RESUMEN
The timely degradation of proteins that regulate the cell cycle is essential for oocyte maturation. Oocytes are equipped to degrade proteins via the ubiquitin-proteasome system. In meiosis, anaphase promoting complex/cyclosome (APC/C), an E3 ubiquitin-ligase, is responsible for the degradation of proteins. Ubiquitin-conjugating enzyme E2 S (UBE2S), an E2 ubiquitin-conjugating enzyme, delivers ubiquitin to APC/C. APC/C has been extensively studied, but the functions of UBE2S in oocyte maturation and mouse fertility are not clear. In this study, we used Ube2s knockout mice to explore the role of UBE2S in mouse oocytes. Ube2s-deleted oocytes were characterized by meiosis I arrest with normal spindle assembly and spindle assembly checkpoint dynamics. However, the absence of UBE2S affected the activity of APC/C. Cyclin B1 and securin are two substrates of APC/C, and their levels were consistently high, resulting in the failure of homologous chromosome separation. Unexpectedly, the oocytes arrested in meiosis I could be fertilized and the embryos could become implanted normally, but died before embryonic day 10.5. In conclusion, our findings reveal an indispensable regulatory role of UBE2S in mouse oocyte meiosis and female fertility.
Asunto(s)
Puntos de Control de la Fase M del Ciclo Celular , Meiosis , Animales , Femenino , Ratones , Ciclosoma-Complejo Promotor de la Anafase/genética , Ciclosoma-Complejo Promotor de la Anafase/metabolismo , Oocitos/metabolismo , Ubiquitinas/metabolismoRESUMEN
BACKGROUND: Members of the nucleotide-binding oligomerization domain, leucine rich repeat and pyrin domain containing (NLRP) family regulate various physiological and pathological processes. However, none have been shown to regulate actin cap formation or spindle translocation during the asymmetric division of oocyte meiosis I. NLRP4E has been reported as a candidate protein in female fertility, but its function is unknown. METHODS: Immunofluorescence, reverse transcription polymerase chain reaction (RT-PCR), and western blotting were employed to examine the localization and expression levels of NLRP4E and related proteins in mouse oocytes. small interfering RNA (siRNA) and antibody transfection were used to knock down NLRP4E and other proteins. Immunoprecipitation (IP)-mass spectrometry was used to identify the potential proteins interacting with NLRP4E. Coimmunoprecipitation (Co-IP) was used to verify the protein interactions. Wild type (WT) or mutant NLRP4E messenger RNA (mRNA) was injected into oocytes for rescue experiments. In vitro phosphorylation was employed to examine the activation of steroid receptor coactivator (SRC) by NLRP4E. RESULTS: NLRP4E was more predominant within oocytes compared with other NLRP4 members. NLRP4E knockdown significantly inhibited actin cap formation and spindle translocation toward the cap region, resulting in the failure of polar body extrusion at the end of meiosis I. Mechanistically, GRIN1, and GANO1 activated NLRP4E by phosphorylation at Ser429 and Thr430; p-NLRP4E is translocated and is accumulated in the actin cap region during spindle translocation. Next, we found that p-NLRP4E directly phosphorylated SRC at Tyr418, while p-SRC negatively regulated p-CDC42-S71, an inactive form of CDC42 that promotes actin cap formation and spindle translocation in the GTP-bound form. CONCLUSIONS: NLRP4E activated by GRIN1 and GANO1 regulates actin cap formation and spindle translocation toward the cap region through upregulation of p-SRC-Tyr418 and downregulation of p-CDC42-S71 during meiosis I.
