Genomic organization and chromosomal localization of the mouse IKBKAP gene.

The autosomal recessive disorder familial dysautonomia (FD) has recently been demonstrated to be caused by mutations in the IKBKAP gene, so named because an initial report suggested that it encoded an IkappaB kinase complex associated protein (IKAP). Two mutations in IKBKAP have been reported to cause FD. The major mutation is a T-->C transition in the donor splice site of intron 20 and the minor mutation is a missense mutation in exon 19 that disrupts a consensus serine/threonine kinase phosphorylation site. We have characterized the cDNA sequences of the mouse, rat and rabbit IKBKAP-encoded mRNAs and determined the genomic organization and chromosomal location of mouse IKBKAP. There is significant homology in the amino acid sequence of IKAP across species and the serine/threonine kinase phosphorylation site altered in the minor FD mutation of IKAP is conserved. The mouse and human IKBKAP genes exhibit significant conservation of their genomic organization and the intron 20 donor splice site sequence, altered in the major FD mutation, is conserved in the human and mouse genes. Mouse IKBKAP is located on the central portion of chromosome 4 and maps to a region in which there is conserved linkage homology between the human and mouse genomes. The homologies observed in the human and mouse sequences should allow, through the process of homologous recombination, for the generation of mice that bear the IKBKAP mutations present in individuals with FD. The characterization of such mice should provide significant information regarding the pathophysiology of FD.     

Genomic organization and chromosomal localization of mouse proteinase 3 (Myeloblastin)

Proteinase 3 (PR3), is a matrix-degrading serine proteinase expressed in different hematopoietic cell lineages. The PR3 protein appears to regulate the myeloid differentiation and was found to be the autoantigen associated with Wegener granulomatosis. We have isolated and characterized the gene for mouse PR3 (mPR3) and determined its chromosomal location. The gene has been localized to Chromosome (Chr) 10. Comparison of mouse PR3 genomic structure with that of its human counterpart indicates that: 1) the mPR3 gene spans 7 kb organized in 5 exons and 4 introns, 2) the codons of His-Asp-Ser of the catalytic site are conserved and spread out over different exons, similar to the human gene, and 3) the gene product encodes a pre-proform of the protein. Knowledge of the structure and chromosomal location of the mPR3 gene may help better the understanding of the temporal and cell-specific expression of mouse PR3. Received: 14 July 1998 / Accepted: 28 October 1998     

Genomic organization and chromosomal localization of the mouse protein kinase C alpha gene

In Chinese hamster extended blocks of telomeric-like repeats were previously detected by in situ hybridization at the pericentromeric region of most chromosomes and short arrays were localized at several interstitial sites. In this work, we analyzed the molecular organization of internal telomeric sequences (ITs) in the Chinese hamster genome. In genomic transfers hybridized with a telomeric probe, multiple Bal31 insensitive fragments were detected. Most of the fragments ranged in size between less than 1 kb and more than 100 kb and some were polymorphic. Fluorescence in situ hybridization experiments on DNA fibers and on elongated chromosomes showed that the pericentromeric ITs are composed of extensive and essentially continuous arrays of telomeric-like sequences. We then isolated three genomic regions which contain short ITs. These ITs are localized at interstitial sites (3q13-15, 3q21-26, 1p26) and are composed of 29-126 bp of (TTAGGG)(n) repeats. A peculiar feature of all the three ITs is the AT richness of the flanking sequences. Since AT-rich DNA is known to be unstable and characteristic of several mammalian fragile sites, we propose that the three ITs were inserted at these sites during the repair of double strand breaks.     

Genomic organization and chromosomal localization of the TAPA-1 gene.

TAPA-1 is a 26-kDa integral membrane protein expressed on many human cell types. Antibodies against TAPA-1 induce homotypic aggregation of cells and can inhibit their growth. The murine homologue of TAPA-1 was cloned from both cDNA and genomic DNA libraries. A very high level of homology was found between human and mouse TAPA-1. The 5' untranslated region of the TAPA-1 gene resembles housekeeping gene promoters with respect to G + C content and the presence of potential Sp1 binding sites. The chromosomal localization of human and murine TAPA-1 genes was determined by Southern blot experiments using DNA from somatic cell hybrids. The genes were found to be part of a conserved syntenic group in mouse chromosome 7 and the short arm of human chromosome 11. The organization of the TAPA-1 gene and the projection of the exon boundaries on the proposed protein structure are presented.     

