ABSTRACT: 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.
