RESUMEN
Monopolar spindle 1 (MPS1), which plays a critical role in somatic mitosis, has also been revealed to be essential for meiosis I in oocytes. Spermatogenesis is an important process involving successive mitosis and meiosis, but the function of MPS1 in spermatogenesis remains unclear. Here, we generated Mps1 conditional knockout mice and found that Ddx4-cre-driven loss of Mps1 in male mice resulted in depletion of undifferentiated spermatogonial cells and subsequently of differentiated spermatogonia and spermatocytes. In addition, Stra8-cre-driven ablation of Mps1 in male mice led to germ cell loss and fertility reduction. Spermatocytes lacking Mps1 have blocked at the zygotene-to-pachytene transition in the prophase of meiosis I, which may be due to decreased H2B ubiquitination level mediated by MDM2. And the expression of many meiotic genes was decreased, while that of apoptotic genes was increased. Moreover, we also detected increased apoptosis in spermatocytes with Mps1 knockout, which may have been the reason why germ cells were lost. Taken together, our findings indicate that MPS1 is required for mitosis of gonocytes and spermatogonia, differentiation of undifferentiated spermatogonia, and progression of meiosis I in spermatocytes.
Asunto(s)
Fertilidad/genética , Proteínas Serina-Treonina Quinasas/fisiología , Espermatogénesis/genética , Animales , Infertilidad Masculina/genética , Masculino , Meiosis/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitosis/genética , Proteínas Serina-Treonina Quinasas/genética , Espermatocitos/fisiología , Espermatogonias/fisiologíaRESUMEN
Gene mutations play important roles in tumour development. In this study, we identified a functional histone H2B mutation H2BL-T11C, causing an amino acid variation from Leu to Pro (L3P, H2BL-L3P). Cells overexpressing H2BL-L3P showed stronger proliferation, colony formation, tumourigenic abilities, and a different cell cycle distribution. Meanwhile, the c-Myc expression was elevated as evident by RNA-seq. We further revealed that an H2BK5ac-H2BK120ubi crosstalk which regulates gene transcription. Moreover, EdU staining demonstrated an important role of c-Myc in accelerating cell cycle progression through the G1/S checkpoint, while treatment with 10058-F4, an inhibitor of the c-Myc/MAX interaction, alleviated the abnormal cell proliferation and cell cycle distribution in vitro and partially inhibited tumour growth in vivo. The mutation of amino acid L3P is associated with tumour progression, suggesting patients carrying this SNP may have higher risk of tumour development.
Asunto(s)
Proliferación Celular/fisiología , Variación Genética/genética , Histonas/genética , Neoplasias/genética , Proteínas Proto-Oncogénicas c-myc/genética , Regulación hacia Arriba/fisiología , Animales , Línea Celular Tumoral , Células HEK293 , Histonas/metabolismo , Humanos , Lentivirus , Leucina/genética , Ratones , Ratones Desnudos , Mutación/genética , Neoplasias/metabolismo , Neoplasias/patología , Nucleótidos/genética , Prolina/genética , Proteínas Proto-Oncogénicas c-myc/biosíntesis , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Cancer cells reprogram their energy metabolic system from the mitochondrial oxidative phosphorylation (OXPHOS) pathway to a glucose-dependent aerobic glycolysis pathway. This metabolic reprogramming phenomenon is known as the Warburg effect, a significant hallmark of cancer. However, the detailed mechanisms underlying this event or triggering this reprogramming remain largely unclear. Here, we found that histone H2B monoubiquitination (H2Bub1) negatively regulates the Warburg effect and tumorigenesis in human lung cancer cells (H1299 and A549 cell lines) likely through controlling the expression of multiple mitochondrial respiratory genes, which are essential for OXPHOS. Moreover, our work also suggested that pyruvate kinase M2 (PKM2), the rate-limiting enzyme of glycolysis, can directly interact with H2B in vivo and in vitro and negatively regulate the level of H2Bub1. The inhibition of cell proliferation and nude mice xenograft of human lung cancer cells induced by PKM2 knockdown can be partially rescued through lowering H2Bub1 levels, which indicates that the oncogenic function of PKM2 is achieved, at least partially, through the control of H2Bub1. Furthermore, PKM2 and H2Bub1 levels are negatively correlated in cancer specimens. Therefore, these findings not only provide a novel mechanism triggering the Warburg effect that is mediated through an epigenetic pathway (H2Bub1) but also reveal a novel metabolic regulator (PKM2) for the epigenetic mark H2Bub1. Thus, the PKM2-H2Bub1 axis may become a promising cancer therapeutic target.
