCLUSION: MDT is crucial in the treatment of the patient. It can promote individualized treatment and improve patient's quality of life.
Asunto(s)
Neoplasias Endometriales , Endometriosis , Sarcoma Estromático Endometrial , Femenino , Humanos , Adulto , Sarcoma Estromático Endometrial/diagnóstico por imagen , Sarcoma Estromático Endometrial/cirugía , Neoplasias Endometriales/diagnóstico por imagen , Neoplasias Endometriales/cirugía , Endometriosis/complicaciones , Endometriosis/diagnóstico por imagen , Endometriosis/patología , Calidad de Vida , Grupo de Atención al PacienteRESUMEN
The advent of base editors (BEs) holds great potential for correcting pathogenic-related point mutations to treat relevant diseases. However, Cas9 nickase (nCas9)-derived BEs lead to DNA double-strand breaks, which can trigger unwanted DNA damage response (DDR). Here, we show that the original version of catalytically dead Cas12a (dCas12a)-conjugated BEs induce a basal level of DNA breaks and minimally activate DDR proteins, including H2AX, ATM, ATR, and p53. By fusing dCas12a with engineered human apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3A (APOBEC3A), we further develop the BEACON (base editing induced by human APOBEC3A and Cas12a without DNA break) system to achieve enhanced deamination efficiency and editing specificity. Efficient C-to-T editing is achieved by BEACON in mammalian cells at levels comparable to AncBE4max, with only low levels of DDR and minimal RNA off-target mutations. Importantly, BEACON induces in vivo base editing in mouse embryos, and targeted C-to-T conversions are detected in F0 mice.
Asunto(s)
Proteínas Bacterianas/metabolismo , Proteínas Asociadas a CRISPR/metabolismo , Daño del ADN , Endodesoxirribonucleasas/metabolismo , Edición Génica/métodos , 17-Hidroxiesteroide Deshidrogenasas/genética , Animales , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Proteínas Bacterianas/genética , Proteínas Asociadas a CRISPR/genética , Citidina/metabolismo , Citidina Desaminasa/genética , Citidina Desaminasa/metabolismo , Replicación del ADN , Desaminación , Endodesoxirribonucleasas/genética , Femenino , Células HEK293 , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Fosforilación , Proteínas/genética , Proteínas/metabolismo , Timidina/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Ubiquitinas/metabolismoRESUMEN
The cerebellum is critical for controlling motor and non-motor functions via cerebellar circuit that is composed of defined cell types, which approximately account for more than half of neurons in mammals. The molecular mechanisms controlling developmental progression and maturation processes of various cerebellar cell types need systematic investigation. Here, we analyzed transcriptome profiles of 21119 single cells of the postnatal mouse cerebellum and identified eight main cell clusters. Functional annotation of differentially expressed genes revealed trajectory hierarchies of granule cells (GCs) at various states and implied roles of mitochondrion and ATPases in the maturation of Purkinje cells (PCs), the sole output cells of the cerebellar cortex. Furthermore, we analyzed gene expression patterns and co-expression networks of 28 ataxia risk genes, and found that most of them are related with biological process of mitochondrion and around half of them are enriched in PCs. Our results also suggested core transcription factors that are correlated with interneuron differentiation and characteristics for the expression of secretory proteins in glia cells, which may participate in neuronal modulation. Thus, this study presents a systematic landscape of cerebellar gene expression in defined cell types and a general gene expression framework for cerebellar development and dysfunction.