Single-cell profiling of lncRNAs in human germ cells and molecular analysis reveals transcriptional regulation of LNC1845 on LHX8
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ABSTRACT:
PROVIDER: PRJNA682049 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR13181234_1.fastq.gz | Fastqsanger.gz | |||
| SRR13181234_2.fastq.gz | Fastqsanger.gz | |||
| SRR13181235_1.fastq.gz | Fastqsanger.gz | |||
| SRR13181235_2.fastq.gz | Fastqsanger.gz | |||
| SRR13181236_1.fastq.gz | Fastqsanger.gz |
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Project description:We report the presence of many long non-coding RNAs (lncRNAs) which were specifically expressed in the germ cells during human gonadal development by single-cell profiling, and the functional characterization of one divergent lncRNA, LNC1845. Functional dissection using in vitro differentiation of human pluripotent stem cells to germ cells revealed the regulatory role of LNC1845 on a transcription factor essential for ovarian follicle development, LHX8. Molecular characterization elucidated that LNC1845 upregulated LHX8 expression by modulating the levels of histone modifications, H3K4me3 and H3K27Ac. Our study provides a comprehensive analysis of lncRNAs in developing human germ cells and a concrete example of lncRNA acting as a cis regulator during female germ cell development.
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Project description:The limited number of in vivo germ cells poses an impediment to genome-wide studies. Here, we applied a small-scale ChIP-Seq method on purified mouse fetal germ cells to generate genome-wide maps of four histone modifications (H3K4me3, H3K27me3, H3K27ac and H2BK20ac), facilitating the identification of active and repressed cis-regulatory elements in germ cells in vivo. Comparison of active chromatin state between somatic, embryonic stem cells (ESC) and germ cells revealed promoters and enhancers needed for stem cell maintenance and germ cell development. The nuclear receptor Nr5a2 motif is enriched at a subset of cis-regulatory regions and we confirm its role in germ cell differentiation. Interestingly, germ cells have comparatively more H3K27me3-marked sites that are absent in ESC and other somatic cell types. These repressed regions are enriched for retrotransposons and MHC genes and this indicates that these loci are specifically silenced in germ cells. Together, our study provides the first genome-wide histone modification maps of in vivo germ cells and revealed the molecular chromatin signatures unique to germ cells. Germ cells were FACS-purified from gonadal single cell suspension based on Pou5f1-GFP expression. ChIP-seq of Histone modification was done for two timepoints in this study: E11.5 (male/female), E13.5 (male). For E13.5 timepoint, two biological replicates were analyzed. In order to validate small scale ChIP-seq method limited number of ES cells were used to check consistency of ChIP-seq data.
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Project description:Non-coding RNAs exert diverse functions in many cell types. In addition to transcription factors from coding genes, non-coding RNAs may also play essential roles in shaping and directing the fate of germ cells. The presence of many long non-coding RNAs (lncRNAs) which are specifically expressed in the germ cells during human gonadal development were reported and one divergent lncRNA, LNC1845, was functionally characterized. Comprehensive bioinformatic analysis of these lncRNAs indicates that divergent lncRNAs occupied the majority of female and male germ cells. Integrating lncRNA expression into the bioinformatic analysis also enhances the cell-type classification of female germ cells. Functional dissection using in vitro differentiation of human pluripotent stem cells to germ cells revealed the regulatory role of LNC1845 on a transcription factor essential for ovarian follicle development, LHX8, by modulating the levels of histone modifications, H3K4me3 and H3K27Ac. Hence, bioinformatical analysis and experimental verification provide a comprehensive analysis of lncRNAs in developing germ cells and elucidate how an lncRNA function as a cis regulator during human germ cell development.
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Project description:Lhx8 is a member of the LIM-homeobox transcription factor family and preferentially expressed in oocytes and germ cells within the mouse ovary. We discovered that Lhx8 knockout females lose oocytes within 7 days after birth. At the time of birth, histological examination shows that Lhx8 deficient (Lhx8(-/-)) ovaries are grossly similar to the newborn wild type ovaries. Lhx8(-/-) ovaries fail to maintain the primordial follicles and the transition from primordial to growing follicles does not occur. Lhx8(-/-) ovaries misexpress oocyte-specific genes such as Gdf9, Pou5f1, and Nobox. Very rapid loss of oocytes may partly be due to drastic the down-regulation of Kit and Kitl in Lhx8(-/-) ovaries. We compared Lhx8(-/-) and wild-type ovaries using Affymetrix 430 2.0 microarray platform. Eighty (44%) of 180 of the genes down-regulated more than 5-fold in Lhx8(-/-) ovaries were preferentially expressed in oocytes, whereas only 3 (2%) of 146 genes up-regulated more than 5-fold in the absence of Lhx8 were preferentially expressed in oocytes. In addition, the comparison of genes regulated in Lhx8(-/-) and Nobox(-/-) newborn ovaries discovered a common set of 34 genes whose expression level is affected in both Lhx8 and Nobox deficient mice. Our findings show that Lhx8 is a critical factor for maintenance and differentiation of the oocyte during early oogenesis and it acts in part by down-regulating the Nobox pathway. This SuperSeries is composed of the following subset Series: GSE7774: Transcriptional changes in Lhx8 Null newborn mouse ovaries GSE7775: Microarray Analyses of Newborn Mouse Ovaries Lacking Nobox GSE7776: Ovarian Transcript Expression in Newborn Mouse Refer to individual Series
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Project description:Background: Long non-coding RNAs (lncRNAs) are an important class of pervasive genes involved in a variety of biological functions. They are aberrantly expressed in many types of diseases. We want to study the lncRNAs profiles in preeclampsia. Preeclampsia has been observed in patients with molar pregnancy where a fetus is absent demonstrating that the placenta is sufficient to cause the condition. So we analyze the lncRNAs profiles in preeclampsia placentas. In this study, we described the lncRNAs profiles in 6 preeclampsia placentas (T) and 5 matched normal pregnancy placentas (N) tissues by microarray. Methodology/Principal Findings: With abundant and varied probes accounting 33,045 LncRNAs in our microarray, the number of lncRNAs that expressed at a certain level could be detected is 28,443. From the data we found there were 738 lncRNAs that differentially expressed (M-bM-^IM-%1.5 fold-change) among preeclampsia placentas compared with matched controls. Up to 18,063 coding transcripts could be detected in placenta samples through 30,215 coding transcripts probes. Coding-non-coding gene co-expression networks (CNC network) were constructed based on the correlation analysis between the differential expressed lncRNAs and mRNAs. According to the GO-Pathway analysis of differential expressed lncRNAs/mRNAs, we choose three lncRNAs to analyze the relationship between lncRNAs and preeclampsia. LOC391533, LOC284100, CEACAMP8 were evaluated by qPCR in 40 of preeclampsia placentas and 40 of controls. The results showed three lncRNAs were aberrantly expressed in preeclampsia placentas compared with controls. Conclusions/Significance: Our study is the first one to determine genome-wide lncRNAs expression patterns in preeclampsia placenta by microarray. The results displayed that clusters of lncRNAs were aberrantly expressed in preeclampsia placenta compared with controls, which revealed that lncRNAs differentially expressed in preeclampsia placenta may exert a partial or key role in preeclampsia development. Misregulation of LOC391533, LOC284100, CEACAMP8 might be associated with preeclampsia. Taken together, this study may provide potential targets for future treatment of preeclampsia and novel insights into preeclampsia biology. LncRNAs/mRNAs profiles in 6 preeclampsia placentas and 5 matched normal pregnancy placentas tissues by microarray using Arraystar v2.0.
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