Isolation and characterization of AINT: a novel ARNT interacting protein expressed during murine embryonic development.

Basic helix-loop-helix-PER-ARNT-SIM (bHLH-PAS) proteins form dimeric transcription factors to mediate diverse biological functions including xenobiotic metabolism, hypoxic response, circadian rhythm and central nervous system midline development. The Ah receptor nuclear translocator protein (ARNT) plays a central role as a common heterodimerization partner. Herein, we describe a novel, embryonically expressed, ARNT interacting protein (AINT) that may be a member of a larger coiled-coil PAS interacting protein family. The AINT C-terminus mediates interaction with the PAS domain of ARNT in yeast and interacts in vitro with ARNT and ARNT2 specifically. AINT localizes to the cytoplasm and overexpression leads to non-nuclear localization of ARNT. A dynamic pattern of AINT mRNA expression during embryogenesis and cerebellum ontogeny supports a role for AINT in development.

Forkhead transcription factor FoxF2 is expressed in mesodermal tissues involved in epithelio-mesenchymal interactions.

The growing family of forkhead transcription factors plays many important roles during embryonic development. In this study we have used in situ hybridization to explore the expression pattern of the forkhead transcription factor gene FoxF2 (FREAC-2, LUN) during mouse and rat embryogenesis, postnatal development, and in adult tissues. We demonstrate that FoxF2 is expressed in the mesenchyme adjacent to the epithelium in alimentary, respiratory, and urinary tracts, similar to FoxF1 (FREAC-1, HFH-8). FoxF2 mRNA was also observed in organs that do not express FoxF1 during embryogenesis, e.g., in the central nervous system, eye, ear, and limb buds. In organs that express both FoxF2 and FoxF1, these transcription factors may have similar functions in epithelio-mesenchymal cross-talk, but the fact that FoxF2 is more widely expressed than FoxF1 suggests that FoxF2 also has an independent role as a developmental regulator.

A forkhead gene, FoxE3, is essential for lens epithelial proliferation and closure of the lens vesicle.

In the mouse mutant dysgenetic lens (dyl) the lens vesicle fails to separate from the ectoderm, causing a fusion between the lens and the cornea. Lack of a proliferating anterior lens epithelium leads to absence of secondary lens fibers and a dysplastic, cataractic lens. We report the cloning of a gene, FoxE3, encoding a forkhead/winged helix transcription factor, which is expressed in the developing lens from the start of lens placode induction and becomes restricted to the anterior proliferating cells when lens fiber differentiation begins. We show that FoxE3 is colocalized with dyl in the mouse genome, that dyl mice have mutations in the part of FoxE3 encoding the DNA-binding domain, and that these mutations cosegregate with the dyl phenotype. During embryonic development, the primordial lens epithelium is formed in an apparently normal way in dyl mutants. However, instead of the proliferation characteristic of a normal lens epithelium, the posterior of these cells fail to divide and show signs of premature differentiation, whereas the most anterior cells are eliminated by apoptosis. This implies that FoxE3 is essential for closure of the lens vesicle and is a factor that promotes survival and proliferation, while preventing differentiation, in the lens epithelium.

The cellular and developmental expression of hrs-2 in rat.

The molecular events underlying vesicular trafficking probably involve the formation and dissolution of protein complexes between integral components of the vesicle and its target membrane. SNAP-25 is associated with the plasma membrane and is a component of a core protein complex thought to be essential for neurotransmitter release. We have previously characterized a protein, hrs-2, that interacts with SNAP-25 and inhibits secretion from permeabilized PC12 cells. The cellular localization and developmental expression patterns of a number of proteins involved in the secretion machinery have been documented. To understand more about the possible cellular role of hrs-2, we have examined hrs-2 distribution, developmental expression and subcellular localization in rat tissues and cell lines. We show herein that the distribution of hrs-2 in brain and periphery parallels that of SNAP-23/25, and that recombinant hrs-2 binds to both SNAP-23 and SNAP-25. Hrs-2 mRNA and protein are found almost ubiquitously in neurons in the brain. Hrs-2 mRNA is expressed in the neural tube at E10 and thereafter mRNA and protein levels remain relatively constant in the whole brain through adulthood. In cultured PC12 cells, endogenous hrs-2 is expressed in the cytoplasm and on the limiting membranes of multivesicular bodies. Overexpression of hrs-2 in mammalian cells results in the appearance of large intracellular compartments that are labelled with hrs-2 antibodies. The wide distribution, the interaction with SNAP-23 and the localization on multivesicular body membranes suggest a general role for hrs-2 in cellular machinery.

Cord blood concentrations of vitamin A in preterm infants.

Plasma vitamin A concentrations were measured in cord blood samples from 56 infants of gestational ages < 33 weeks. Outcome was followed prospectively. Mothers' dietary habits and use of multivitamins during pregnancy were evaluated by means of a questionnaire. Vitamin A concentrations less than 1.05 micromol/l (low) were measured in 22, but levels below 0.7 micromol/l (deficient) only in two cases. The concentrations were not correlated with the infants' gestational ages. Infants with low concentrations were significantly more often multiplets compared to those with normal levels and the vitamin A concentrations of the multiplets were significantly lower than those of the singletons. The outcome measures used and the mothers' dietary habits and multivitamin use were similar in cases with low and normal vitamin A concentrations. Multiple gestation seems to be correlated with low plasma vitamin A concentrations in preterm infants at birth, and a complete assessment of vitamin A status to detect possible deficiency might be indicated in these cases.

FREAC-1 contains a cell-type-specific transcriptional activation domain and is expressed in epithelial-mesenchymal interfaces.

The forkhead transcription factor FREAC-1 is a potent transcriptional activator. We have localized a transcriptional activation domain in the C-terminus of FREAC-1 and another one to a stretch of approximately 60 amino acids in the central part of the protein. While the C-terminal activation domain activates in all cell lines tested, the activation domain in the central part of the protein is functional only in cell lines derived from lung. This cell-type-specific activity is retained when the activation domain is fused to the heterologous DNA binding domain of Gal4. The human FREAC-1 gene was found to consist of two exons separated by an intron of 1.2 kb. Exon 1 encodes the forkhead DNA binding domain and the cell-type-specific activation domain. Exon 2 encodes the general activation domain. The distribution of FREAC-1 expression during embryogenesis was investigated by in situ hybridization. FREAC-1 mRNA was found in mesenchyme in immediate proximity to endodermal epithelia throughout the digestive, urinary, and respiratory tracts. Mesenchyme surrounding the notochord and adjacent to the ectodermal epithelia of the oral cavity and developing teeth also expresses FREAC-1. The pattern of FREAC-1 expression, with highest levels in the mesenchyme next to the epithelium and gradually diminishing as the distance from the epithelium increases, suggests that FREAC-1 expression is a response to epithelial paracrine signaling and that FREAC-1 may play a role in epitheliomesenchymal interactions.