Genomic organization and chromosomal localization of the human casein gene family

Five yeast artificial chromosome (YAC) clones containing the human casein gene family were isolated and characterized to study the control mechanisms for the expression of these genes. Partial restriction analysis in conjunction with the chromosomal fragmentation method and fluorescence in situ hybridization (FISH) analysis were performed to construct a detailed physical map of the casein gene family and to determine the chromosomal localization of these genes. The isolated YAC clones 748F3, 750D11, 882G11, 886B3 and 960D2 were 1.2 Mb, 860 kb, 800 kb 1.5 Mb and 1.5 Mb in size, respectively. The clones 748F3, 882G11, 886B3 and 960D2 contained the entire casein gene family, while the κ-casein gene was absent in 750D11. The human αS1-, β- and κ-casein genes were found to be closely linked and arranged in the order αS1-β-κ. The distance between αS1 and β, and between αS1 and κ was approximately 10 and 300 kb, respectively. The β-casein gene was oriented in the opposite direction to the αS1- and κ-casein genes. The casein gene family was localized to chromosome 4q21.1 by FISH analysis. Received: 7 July 1996 / Revised: 29 October 1996     

Genomic structure and chromosomal localization of the mouse persyn gene

Synucleins are a family of small intracellular proteins expressed mainly in the nervous system. The involvement of synucleins in neurodegeneration and malignancy has been demonstrated, but the physiological functions of these proteins remain elusive. Further studies including generation of animals with modified persyn expression are necessary to clarify the functions of these proteins and the mechanisms of their involvement in human diseases. We cloned and determined the organization and chromosomal localization of the mouse gene coding for persyn, a member of the synuclein family. The gene is composed of five exons, and its general structure is very similar to that of the human persyn gene. Using fluorescence in situ hybridization, we assigned the persyn gene to the boundary of bands B and C on mouse chromosome 14. We found a fragment of the gene that directs expression of the persyn protein in sensory neurons and could be used for generation of transgenic animals.     

Genomic structure and chromosomal localization of the mouse gene Punc

The mouse gene Punc encodes a member of the immunoglobulin superfamily of cell surface proteins. It is highly expressed in the developing embryo in nervous system and limb buds. At mid-gestation, however, expression levels of Punc decrease sharply. To allow investigation of such a regulatory mechanism, the genomic locus encompassing the Punc gene was cloned, characterized, and mapped. Fluorescent in situ hybridization was used to determine the chromosomal location of the Punc gene of mouse and human. Mouse Punc maps to Chromosome (Chr) 9 in the region D-E1, whereas the human PUNC gene is localized to Chr 15 at 15q22.3-23, a region known to be syntenic to mouse 9D-E1. The human PUNC gene therefore maps close to a genetic locus that is linked to Bardet-Biedl Syndrome, an autosomal recessive human disorder. Confirmation for the location of human PUNC was obtained through sequence relationships between mouse Punc cDNA, human PUNC cDNA, genomic sequence upstream of the murine Punc gene, and human STS markers that had been previously mapped on Chr 15. The STS sequence WI-14920 is in fact derived from the 3′-untranslated region of the human PUNC gene. WI-14920 had been placed at 228cR from the top of the Chr 15 linkage group, which provided positional information for the human PUNC gene at high resolution. Thus, this study identifies PUNC as the gene corresponding to a previously anonymous marker and serves as a basis to investigate its role in genetic disorders. Received: 8 July 1998 / Accepted: 14 October 1998     

Genomic organization and chromosomal localization of the murine epididymal retinoic acid-binding protein (mE-RABP) gene

The murine epididymal retinoic acid-binding protein (mE-RABP) is specifically synthesized in the mouse mid/distal caput epididymidis and secreted in the lumen. In this report, we have demonstrated by Southern blot analysis of genomic DNA that mE-RABP is encoded by a single-copy gene. A mouse 129/SvJ genomic bacterial artificial chromosome (BAC) library was screened using a cDNA encoding the minor form of mE-RABP. One positive BAC clone was characterized and sequenced to determine the nucleotide sequence of the entire mE-RABP gene. The molecular cloning of the mE-RABP gene completes the characterization of the 20.5-kDa–predicted preprotein leading to the minor and major forms of mE-RABP. Comparison of the DNA sequence of the promoter and coding regions with that of the rat epididymal secretory protein I (ESP I) gene showed that the mE-RABP gene is the orthologue of the ESP I gene that encodes a rat epididymal retinoic acid-binding protein. Several regulatory elements, including a putative androgen receptor binding site, “CACCC-boxes,” NF-1, Oct-1, and SP-1 recognition sites, are conserved in the proximal promoter. Analysis of the nucleotide sequence of the mE-RABP gene revealed the presence of seven exons and showed that the genomic organization is highly related to other genes encoding lipocalins. The mE-RABP gene was mapped by fluorescent in situ hybridization to the [A3-B] region of the murine chromosome 2. Our data, combined with that of others, suggest that the proximal segment of the mouse chromosome 2 may be a rich region for genes encoding lipocalins with a genomic organization highly related to the mE-RABP gene. Mol. Reprod. Dev. 50:387–395, 1998. © 1998 Wiley-Liss, Inc.