Pleiotropic functions of pituitary adenylyl cyclase-activating polypeptide on retinal ontogenesis: involvement of KLF4 in the control of progenitor cell proliferation.

We showed previously that the neuropeptide pituitary adenylyl cyclase-activating polypeptide (PACAP) negatively regulates proliferation of postnatal rat retinal progenitor cells through the downregulation of cyclin D1 in a cAMP/protein kinase A dependent manner. In the present study, we describe by microarray analysis several putative PACAP targets regulated by different transcription factor families. One of these families is the Sp/Klf family of transcriptional factors capable of regulating cyclin D1, and among members, we demonstrate by immunocytochemistry that KLF4 is expressed throughout rat retinal development by retinal progenitor cells and in most differentiated cell types. Using retinal explants preparations, PACAP treatment can transiently increase Klf4 mRNA levels; from electrophoretic mobility shift assays, PACAP is also able to increase the nuclear KLF4 content. From these results, we suggest that KLF4 may be involved in the anti-proliferative effects of PACAP as one mechanism regulating progenitor cell transition from proliferation to differentiation throughout retinal development.

The Krüppel-like factor 4 controls biosynthesis of thyrotropin-releasing hormone during hypothalamus development.

Embryonic neurogenesis is controlled by the activation of specific genetic programs. In the hypothalamus, neuronal thyrotropin-releasing hormone (TRH) populations control important physiological process, including energy homeostasis and autonomic function; however, the genetic program leading to the TRH expression is poorly understood. Here, we show that the Klf4 gene, encoding the transcription factor Krüppel-like factor 4 (Klf4), was expressed in the rat hypothalamus during development and regulated Trh expression. In rat fetal hypothalamic cells Klf4 regulated Trh promoter activity through CACCC and GC motifs present on the Trh gene promoter. Accordingly, hypothalamic Trh expression was down-regulated at embryonic day 15 in the Klf4(-/-) mice resulting in diminished bioactive peptide levels. Although at the neonatal stage the Trh transcript levels of the Klf4(-/-) mice were normal, the reduction in peptide levels persisted. Thus, our data indicate that Klf4 plays a key role in the maturation of TRH expression in hypothalamic neurons.