Essential roles of retinoic acid signaling in interdigital apoptosis and control of BMP-7 expression in mouse autopods

We previously reported that mice lacking the RAR gamma gene and one or both alleles of the RAR beta gene (i.e., RAR beta(+/-)/RAR gamma(-/-) and RAR b eta(-/-)/RAR gamma(-/-) mutants) display a severe and fully penetrant inter digital webbing (soft tissue syndactyly), caused by the persistence of the fetal interdigital mesenchyme (Ghyselinck ct al., 1997, Int. J. Dev. Biol. 41, 425-447). In the present study, these compound mutants were used to inv estigate the cellular and molecular mechanisms involved ill retinoic acid ( RA)-dependent formation of the interdigital necrotic zones (INZs). The muta nt INZs show a marked decrease in the number of apoptotic cells accompanied by an increase of cell proliferation. This marked decrease was not paralle led by a reduction of the number of macrophages, indicating that the chemot actic cues which normally attract these cells into the INZs were not affect ed. The expression of a number of genes known to be involved in the establi shment of the INZs, the patterning of the autopod, and/or the initiation of apoptosis was also unaffected. These genes included BMP-2, BMP-4, Msx-1, M sx-2, 5' members of Hox complexes, Bcl2, Bar, and p53. In contrast the muta nt INZs displayed a specific, graded, down-regulation of tissue transglutam inase (tTG) promoter activity and of stromelysin-3 expression upon the remo val of one or both alleles of the RAR beta gene from the RAR gamma null gen etic background. As retinoic acid response elements are present in the prom oter regions of both tTG and stromelysin-3 genes,we propose that RA might i ncrease the amount of cell death in the INZs through a direct modulation of tTG expression and that it also contributes to the process of tissue remod eling, which accompanies cell death, through an up-regulation of stromelysi n-3 expression ill the INZs. Approximately 10% of the RAR beta(-/-) /RAR ga mma(-/-) mutants displayed a supernumerary preaxial digit on hindfeet, whic h is also a feature of the BMP-7 null phenotype (Dudley ct at, 1995, Genes Dev. 9, 2795-2807; Luo ct at, 1995, Genes Dev. 9, 2808-2820). BMP-7 was glo bally down-regulated at an early stage in the autopods of these RAR double null mutants, prior to the appearance of the digital rays. Therefore, RA ma y exert some of its effects on anteroposterior autopod patterning through c ontrolling BMP-7 expression. (C) 1999 Academic Press.