Asunto(s)
Actinas , Meiosis , Oocitos , Proteína de Unión al GTP cdc42 , Animales , Oocitos/metabolismo , Ratones , Femenino , Actinas/metabolismo , Actinas/genética , Proteína de Unión al GTP cdc42/metabolismo , Proteína de Unión al GTP cdc42/genética , Fosforilación , Huso Acromático/metabolismoRESUMEN
microRNA (miR)-141-3p has context-dependent effects on tumor progression. In this study, we attempted to explore the expression and function of miR-141-3p in cervical cancer. We found that miR-141-3p expression was significantly increased in cervical cancer specimens relative to normal cervical tissues. Moreover, miR-141-3p levels were associated with tumor size and lymph node metastasis status. Ectopic expression of miR-141-3p significantly increased cervical cancer cell proliferation, colony formation, invasion, and epithelial to mesenchymal transition, whereas depletion of miR-141-3p suppressed cervical cancer cell proliferation and invasion. FOXA2 was identified to be a target of miR-141-3p. Overexpression of miR-141-3p led to a marked inhibition of endogenous FOXA2 in cervical cancer cells. FOXA2 silencing phenocopied the effects of miR-141-3p overexpression on cervical cancer cell proliferation and invasion. Enforced expression of FOXA2 blocked the effects of miR-141-3p on cervical cancer cell proliferation and invasion. miR-141-3p overexpression significantly accelerated the growth of xenograft tumors, which was accompanied by a striking reduction in FOXA2 expression. miR-141-3p acts as an oncogene in cervical cancer largely through repression of FOXA2. Targeting miR-141-3p may represent a potential therapeutic strategy for cervical cancer.
Asunto(s)
Carcinogénesis/genética , Factor Nuclear 3-beta del Hepatocito/genética , MicroARNs/genética , Neoplasias del Cuello Uterino/genética , Adulto , Anciano , Animales , Línea Celular Tumoral , Proliferación Celular/genética , Regulación hacia Abajo , Transición Epitelial-Mesenquimal/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Ratones Endogámicos BALB C , Persona de Mediana Edad , Invasividad Neoplásica/genética , Regulación hacia Arriba , Neoplasias del Cuello Uterino/patologíaRESUMEN
To investigate the influences of fresh embryo transfer (ET) and frozen embryo transfer (FET) on neonatal birthweight and the expression of imprinted genes PEG10 and L3MBTL1 in the placenta after in vitro fertilization-embryo transfer (IVF-ET), we analyzed the neonatal birthweight between fresh ET and FET transfer cycles. Then, we collected placentas delivered by fresh ET and FET, and real-time quantitative PCR, Western blotting and immunohistochemistry were used to detect the expression of PEG10 and L3MBTL1. The mean neonatal birthweight of fresh ET was lower than that of FET(3348.48 ± 521.05 vs. 3450.34 ± 524.13, P < 0.001). The risks of low birthweight (LBW) and small-for-gestational age (SGA) were lower after FET (3.9 % vs. 5.4 %; 7.2 % vs. 10.3 %), with adjusted rate ratios of 0.74 (95 % CI, 0.59-0.93; P = 0.003) and 0.70 (95 % CI, 0.59-0.84; P < 0.001), respectively. FET resulted in higher frequencies of macrosomia and large-for-gestational age (LGA) (14.2 % vs. 10.3; 11.0 % vs. 7.1 %) than fresh ET, with adjusted rate ratios of 1.45 (95 % CI, 1.26-1.68; P < 0.001) and 1.62 (95 % CI, 1.37-1.91; P < 0.001), respectively. We also observed PEG10 mRNA and protein expression levels in placentas delivered by fresh ET and FET were significantly different, but there were no significant differences in L3MBTL1 between the two groups. Fresh ET may affect the expression of the imprinted gene PEG10 in the placenta and adverse to placental implantation and development, resulting to increasing incidences of LBW and SGA.
Asunto(s)
Criopreservación , Fertilización In Vitro , Proteínas Reguladoras de la Apoptosis , Peso al Nacer , Proteínas de Unión al ADN , Transferencia de Embrión , Femenino , Retardo del Crecimiento Fetal , Humanos , Recién Nacido , Placenta , Embarazo , Proteínas de Unión al ARN , Proteínas Represoras , Estudios Retrospectivos , Proteínas Supresoras de TumorRESUMEN
Immunoglobulins are key humoral immune molecules produced and secreted by B lymphocytes at various stages of differentiation. No research has reported whether immunoglobulins are present in the non-proliferative female germ cells-oocytes-and whether they are functionally important for oocyte quality, self-protection, and survival. Herein, we found that IgG was present in the oocytes of immunodeficient mice; the IgG-VDJ regions were highly variable between different oocytes, and H3K27Ac bound and regulated the IgG promoter region. Next, IgG mRNA and protein levels increased in response to LPS, and this increment was mediated by CR2 on the oocyte membrane. Finally, we revealed three aspects of the functional relevance of oocyte IgG: first, oocytes could upregulate IgG to counteract the increased ROS level induced by CSF1; second, oocytes could upregulate IgG in response to injected virus ssRNA to maintain mitochondrial integrity; third, upon bacterial infection, oocytes could secrete IgG, subsequently encompassing the bacteria, thus increasing survival compared to somatic cells. This study reveals for the first time that the female germ cells, oocytes, can independently adjust intrinsic IgG production to survive in adverse environments.
