ABSTRACT
Long noncoding RNAs (lncRNAs) have been found to play an important role in the occurrence and development of endometrial carcinoma (EC). Here, using RNA sequencing analysis, we systemically screened and identified the lncRNA eukaryotic translation initiation factor 1A, X-linked (EIF1AX)-AS1, which is aberrantly downregulated in clinical EC tissues and closely correlated with tumor type. EIF1AX-AS1 markedly inhibited EC cell proliferation and promoted apoptosis in vitro and in vivo. Mechanistically, EIF1AX-AS1 interacts with EIF1AX mRNA and poly C binding protein 1 (PCBP1), which promote EIF1AX mRNA degradation. Intriguingly, by interacting with internal ribosome entry site-related protein Y-box binding protein 1 (YBX-1), EIF1AX promotes c-Myc translation through the internal ribosome entry site pathway. c-Myc promotes EIF1AX transcription and thus forms a feed-forward loop to regulate EC cell proliferation. Taken together, these data reveal new insights into the biology driving EC proliferation and highlights the potential of lncRNAs as biomarkers for prognosis and future therapeutic targets for cancer.
Subject(s)
Endometrial Neoplasms , RNA, Long Noncoding , Apoptosis/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Endometrial Neoplasms/metabolism , Female , Gene Expression Regulation, Neoplastic , Humans , Internal Ribosome Entry Sites , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , RNA, Messenger/genetics , Xenograft Model Antitumor AssaysABSTRACT
Preimplantation embryo development is characterized by drastic nuclear reprogramming and dynamic stage-specific gene expression. Key regulators of this earliest developmental stage have not been revealed. In the present study, a "non-classical" nuclear-localization pattern of eIF1A was observed during early developmental stages of mouse preimplantation embryo before late-morula. In particular, eIF1A is most highly expressed in the nuclear of 2-cell embryo. Knockdown eIF1A by siRNA microinjection affected the development of mouse preimplantation embryo, resulted in decreased blastocyst formation rate. CDX2 protein expression level significantly down-regulated after eIF1A knockdown in morula stage. In addition, the mRNA expression level of Hsp70.1 was also decreased in 2-cell embryo. The results indicate an indispensable role of eIF1A in mouse preimplantation embryos.
Subject(s)
Cell Nucleus/metabolism , Embryonic Development , Eukaryotic Initiation Factor-1/metabolism , Animals , Biomarkers/metabolism , Eukaryotic Initiation Factor-1/genetics , Female , Gene Expression Regulation, Developmental , Gene Knockdown Techniques , Genome , Male , Mice , Transcription Factors/metabolism , Zygote/metabolismABSTRACT
BACKGROUND: The tricho-rhino-phalangeal syndrome-1 gene (Trps1) is an atypical GATA family member. Although current studies of Trps1 mainly focus on tumors, whether Trps1 plays a role in the male reproductive system remains unknown. OBJECTIVES: The purpose of this study was to elucidate the function of Trps1 in Leydig cells, indicating its regulatory mechanism on the cell cycle. METHODS: Gene-silencing technology, RNA-seq, RT-qPCR, and western blotting were used to evaluate the function of Trps1 in mouse primary Leydig cells and MLTC-1 cells. In addition, ChIP-base sets and ChIP-qPCR were employed to further assess the regulatory mechanism of Trps1 in MLTC-1 cells. RESULTS: Knockdown of Trps1 in Leydig cells significantly suppressed phosphorylation of Src and Akt and expression of Ccnd1, which was accompanied by impairment of cell proliferative ability. Trps1 may affect the cell cycle through the Src/Akt/Ccnd1 signaling pathway. In addition, Trps1 may bind to the promoter of Srcin1 to regulate its transcription, thus influencing Src phosphorylation levels and the proliferation of Leydig cells. DISCUSSION AND CONCLUSION: Src increases in Leydig cells during pubertal development, suggesting its functional involvement in differentiated adult Leydig cells. Inhibition of the Src/Akt pathway would reduce Ccnd1 expression. In the present study, we found that Trps1 may regulate the phosphorylation level of Src and Akt through Srcin1, targeting Ccnd1 to influence mouse Leydig cell proliferation. These findings shed light on the regulation of Trps1 on cell proliferation and differentiation of mouse Leydig cells.