Asunto(s)
Epigénesis Genética , Histonas/metabolismo , Ubiquitinación , Efecto Warburg en Oncología , Animales , Carcinogénesis/genética , Carcinogénesis/patología , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Respiración de la Célula/genética , Células HEK293 , Humanos , Masculino , Proteínas de la Membrana/metabolismo , Ratones Endogámicos BALB C , Ratones Desnudos , Mitocondrias/genética , Simulación del Acoplamiento Molecular , Unión Proteica , Hormonas Tiroideas/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas de Unión a Hormona TiroideRESUMEN
BACKGROUND: Histone modifications are critical in regulating neuronal processes. However, the impacts of individual histone modifications on learning and memory are elusive. Here, we investigated the contributions of histone H3 lysine modifications to learning and memory in Drosophila by using histone lysine-to-alanine mutants. RESULTS: Behavioural analysis indicated that compared to the H3WT group, mutants overexpressing H3K23A displayed impaired courtship learning. Chromatin immunoprecipitation analysis of H3K23A mutants showed that H3K23 acetylation (H3K23ac) levels were decreased on learning-related genes. Knockdown of CREB-binding protein (CBP) decreased H3K23ac levels, attenuated the expression of learning-related genes, led to a courtship learning defect and altered development of the mushroom bodies. A decline in courtship learning ability was observed in both larvae and adult treatments with ICG-001. Furthermore, treatment of Drosophila overexpressing mutated H3K23A with a CBP inhibitor did not aggravate the learning defect. CONCLUSIONS: H3K23ac, catalysed by the acetyltransferases dCBP, contributes to Drosophila learning, likely by controlling the expression of specific genes. This is a novel epigenetic regulatory mechanism underlying neuronal behaviours.
Asunto(s)
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Histonas/metabolismo , Lisina/metabolismo , Factores de Transcripción p300-CBP/metabolismo , Acetilación , Animales , Cortejo , Femenino , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Histonas/genética , Aprendizaje , Masculino , Mutación , Neuronas/metabolismoRESUMEN
Dysregulation of the homeostatic balance of histone H3 di- and tri-methyl lysine 27 (H3K27me2/3) levels caused by the mis-sense mutation of histone H3 (H3K27M) is reported to be associated with various types of cancers. In this study, we found that reduction in H3K27me2/3 caused by H3.1K27M, a mutation of H3 variants found in patients with diffuse intrinsic pontine glioma (DIPG), dramatically attenuated the presence of 53BP1 (also known as TP53BP1) foci and the capability of non-homologous end joining (NHEJ) in human dermal fibroblasts. H3.1K27M mutant cells showed increased rates of genomic insertions/deletions and copy number variations, as well as an increase in p53-dependent apoptosis. We further showed that both hypo-H3K27me2/3 and H3.1K27M interacted with FANCD2, a central player in the choice of DNA repair pathway. H3.1K27M triggered the accumulation of FANCD2 on chromatin, suggesting an interaction between H3.1K27M and FANCD2. Interestingly, knockdown of FANCD2 in H3.1K27M cells recovered the number of 53BP1-positive foci, NHEJ efficiency and apoptosis rate. Although these findings in HDF cells may differ from the endogenous regulation of the H3.1K27M mutant in the specific tumor context of DIPG, our results suggest a new model by which H3K27me2/3 facilitates NHEJ and the maintenance of genome stability.This article has an associated First Person interview with the first author of the paper.
Asunto(s)
Cromatina/metabolismo , Reparación del ADN por Unión de Extremidades , Enzimas Reparadoras del ADN/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/metabolismo , Histonas/metabolismo , Neoplasias del Tronco Encefálico/genética , Neoplasias del Tronco Encefálico/metabolismo , Línea Celular , Cromatina/genética , Reparación del ADN , Enzimas Reparadoras del ADN/genética , Proteínas de Unión al ADN/genética , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Fibroblastos , Inestabilidad Genómica , Glioma/genética , Glioma/metabolismo , Células HEK293 , Histonas/genética , Humanos , Metilación , Proteína 1 de Unión al Supresor Tumoral P53/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismoRESUMEN
Autophagy is an evolutionarily conserved cellular process that primarily participates in lysosome-mediated protein degradation. Although autophagy is a cytoplasmic event, how epigenetic pathways are involved in the regulation of autophagy remains incompletely understood. Here, we found that H2B monoubiquitination (H2Bub1) is down-regulated in cells under starvation conditions and that the decrease in H2Bub1 results in the activation of autophagy. We also identified that the deubiquitinase USP44 is responsible for the starvation-induced decrease in H2Bub1. Furthermore, the changes in H2Bub1 affect the transcription of genes involved in the regulation of autophagy. Therefore, this study reveals a novel epigenetic pathway for the regulation of autophagy through H2Bub1.