Asunto(s)
Células Germinativas , Oocitos , Femenino , Ratones , Animales , Oocitos/metabolismo , Diferenciación Celular , ARN Mensajero/metabolismo , Inmunoglobulina G/metabolismoRESUMEN
OBJECTIVE: To determine whether singleton pregnancy achieved after preimplantation genetic testing (PGT) is associated with a higher risk of adverse perinatal outcomes than in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) singleton pregnancy. DESIGN: A retrospective cohort study. SETTING: A university-affiliated fertility center. PATIENT(S): This cohort study included singleton live births resulting from PGT (n = 232) and IVF/ICSI singleton pregnancies (n = 2,829) with single frozen-thawed blastocyst transfer. Multiple baseline covariates were used for propensity score matching, yielding 214 PGT singleton pregnancies matched to 617 IVF/ICSI singleton pregnancies. INTERVENTION(S): Trophectoderm biopsy. MAIN OUTCOME MEASURE(S): The primary outcome was gestational hypertension, and various clinical perinatal secondary outcomes related to maternal and neonatal health were measured. RESULT(S): Compared with IVF/ICSI singleton pregnancy, PGT singleton pregnancy was associated with a significantly higher risk of gestational hypertension (adjusted odds ratio, 2.58; 95% confidence interval, 1.32, 5.05). In the matched sample, the risk of gestational hypertension remained higher with PGT singleton pregnancy (odds ratio, 2.33; 95% confidence interval, 1.04, 5.22) than with IVF/ICSI singleton pregnancy. No statistical differences were noted in any other measured outcomes between the groups. CONCLUSION(S): The perinatal outcomes of PGT and IVF/ICSI singleton pregnancies were similar except for the observed potentially higher risk of gestational hypertension with PGT singleton pregnancy. However, because the data on PGT singleton pregnancies are limited, this conclusion warrants further investigation.
Asunto(s)
Criopreservación/tendencias , Transferencia de Embrión/tendencias , Pruebas Genéticas/tendencias , Nacimiento Vivo/epidemiología , Diagnóstico Preimplantación/tendencias , Puntaje de Propensión , Adulto , Estudios de Cohortes , Criopreservación/métodos , Transferencia de Embrión/métodos , Femenino , Fertilización In Vitro , Congelación , Pruebas Genéticas/métodos , Humanos , Hipertensión Inducida en el Embarazo/diagnóstico , Hipertensión Inducida en el Embarazo/epidemiología , Recién Nacido , Masculino , Embarazo , Diagnóstico Preimplantación/métodos , Estudios RetrospectivosRESUMEN
Many integral membrane proteins might act as indispensable coordinators in specific functional microdomains to maintain the normal operation of known receptors, such as Notch. Gm364 is a multi-pass transmembrane protein that has been screened as a potential female fertility factor. However, there have been no reports to date about its function in female fertility. Here, we found that global knockout of Gm364 decreased the numbers of primordial follicles and growing follicles, impaired oocyte quality as indicated by increased ROS and γ-H2AX, decreased mitochondrial membrane potential, decreased oocyte maturation, and increased aneuploidy. Mechanistically, Gm364 directly binds and anchors MIB2, a ubiquitin ligase, on the membrane. Subsequently, membrane MIB2 ubiquitinates and activates DLL3. Next, the activated DLL3 binds and activates Notch2, which is subsequently cleaved within the cytoplasm to produce NICD2, the intracellular active domain of Notch2. Finally, NICD2 can directly activate AKT within the cytoplasm to regulate oocyte meiosis and quality.