Subject(s)
Cell Proliferation/genetics , Cyclin D1/physiology , Leydig Cells/metabolism , Repressor Proteins/physiology , Animals , Cell Cycle/genetics , Cell Differentiation/genetics , Male , Mice , Signal Transduction/geneticsABSTRACT
The first cell lineage differentiation occurs during the development of mouse 8-cell embryo to blastocyst. Akt is a potent kinase whose role during blastocyst formation has not been elucidated. In the present study, immunofluorescence results showed that the Akt protein was specifically localized to the outer cells of the morula. Akt-specific inhibitor MK2206 significantly inhibited mouse blastocyst formation and resulted in decreased expression of the trophectoderm marker Cdx2 and led to granular distribution of ERα in the cytoplasm. Furthermore, knockdown of ERα by siRNA microinjection can also lead to a decrease in the development rate of mouse blastocysts, accompanied by a decrease in the expression level of Yap protein. We conclude that Akt may be indispensable for the first cell lineage differentiation of mouse.
Subject(s)
Cell Differentiation , Cell Lineage , Embryo, Mammalian/cytology , Proto-Oncogene Proteins c-akt/physiology , Adaptor Proteins, Signal Transducing/metabolism , Animals , Blastocyst/cytology , Cell Cycle Proteins/metabolism , Embryonic Development , Gene Expression Regulation, Developmental , Mice , Morula/chemistry , YAP-Signaling ProteinsABSTRACT
Infertility caused by environmental pollution is becoming a global problem, but an effective prevention or treatment is lacking. Icariin (ICA) is a flavonoid used in traditional Chinese medicine. The present study investigated the possible roles of ICA in preventing testicular dysfunction caused by di(2-ethylhexyl) phthalate (DEHP), one of the most studied environmental endocrine disruptors. Cultured mouse Leydig cells were pretreated with ICA and exposed to DEHP to determine ICA effects upon cell proliferation, reactive oxygen species (ROS) levels, mitochondrial membrane potential (Δψm), testosterone levels and the expression of transcription factor SF-1 and steroidogenic enzymes (CYP11, 3ß-HSD and 17ß-HSD), which play critical roles in androgen production. Our results showed that ICA reversed the adverse effect of DEHP on Leydig cell proliferation, and decreased ROS levels and elevated Δψm levels. Also, ICA promoted testosterone production and up-regulated the expression of SF-1 and steroidogenic enzymes. We investigated ICA actions in vivo, using male mice administrated DEHP followed by ICA. Exposure to DEHP decreased epididymal sperm counts and disrupted seminiferous tubules, and both of these effects were reversed by ICA treatment. These results showed that the mechanisms of ICA in protecting mouse testes against DEHP-induced damage involves the prevention of ROS accumulation and promotion of testosterone secretion.
Subject(s)
Diethylhexyl Phthalate/adverse effects , Flavonoids/pharmacology , Leydig Cells/drug effects , Phthalic Acids/adverse effects , Protective Agents/pharmacology , Testosterone/metabolism , Animals , Cell Proliferation/drug effects , Endocrine Disruptors/metabolism , Female , Leydig Cells/metabolism , Male , Mice , Mice, Inbred ICR , Pregnancy , Prenatal Exposure Delayed Effects/metabolism , Reactive Oxygen Species/metabolism , Spermatozoa/drug effects , Spermatozoa/metabolism , Testis/drug effects , Testis/metabolismABSTRACT
Zygotic genome activation (ZGA) is one of the most critical events at the beginning of mammalian preimplantation embryo development (PED). The mechanisms underlying mouse ZGA remain unclear although it has been widely studied. In the present study, we identified that tricho-rhino-phalangeal syndrome 1 (TRPS1), an atypical GATA family member, is an important factor for ZGA in mouse PED. We found that the Trps1 mRNA level peaked at the one-cell stage while TRPS1 protein did so at the two/four-cell stage. Knockdown of Trps1 by the microinjection of Trps1 siRNA reduced the developmental rate of mouse preimplantation embryos by approximately 30%, and increased the expression of ZGA marker genes MuERV-L and Zscan4d via suppressing the expression of major histone markers H3K4me3 and H3K27me3. Furthermore, Trps1 knockdown decreased the expression of Sox2 but increased Oct4 expression. We conclude that TRPS1 may be indispensable for zygotic genome activation during mouse PED.
