Paralogous mouse Hox genes, Hoxa9, Hoxb9, and Hoxd9, function together to control development of the mammary gland in response to pregnancy

Abstract

Although the role of Hox genes in patterning the mammalian body plan has been studied extensively during embryonic and fetal development, relatively little is known concerning Hox gene function in adult animals. Analysis of mice with mutant Hoxa9, Hoxb9, and Hoxd9 genes shows that these paralogous genes are required for mediating the expansion and/or differentiation of the mammary epithelium ductal system in response to pregnancy. Mothers with these three mutant genes cannot raise their own pups, but the pups can be rescued by fostering by wild-type mothers. Histologically, the mammary glands of the mutant mothers seem normal before pregnancy but do not develop properly in response to pregnancy and parturition. Hoxa9, Hoxb9, and Hoxd9 are expressed normally in adult mammary glands, suggesting a direct role for these genes in the development of mammary tissue after pregnancy. Because loss-of-function mutations in these Hox genes cause hypoplasia of the mammary gland after pregnancy, it may be productive to look for misexpression of these genes in mammary carcinomas.

Figure 1

The percentage of newborn pups that survived to weaning decreased dramatically when the females carried a greater number of mutant Hox 9 alleles. About 94% of the pups born to control females survived to weaning. The newborn survival rate dropped to 9% for pups of Aabbdd females and to 0% for pups of aaBbdd and aabbdd females.

Figure 2

Hypoplasia is evident in the mammary glands of the Aabbdd, aaBbdd, and aabbdd mutant females during pregnancy and around parturition. A and B are whole-mount preparations (×2.5). C, D, G, and H are sections under low magnification (×62). E, F, I, and J are sections under high magnification (×250). A, C, and E are from the fourth mammary glands of wild-type females at 12.5 days of pregnancy. B, D, and F are from the fourth mammary glands of aabbdd females at 12.5 days of pregnancy . G and I are from the fourth mammary gland of a wild-type female shortly post partum. H and J are from the fourth mammary gland of an aabbdd female shortly post partum. The mammary epithelia (e) include mammary ducts (d) and lobuloalveolar structures (a) in pregnant and lactating females. The mammary stroma (s) consist mainly of fat cells (f) and fibroblasts. The arrow in J points to an intracellular oil droplet in an alveolar cell.

Figure 3

RNA whole-mount in situ hybridization on E12.5 Hoxb9 (A) and Hoxd9 (B) embryos. Arrows point to the dense mesenchyme surrounding the primitive nipples. (C) Results of reverse transcription–PCR analyses with templates from various sources: the mammary gland of a virgin mouse (V), the mammary gland from a mouse at 6.5 days of pregnancy (P6.5), the mammary gland from a mouse at 12.5 days of pregnancy (P12.5), the mammary gland from a mouse that had been lactating for 2 days (L2), the mammary gland from a mouse that had been lactating for 7 days (L7), E12.5 embryos without heads (E), adult liver (Lvr), and a no-template control (N).