Activation of the nuclear factor of activated T-cell (NFAT) family of transcription factors is associated with changes in gene expression and myocyte function in adult cardiac and skeletal muscle. However, the role of NFATs in normal embryonic heart development is not well characterized. In this report, the function of NFATc3 and NFATc4 in embryonic heart development was examined in mice with targeted disruption of both nfatc3 and nfatc4 genes. The nfatc3-/-nfatc4-/- mice demonstrate embryonic lethality after embryonic day 10.5 and have thin ventricles, pericardial effusion, and a reduction in ventricular myocyte proliferation. Cardiac mitochondria are swollen with abnormal cristae, indicative of metabolic failure, but hallmarks of apoptosis are not evident. Furthermore, enzymatic activity of complex II and IV of the respiratory chain and mitochondrial oxidative activity are reduced in nfatc3-/-nfatc4-/- cardiomyocytes. Cardiac-specific expression of constitutively active NFATc4 in nfatc3-/-nfatc4-/- embryos prolongs embryonic viability to embryonic day 12 and preserves ventricular myocyte proliferation, compact zone density, and trabecular formation. The rescued embryos also maintain cardiac mitochondrial ultrastructure and complex II enzyme activity. Together, these data support the hypothesis that loss of NFAT activity in the heart results in a deficiency in mitochondrial energy metabolism required for cardiac morphogenesis and function.

In Drosophila, the Dorsal protein establishes the embryonic dorso-ventral axis during development. Here we show that the vertebrate homologue of Dorsal, nuclear factor-kappa B (NF-kappaB), is vital for the formation of the proximo-distal organizer of the developing limb bud, the apical ectodermal ridge (AER). Transcription of the NF-kappaB proto-oncogene c-rel is regulated, in part, during morphogenesis of the limb bud by AER-derived signals such as fibroblast growth factors. Interruption of NF-kappaB activity using viral-mediated delivery of an inhibitor results in a highly dysmorphic AER, reduction in overall limb size, loss of distal elements and reversal in the direction of limb outgrowth. Furthermore, inhibition of NF-kappaB activity in limb mesenchyme leads to a reduction in expression of Sonic hedgehog and Twist but derepresses expression of the bone morphogenetic protein-4 gene. These results are the first evidence that vertebrate NF-kappaB proteins act to transmit growth factor signals between the ectoderm and the underlying mesenchyme during embryonic limb formation.

The nuclear factor-B (NF-B) family of transcription factors has been shown to regulate proliferation in several cell types. Although recent studies have demonstrated aberrant expression or activity of NF-B in human breast cancer cell lines and tumors, little is known regarding the precise role of NF-B in normal proliferation and development of the mammary epithelium. We investigated the function of NF-B during murine early postnatal mammary gland development by observing the consequences of increased NF-B activity in mouse mammary epithelium lacking the gene encoding IB␣, a major inhibitor of NF-B. Mammary tissue containing epithelium from inhibitor B␣ (IB␣)-deficient female donors was transplanted into the gland-free mammary stroma of wild-type mice, resulting in an increase in lateral ductal branching and pervasive intraductal hyperplasia. A two-to threefold increase in epithelial cell number was observed in IB␣-deficient epithelium compared with controls. Epithelial cell proliferation was strikingly increased in IB␣-deficient epithelium, and no alteration in apoptosis was detected. The extracellular matrix adjacent to IB␣-deficient epithelium was reduced. Consistent with in vivo data, a fourfold increase in epithelial branching was also observed in purified IB␣-deficient primary epithelial cells in three-dimensional culture. These data demonstrate that NF-B positively regulates mammary epithelial proliferation, branching, and functions in maintenance of normal epithelial architecture during early postnatal development. INTRODUCTIONThe mammary gland is an organ designed to deliver nourishment and passive immunity to infant mammals. It consists of an epithelium that synthesizes and secretes lipid and milk proteins, as well as a fatty stroma that provides support and local growth regulatory cues to the epithelium (reviewed by Medina, 1996). Although the mammary gland rudiment is established during embryogenesis, the majority of mammary gland development occurs postnatally. During puberty, the epithelium proliferates and branches in response to hormonal signals, eventually extending throughout the entire stroma. More extensive growth and differentiation of the epithelium occurs during each round of pregnancy. The distal tips of each epithelial branch proliferate and differentiate into lobuloalveoli, which synthesize and secrete milk during lactation. Upon cessation of nursing, the majority of the epithelium undergoes apoptosis in a process called involution (reviewed by Furth, 1999). After involution, the epithelium remains relatively quiescent until the next pregnancy, when the morphogenetic cycle is repeated.The nuclear factor-B (NF-B) family of transcription factors regulates growth, differentiation, and apoptosis in several tissues, including lymphocytes, embryonic limb, lung and liver, skin, and bone (Beg et al., 1995;Klement et al., 1996;Boothby et al., 1997;Franzoso et al., 1997;Bushdid et al., 1998;Kanegae et al., 1998;Seitz et al., 1998;Bendall et al., 1999;Hu et al., 1999; Li et al., 1999a,b,c;Takeda et...

In summary, embryonic erythroblasts are the primary target of ART toxicity in the rat embryo after in vivo treatment, preceding embryolethality and malformations.

It is becoming increasingly recognized that the ubiquitous, inducible transcription factor nuclear factor-kappaB (NF-kappaB) is involved in developmental processes. For example, NF-kappaB acts as a mediator of epithelial-mesenchymal interactions in the developing chick limb. We investigated the role of NF-kappaB in directing the branching morphogenesis of the developing chick lung, a process which relies on epithelial-mesenchymal communication. High level expression of relA was found in the mesenchyme surrounding the nonbranching structures of the lung but was not detected either in the mesenchyme surrounding the branching structures of the distal lung or in the developing lung epithelium. Specific inhibition of mesenchymal NF-kappaB in lung cultures resulted in increased epithelial budding. Conversely, expression of a trans-dominant activator of NF-kappaB in the lung mesenchyme repressed budding. Ectopic expression of RelA was sufficient to inhibit the ability of the distal mesenchyme to induce epithelial bud formation. Cellular proliferation in the mesenchyme was inhibited by hyperactivation of NF-kappaB in the mesenchyme of lung cultures. Interestingly, increased NF-kappaB activity in the mesenchyme also decreased the proliferation of the associated epithelium, while inhibition of NF-kappaB activity increased cellular proliferation in lung cultures. Expression patterns of several genes which are known to influence lung branching morphogenesis were altered in response to changes in mesenchymal NF-kappaB activity, including fgf10, bmp-4, and tgf-beta1. Thus NF-kappaB represents the first transcription factor reported to function within the lung mesenchyme to limit growth and branching of the adjacent epithelium.