Asunto(s)
Autofagia/genética , Epigénesis Genética , Histonas/metabolismo , Ubiquitinación/genética , Animales , Proteínas Relacionadas con la Autofagia/antagonistas & inhibidores , Proteínas Relacionadas con la Autofagia/genética , Proteínas Relacionadas con la Autofagia/metabolismo , Diferenciación Celular , ADN (Citosina-5-)-Metiltransferasas/metabolismo , ADN Metiltransferasa 3A , Regulación hacia Abajo , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Endopeptidasas/genética , Endopeptidasas/metabolismo , Técnicas de Silenciamiento del Gen , Células HEK293 , Células HeLa , Histonas/química , Histonas/genética , Humanos , Ratones , Modelos Biológicos , ARN Interferente Pequeño/genética , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Ubiquitina Tiolesterasa , Proteasas Ubiquitina-Específicas/antagonistas & inhibidores , Proteasas Ubiquitina-Específicas/genética , Proteasas Ubiquitina-Específicas/metabolismo , ADN Metiltransferasa 3BRESUMEN
Epigenetics plays critical roles in controlling stem cell self-renewal and differentiation. Histone H1 is one of the most critical chromatin regulators, but its role in adult stem cell regulation remains unclear. Here we report that H1 is intrinsically required in the regulation of germline stem cells (GSCs) in the Drosophila ovary. The loss of H1 from GSCs causes their premature differentiation through activation of the key GSC differentiation factor bam. Interestingly, the acetylated H4 lysine 16 (H4K16ac) is selectively augmented in the H1-depleted GSCs. Furthermore, overexpression of mof reduces H1 association on chromatin. In contrast, the knocking down of mof significantly rescues the GSC loss phenotype. Taken together, these results suggest that H1 functions intrinsically to promote GSC self-renewal by antagonizing MOF function. Since H1 and H4K16 acetylation are highly conserved from fly to human, the findings from this study might be applicable to stem cells in other systems.
Asunto(s)
Autorrenovación de las Células , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Células Germinativas/metabolismo , Histonas/metabolismo , Secuencias de Aminoácidos , Animales , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/química , Drosophila melanogaster/genética , Epigénesis Genética , Femenino , Células Germinativas/citología , Histonas/química , Histonas/genética , Masculino , Ovario/química , Ovario/metabolismoRESUMEN
Chromatin is a highly organized and dynamic structure in eukaryotic cells. The change of chromatin structure is essential in many cellular processes, such as gene transcription, DNA damage repair and others. Anti-silencing function 1 (ASF1) is a histone chaperone that participates in chromatin higher-order organization and is required for appropriate chromatin assembly. In this study, we identified the E2 ubiquitin-conjugating enzyme RAD6 as an evolutionary conserved interacting protein of ASF1 in D. melanogaster and H. sapiens that promotes the turnover of ASF1A by cooperating with a well-known E3 ligase, MDM2, via ubiquitin-proteasome pathway in H. sapiens. Further functional analyses indicated that the interplay between RAD6 and ASF1A associates with tumorigenesis. Together, these data suggest that the RAD6-MDM2 ubiquitin ligase machinery is critical for the degradation of chromatin-related proteins.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Enzimas Ubiquitina-Conjugadoras/metabolismo , Animales , Western Blotting , Proteínas de Ciclo Celular/genética , Línea Celular , Línea Celular Tumoral , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Regulación Neoplásica de la Expresión Génica , Células HEK293 , Células Hep G2 , Histonas/metabolismo , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Microscopía Confocal , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Proteínas Proto-Oncogénicas c-mdm2/genética , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Enzimas Ubiquitina-Conjugadoras/genética , UbiquitinaciónRESUMEN
Histone H2B monoubiquitination is a key histone modification that has significant effects on chromatin higher-order structure and gene transcription. Multiple biological processes have been suggested to be tightly related to the dynamics of H2B monoubiquitination. However, a comprehensive understanding of biological roles of H2B monoubiquitination is still poorly understood. In the present study, we developed an efficient tool to disrupt endogenous H2B monoubiquitination levels by using an H2BK120R mutant construct expressed in human cells. Genome-wide microarray analysis of these cells revealed a potential global view of biological functions of H2B monoubiquitination. Bioinformatics analysis of our data demonstrated that while H2B monoubiquitination expectedly affected a number of previously reported biological pathways, we also uncovered the influence of this histone modification on many novel biological processes. Therefore, our work provided valuable information for understanding the role of H2B monoubiquitination and indicated potential directions for its further studies.
