RESUMEN
Faithful chromosome segregation is required for both mitotic and meiotic cell divisions and is regulated by multiple mechanisms including the anaphase-promoting complex/cyclosome (APC/C), which is the largest known E3 ubiquitin-ligase complex and has been implicated in regulating chromosome segregation in both mitosis and meiosis in animals. However, the role of the APC/C during plant meiosis remains largely unknown. Here, we show that Arabidopsis APC8 is required for male meiosis. We used a combination of genetic analyses, cytology and immunolocalisation to define the function of AtAPC8 in male meiosis. Meiocytes from apc8-1 plants exhibit several meiotic defects including improper alignment of bivalents at metaphase I, unequal chromosome segregation during anaphase II, and subsequent formation of polyads. Immunolocalisation using an antitubulin antibody showed that APC8 is required for normal spindle morphology. We also observed mitotic defects in apc8-1, including abnormal sister chromatid segregation and microtubule morphology. Our results demonstrate that Arabidopsis APC/C is required for meiotic chromosome segregation and that APC/C-mediated regulation of meiotic chromosome segregation is a conserved mechanism among eukaryotes.
Asunto(s)
Ciclosoma-Complejo Promotor de la Anafase/metabolismo , Subunidad Apc8 del Ciclosoma-Complejo Promotor de la Anafase/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/citología , Arabidopsis/metabolismo , Meiosis , Subunidad Apc8 del Ciclosoma-Complejo Promotor de la Anafase/genética , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Ciclo Celular/metabolismo , Cromátides/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Segregación Cromosómica , Cromosomas de las Plantas/genética , Secuencia Conservada , Variación Genética , Cinetocoros/metabolismo , Microtúbulos/metabolismo , Mitosis , Modelos Biológicos , Fenotipo , Mutación Puntual/genética , Huso Acromático/metabolismo , CohesinasRESUMEN
Cell division requires proper spindle assembly; a surveillance pathway, the spindle assembly checkpoint (SAC), monitors whether the spindle is normal and correctly attached to kinetochores. The SAC proteins regulate mitotic chromosome segregation by affecting CDC20 (Cell Division Cycle 20) function. However, it is unclear whether CDC20 regulates meiotic spindle assembly and proper homolog segregation. Here, we show that the Arabidopsis thaliana CDC20.1 gene is indispensable for meiosis and male fertility. We demonstrate that cdc20.1 meiotic chromosomes align asynchronously and segregate unequally and the metaphase I spindle has aberrant morphology. Comparison of the distribution of meiotic stages at different time points between the wild type and cdc20.1 reveals a delay of meiotic progression from diakinesis to anaphase I. Furthermore, cdc20.1 meiocytes exhibit an abnormal distribution of a histone H3 phosphorylation mark mediated by the Aurora kinase, providing evidence that CDC20.1 regulates Aurora localization for meiotic chromosome segregation. Further evidence that CDC20.1 and Aurora are functionally related was provided by meiosis-specific knockdown of At-Aurora1 expression, resulting in meiotic chromosome segregation defects similar to those of cdc20.1. Taken together, these results suggest a critical role for CDC20.1 in SAC-dependent meiotic chromosome segregation.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Proteínas Cdc20/genética , Puntos de Control del Ciclo Celular , Segregación Cromosómica , Meiosis , Anafase , Ciclosoma-Complejo Promotor de la Anafase/genética , Ciclosoma-Complejo Promotor de la Anafase/metabolismo , Arabidopsis/citología , Arabidopsis/fisiología , Proteínas de Arabidopsis/metabolismo , Proteínas Cdc20/metabolismo , Técnicas de Silenciamiento del Gen , Cinetocoros/metabolismo , Metafase , Infertilidad Vegetal , Huso Acromático/metabolismoRESUMEN
Cancer stem cells (CSCs) are known to contribute to the progression of colorectal cancer (CRC). However, understanding of the molecular mechanisms and key factors involved in CRC is still insufficient to identify therapeutic targets against colorectal CSCs. In an effort to identify such mechanisms, we conducted bioinformatics analyses to evaluate the expression patterns in tumor and normal colorectal tissues, leading us to focus on the role of the ZNF217/Notch1 axis in mediating stem cell properties in CRC. Our findings revealed that ZNF217 overexpression activated self-renewal ability, expression of colorectal CSC markers, and Notch signaling in CRC. Dual-luciferase reporter assay suggested a role for ZNF217 in targeting Notch1 to activate Notch signaling. We observed that the promotional effects of Notch signaling, as well as CSC markers, under ZNF217 overexpression were attenuated after Notch1 knockdown. In addition to in vitro data, our in vivo results confirmed the inhibitory effect of sulforaphane on the tumorigenicity of CSCs, depicted the suppressive role of sulforaphane on colorectal CSCs mediated by the ZNF217/Notch1 axis, thereby providing new targetable vulnerabilities and therapeutic strategies for CRC.