Project description:Purpose: To identify the changes in postnatal mouse conjunctival forniceal gene expression and their regulation by Klf4 around eye opening stage when the goblet cells first appear. Methods: Laser-capture-microdissection was used to collect conjunctival forniceal epithelial cells from postnatal-day (PN) 9, PN14 and PN20 wild-type (WT), and PN14 Klf4-conditional null (Klf4CN) mice, where goblet cells are absent, developing, present, and missing, respectively. Microarrays were used to compare gene expression among these four groups. Expression of selected genes was validated by Q-RT-PCR, and spatiotemporal expression assessed by in situ hybridization. Results: We identified 668, 251, 1160 and 139 genes that were upregulated and 492, 377, 1419 and 57 genes that were downregulated between PN9 and PN14, PN14 and PN20, PN9 and PN20, and PN14 WT and Klf4CN conjunctiva, respectively. Transcription factors Spdef, FoxA1 and FoxA3 that regulate goblet cell development in other mucosal epithelia, and epithelial specific Ets (ESE) transcription factor family members were upregulated during conjunctival development. Mesenchymal-epithelial transition (MET) was favored and diverse pathways related to glycoprotein biosynthesis, mucosal immunity, signaling, endocytic and neural regulation were affected during conjunctival development. Conjunctival Klf4-target genes differed significantly from the previously identified corneal Klf4-target genes, implying tissue-dependant regulatory targets for Klf4. Conclusions: We have identified the changes in gene expression accompanying mouse conjunctival development and the role of Klf4 in this process. These studies provide new probes to study conjunctival epithelial development and function, and reveal that the gene regulatory network required for goblet cell development is conserved across different mucosal epithelia. Three independent samples in each of four developmental groups
Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other
Project description:Purpose: To identify the changes in postnatal mouse conjunctival forniceal gene expression and their regulation by Klf4 around eye opening stage when the goblet cells first appear. Methods: Laser-capture-microdissection was used to collect conjunctival forniceal epithelial cells from postnatal-day (PN) 9, PN14 and PN20 wild-type (WT), and PN14 Klf4-conditional null (Klf4CN) mice, where goblet cells are absent, developing, present, and missing, respectively. Microarrays were used to compare gene expression among these four groups. Expression of selected genes was validated by Q-RT-PCR, and spatiotemporal expression assessed by in situ hybridization. Results: We identified 668, 251, 1160 and 139 genes that were upregulated and 492, 377, 1419 and 57 genes that were downregulated between PN9 and PN14, PN14 and PN20, PN9 and PN20, and PN14 WT and Klf4CN conjunctiva, respectively. Transcription factors Spdef, FoxA1 and FoxA3 that regulate goblet cell development in other mucosal epithelia, and epithelial specific Ets (ESE) transcription factor family members were upregulated during conjunctival development. Mesenchymal-epithelial transition (MET) was favored and diverse pathways related to glycoprotein biosynthesis, mucosal immunity, signaling, endocytic and neural regulation were affected during conjunctival development. Conjunctival Klf4-target genes differed significantly from the previously identified corneal Klf4-target genes, implying tissue-dependant regulatory targets for Klf4. Conclusions: We have identified the changes in gene expression accompanying mouse conjunctival development and the role of Klf4 in this process. These studies provide new probes to study conjunctival epithelial development and function, and reveal that the gene regulatory network required for goblet cell development is conserved across different mucosal epithelia.
Project description:To study effect of VRK1 deletion on spermatogenesis of the mouse, transciptomic analysis of genes in postnatal 8-day testicular cells of wild type and VRK1-deficient Mus musculus was performed.