Asunto(s)
Histonas/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Ubiquitinación , Animales , Diferenciación Celular/genética , Cromatina/metabolismo , Daño del ADN/genética , Reparación del ADN/genética , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HEK293 , Células HeLa , Histonas/genética , Humanos , Ratones , Células Madre Embrionarias de Ratones/citología , Proteínas Mutantes/metabolismo , Mutación/genética , Ubiquitina/metabolismoRESUMEN
This article has been withdrawn by the authors.
RESUMEN
Efficient DNA double-strand break (DSB) repair is critical for the maintenance of genome stability. Unrepaired or misrepaired DSBs cause chromosomal rearrangements that can result in severe consequences, such as tumorigenesis. RAD6 is an E2 ubiquitin-conjugating enzyme that plays a pivotal role in repairing UV-induced DNA damage. Here, we present evidence that RAD6 is also required for DNA DSB repair via homologous recombination (HR) by specifically regulating the degradation of heterochromatin protein 1α (HP1α). Our study indicates that RAD6 physically interacts with HP1α and ubiquitinates HP1α at residue K154, thereby promoting HP1α degradation through the autophagy pathway and eventually leading to an open chromatin structure that facilitates efficient HR DSB repair. Furthermore, bioinformatics studies have indicated that the expression of RAD6 and HP1α exhibits an inverse relationship and correlates with the survival rate of patients.
Asunto(s)
Autofagia/genética , Proteínas Cromosómicas no Histona/metabolismo , Heterocromatina/metabolismo , Reparación del ADN por Recombinación/genética , Enzimas Ubiquitina-Conjugadoras/metabolismo , Línea Celular , Homólogo de la Proteína Chromobox 5 , HumanosRESUMEN
Protein ubiquitinylation regulates protein stability and activity. RAD6, an E2 ubiquitin-conjugating enzyme, which that has been substantially biochemically characterized, functions in a number of biologically relevant pathways, including cell cycle progression. In this study, we show that RAD6 promotes the G1-S transition and cell proliferation by regulating the expression of cyclin D1 (CCND1) in human cells. Furthermore, our data indicate that RAD6 influences the transcription of CCND1 by increasing monoubiquitinylation of histone H2B and trimethylation of H3K4 in the CCND1 promoter region. Our study presents, for the first time, an evidence for the function of RAD6 in cell cycle progression and cell proliferation in human cells, raising the possibility that RAD6 could be a new target for molecular diagnosis and prognosis in cancer therapeutics.
Asunto(s)
Ciclina D1/metabolismo , Enzimas Ubiquitina-Conjugadoras/metabolismo , Línea Celular , Proliferación Celular , Ciclina D1/genética , Fase G1 , Células HeLa , Histonas/metabolismo , Humanos , Regiones Promotoras Genéticas , Interferencia de ARN , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Fase S , Transcripción Genética , Enzimas Ubiquitina-Conjugadoras/antagonistas & inhibidores , Enzimas Ubiquitina-Conjugadoras/genética , Ubiquitinación , Regulación hacia ArribaRESUMEN
OBJECTIVES: To test the serum sexual hormone and sperm cytology in aspermia patients. METHODS: Radioimmunoassay (RIA) and Wright-Giemsa stain were used to detect serum sexual hormone and sperm cytology in 45 aspermia patients. RESULTS: Among the 45 patients, spermatogenetic cells were detected in 16 patients(35.6%). The level of serum testerone were decreased in 16 patients, and FSH, LH were increased in 13 cases as well as that of PRL in 4 cases. CONCLUSIONS: These results suggested that the testis function was demaged, the ratio of T/LH could further reflect the function of Leydig cells, the detection of PRL were significant only in diagnosis of aspermia induced by hypermia. It is an important index in identifying obstructive and non-obstructive aspermia and has important significance in estimating the extention of testis demage and commanding the clinical treatment of serum hormone detection and sperm cell analysis.
