The rat peptidylarginine deiminase-encoding gene: structural analysis and the 5'-flanking sequence.

Genomic clones of the rat peptidylarginine deiminase (PAD)-encoding gene (PAD) were isolated, and the gene organization was analyzed by restriction mapping and nucleotide sequencing. The PAD spans more than 50 kb and contains 16 exons and 15 introns. The lengths of the introns from 0.5 kb to more than 16.5 kb. A 1.7-kb sequence in the 5'-flanking region was determined. S1 nuclease mapping revealed two putative cap sites 79 and 81 bp upstream from the N-terminal ATG codon of PAD, which had been determined by amino acid sequence analysis. This ATG was confirmed to be the translation start site, since no other ATG codon was found in the open reading frame downstream from the cap sites. The 5'-flanking sequence contains four potential SP1-binding sites, a putative Pit-1/GHF-1-binding site, four short sequences either identical or homologous to the sequences in the promoter regions of rat or human growth hormone encoding genes, as well as a sequence similar to an estrogen-responsive element. However, neither a typical TATAA box, nor CCAAT box is present. These results provide important clues for elucidating the mechanism of female-specific and/or sex cycle-dependent gene expression.     

Human spermidine synthase gene: structure and chromosomal localization

The human spermidine synthase (EC 2.5.1.16) gene was isolated from a genomic library constructed with DNA obtained from a human immunoglobulin G (IgG) myeloma cell line. Subsequent sequence analyses revealed that the gene comprised of 5,818 nucleotides from the cap site to the last A of the putative polyadenylation signal with 8 exons and 7 intervening sequences. The 5'-flanking region of the gene was extremely GC rich, lacking any TATA box but containing CCAAT consensus sequences. No perfect consensus sequence for the cAMP-responsive element for the AP-1 binding site was found, yet the gene contained seven AP-2 binding site consensus sequences. The putative polyadenylation signal was an unusual AATACA instead of AATAAA. Polymerase chain reaction analysis with DNA obtained from human x hamster somatic cell hybrids indicated that human spermidine synthase genomic sequences segregate with human chromosome 1. Transfection of the genomic clone into Chinese hamster ovary cells displaying a low endogenous spermidine synthase activity revealed that the gene was transiently expressed and hence in all likelihood represents a functional gene.     

Genomic organization and chromosomal location of the mouse vasoactive intestinal polypeptide 1 (VPAC1) receptor.

The gene encoding the mouse vasoactive intestinal polypeptide type 1 (VPAC1) receptor was cloned, and its structural organization was determined. The gene (Vipr1) is more than 16 kb in length and is divided into 13 exons. The 5'-flanking region is highly GC-rich and lacks an apparent TATA box, but contains a CCAAT box, three potential Sp1-binding sites, and two potential AP-2-binding sites. Promoter analysis of the 5'-flanking region of Vipr1 using a luciferase gene reporter system revealed that the isolated 5'-flanking region has functional promoter activity. The mouse Vipr1 gene is encoded by a single gene, which was mapped to the distal region of mouse chromosome 9. This region is syntenic with human chromosome 3p, where the human VPAC1 receptor gene has been mapped.     

Cloning, genomic organization, and tissue-specific expression of the RASL11B gene

RASL11B is a member of the small GTPase protein family with a high degree of similarity to RAS proteins. Cloning of RASL11B mRNA and in silico analyses revealed that the human RASL11B gene spans about 4.5 kb and comprises four exons on chromosomal locus 4q12. The proximal 5'-flanking region of the gene lacks a TATA box but is GC-rich and contains a CCAAT box and several Sp1 sites. Consistent with this, the RASL11B gene was found to be expressed in all tissues investigated, with highest levels in placenta and in primary macrophages. The predicted RASL11B protein has no typical prenylation signal, indicating that it is probably not anchored to cellular membranes. RASL11B was induced during maturation of THP-1 monocytic cells into macrophage-like cells and in coronary artery smooth muscle cells after treatment with TGF-beta1. These results indicate that RASL11B may play a role in TGF-beta1-mediated developmental processes and in pathophysiologies such as inflammation, cancer, and arteriosclerosis.