Temporospatial gene expression of Prx1 and Prx2 is involved in morphogenesis of cranial placode-derived tissues through epithelio-mesenchymal interaction during rat embryogenesis

Abstract

Paired-related homeobox transcription factors, PRX1 and PRX2, are verified to play essential roles in limb, heart and craniofacial development by analyses of knockout animals. Their gene expression in the embryonic primordia derived from the mesoderm and neural crest is confirmed by in situ hybridization. Nevertheless, a detailed localization of PRX1 and PRX2 was not carried out because of a lack of specific antibodies for each factor. We have previously confirmed the presence of PRX proteins in rat embryonic pituitary by using an antibody that recognizes both PRX1 and PRX2. However, the pituitary originates in the cranial placodes, not the mesoderm or neural crest. In this study, we analyze the temporospatial distribution of PRX1 and PRX2 with novel antibodies specific for each factor, together with a stem/progenitor marker SOX2 (sex-determining region Y-box 2) in the primordia formed by epithelio-mesenchymal interaction. We observe immunoreactive signals of both PRX proteins in rat embryo, showing a similar pattern to that obtained by in situ hybridization. In early embryogenesis, PRX proteins are not co-localized with SOX2 but PRX2 and/or PRX1-positive cells are present in the border or periphery of SOX2-positive primordia originating in the cranial placode. During advanced embryogenesis, either PRX2-positive cells become condensed in the border of SOX2-positive cells or PRX1 and/or PRX2 become co-localized with SOX2. Our results suggest that PRX proteins, especially PRX2, play a role in the morphogenesis of the primordial tissues formed by the epithelio-mesenchymal interaction and that neural crest cells contribute to the morphogenesis of tissues derived from the cranial placode.

Molecular Cloning of Rat and Porcine Retina-derived POU Domain Factor 1 (POU6F2) from a Pituitary cDNA Library

“…The clone showed conservation in the POU-specific and homeobox domains very similar to those of humans. A transfection assay in CHO cells was performed to examine whether the rat RPF1 regulates genes; Prop1 , Prrx1 and Prrx2 were chosen, and we sought to determine if they might play important roles in the pituitary stem/progenitor cells [9, 15, 23, 24]. Rat RPF1 showed significant stimulation of Prop1 (–2997/+21), by 3.6-fold, and significant repression of Prrx2 (–372/+21), by 0.61-fold (Fig.…”

“…We recently demonstrated that the transcription factors PROP1, PRRX1 and PRRX2 are present in pituitary stem/progenitor cells and are involved in the differentiation and development of pituitary hormone-producing cells [9, 15, 23, 24]. Moreover, transfection assays showed that RPF1 is able to modulate gene expression of Prop1 by stimulating it and that of Prrx2 by repressing it.…”

“…Moreover, transfection assays showed that RPF1 is able to modulate gene expression of Prop1 by stimulating it and that of Prrx2 by repressing it. Considering the results in the CHO cell line directly, RPF1, which is expressed in the early embryonic pituitary, may stimulate expression of Prop1 in pituitary primordium cells [29] and inversely repress that of Prrx2 appearing in a small number of postnatal pituitary cells [15]. Cloning of pituitary RPF1 thus provides a novel clue for clarifying the molecular mechanism of pituitary organogenesis.…”

“…Cloning of pituitary RPF1 thus provides a novel clue for clarifying the molecular mechanism of pituitary organogenesis. We observed that PRRX1 appears in embryonic pituitary stem/progenitor cells around day E15.5 in the rat by changing Prop1 expression [15], while RPF1 did not show any effect on the Prrx1 promoter. Hence, the mechanism of Prrx1 expression in the PROP1-positive cells remains to be clarified.…”

“…Subsequently, PRRX2 was first cloned from the mouse adult spleen [10], and is now known as a mesenchymal transcription factor that plays important roles in craniofacial and limb morphogenesis [11,12,13]. We demonstrated that PRRX2, as well as its close relative PRRX1, is present in the stem/progenitor cells of the anterior pituitary [14,15,16]. Finally, LHX2 is a member of the LIM-homeodomain protein family and has been previously identified as a transcription factor for Cga [17].…”