Dataset Information

Profiling gene expression during the differentiation and development of the murine embryonic gonad.

ABSTRACT: The application of microarray technology to the study of mammalian organogenesis can provide greater insights into the steps necessary to elicit a functionally competent tissue. To this end, a temporal profile of gene expression was generated with the purpose of identifying changes in gene expression occurring within the developing male and female embryonic gonad. Gonad tissue was collected from mouse embryos at 11.5, 12.5, 14.5, 16.5, and 18.5 days postcoitum (dpc) and relative steady-state levels of mRNA were determined using the Affymetrix MGU74v2 microarray platform. Statistical analysis produced 3693 transcripts exhibiting differential expression during male and/or female gonad development. At 11.5 dpc, the gonad is morphologically indifferent, but at 12.5 dpc, transitions to a male or female phenotype are discernible by the appearance of testicular cords. A number of genes are expressed during this period and many share similar expression profiles in both sexes. As expected, the expression of two well-known sex determination genes, specifically Sry and Sox9, is unique to the testis. Beyond 12.5 dpc, differential gene expression becomes increasingly evident as the male and female tissue morphologically and physiologically diverges. This is evident by two unique waves of transcriptional activity occurring after 14.5 dpc in the male and female. With this study, a large number of transcripts comprising the murine transcriptome can be examined throughout male and female embryonic gonad development and allow for a more complete description of gonad differentiation and development.

SUBMITTER: Small CL

PROVIDER: S-EPMC3217241 | biostudies-literature | 2005 Feb

REPOSITORIES: biostudies-literature

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Profiling gene expression during the differentiation and development of the murine embryonic gonad.

Small Christopher L CL Shima James E JE Uzumcu Mehmet M Skinner Michael K MK Griswold Michael D MD

Biology of reproduction 20041020 2

The application of microarray technology to the study of mammalian organogenesis can provide greater insights into the steps necessary to elicit a functionally competent tissue. To this end, a temporal profile of gene expression was generated with the purpose of identifying changes in gene expression occurring within the developing male and female embryonic gonad. Gonad tissue was collected from mouse embryos at 11.5, 12.5, 14.5, 16.5, and 18.5 days postcoitum (dpc) and relative steady-state lev ...[more]

PMID: 15496517

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Project description:BACKGROUND: In birds as in mammals, a genetic switch determines whether the undifferentiated gonad develops into an ovary or a testis. However, understanding of the molecular pathway(s) involved in gonad differentiation is still incomplete. METHODOLOGY/PRINCIPAL FINDINGS: With the aim of improving characterization of the molecular pathway(s) involved in gonad differentiation in the chicken embryo, we developed a large scale real time reverse transcription polymerase chain reaction approach on 110 selected genes for evaluation of their expression profiles during chicken gonad differentiation between days 5.5 and 19 of incubation. Hierarchical clustering analysis of the resulting datasets discriminated gene clusters expressed preferentially in the ovary or the testis, and/or at early or later periods of embryonic gonad development. Fitting a linear model and testing the comparisons of interest allowed the identification of new potential actors of gonad differentiation, such as Z-linked ADAMTS12, LOC427192 (corresponding to NIM1 protein) and CFC1, that are upregulated in the developing testis, and BMP3 and Z-linked ADAMTSL1, that are preferentially expressed in the developing ovary. Interestingly, the expression patterns of several members of the transforming growth factor ? family were sexually dimorphic, with inhibin subunits upregulated in the testis, and bone morphogenetic protein subfamily members including BMP2, BMP3, BMP4 and BMP7, upregulated in the ovary. This study also highlighted several genes displaying asymmetric expression profiles such as GREM1 and BMP3 that are potentially involved in different aspects of gonad left-right asymmetry. CONCLUSION/SIGNIFICANCE: This study supports the overall conservation of vertebrate sex differentiation pathways but also reveals some particular feature of gene expression patterns during gonad development in the chicken. In particular, our study revealed new candidate genes which may be potential actors of chicken gonad differentiation and provides evidence of the preferential expression of BMPs in the developing ovary and Inhibin/Activin subunits in the developing testis.

Project description:BackgroundUnderstanding the mechanisms controlling stem cell differentiation is the key to future advances in tissue and organ regeneration. Embryonic stem (ES) cell differentiation can be triggered by embryoid body (EB) formation, which involves ES cell aggregation in suspension. EB growth in the absence of leukaemia inhibitory factor (LIF) leads EBs to mimic early embryonic development, giving rise to markers representative of endoderm, mesoderm and ectoderm. Here, we have used microarrays to investigate differences in gene expression between 3 undifferentiated ES cell lines, and also between undifferentiated ES cells and Day 1-4 EBs.ResultsAn initial array study identified 4 gene expression changes between 3 undifferentiated ES cell lines. Tissue culture conditions for ES differentiation were then optimized to give the maximum range of gene expression and growth. -Undifferentiated ES cells and EBs cultured with and without LIF at each day for 4 days were subjected to microarray analysis. -Differential expression of 23 genes was identified. 13 of these were also differentially regulated in a separate array comparison between undifferentiated ES cells and compartments of very early embryos. A high degree of inter-replicate variability was noted when confirming array results. Using a panel of marker genes, RNA amplification and RT-PCR, we examined expression pattern variation between individual -D4-Lif EBs. We found that individual EBs selected from the same dish were highly variable in gene expression profile.ConclusionES cell lines derived from different mouse strains and carrying different genetic modifications are almost invariant in gene expression profile under conditions used to maintain pluripotency. Tissue culture conditions that give the widest range of gene expression and maximise EB growth involve the use of 20% serum and starting cell numbers of 1000 per EB. 23 genes of importance to early development have been identified; more than half of these are also identified using similar studies, thus validating our results. EBs cultured in the same dish vary widely in terms of their gene expression (and hence, undoubtedly, in their future differentiation potential). This may explain some of the inherent variability in differentiation protocols that use EBs.

Dataset Information

Profiling gene expression during the differentiation and development of the murine embryonic gonad.

Publications

Profiling gene expression during the differentiation and development of the murine embryonic gonad.

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