A. Mouse Homeobox And POU-Domain Genes. A novel mouse homeobox gene NKx-1, was cloned and approximately 8 kb was sequenced. The deduced amino acid sequence of the Nkx-1 homeodomain differs from the Drosophila NK-1 homeodomain by only 3 of 60 amino acid residues. Both Nkx-1 and NK-1 proteins contain an acidic region before the homeodomain. Northern analysis revealed one major band of Nkx-1 poly A+ RNA in brain and trace bands in testes and spleen. The abundance of Nkx-1 poly A+ RNA is highest in 10 day mouse embryos and progressively decreases thereafter. In situ hybridization with sections of 14-day mouse embryos revealed Nkx-1 RNA in the mesencephalon, myelencephalon, spinal cord, vertebrae, and ribs. Five additional novel mouse homeobox genes were cloned and partially sequenced. A novel mouse POU-domain gene related to Brain-3 POU-domain CDNA was cloned and about 3 kb was sequenced. The expression of Brain-1, Brain-2, Brain-4 and Skip POU-domain genes in the mouse nervous system was determined by in situ hybridization as a function of developmental age. B. Regulation of a Calcium Channel a-1 Subunit Gene. A nucleotide sequence was found in the 5 -upstream regulatory region of a voltage-sensitive calcium channel alpha-1 subunit gene that is a powerful activator of an enhancerless chloramphenicol acetyltransferase reporter gene. A CDNA expression library was screened for recombinants that direct the synthesis of proteins that bind to this nucleotide sequence. Seven kinds of clones were found that encode proteins that bind to oligonucleotides with appropriate sequence specificity. C. Promotor And Enhancer Selection. CDNA clones were obtained that encode proteins that specifically bind to oligonucleotide sequences by a method that selects for gene regulatory sequences. Partial sequences of the CDNA clones were obtained.
