1. Organization of the mouse cardiac natriuretic peptide locus encoding BNP and ANP
H Huang, S W John, M E Steinhelper J Mol Cell Cardiol. 1996 Aug;28(8):1823-8. doi: 10.1006/jmcc.1996.0172.
The genes encoding the mouse atrial natriuretic peptide and B-type natriuretic peptide were previously shown to be physically linked on mouse chromosome 4 (Steinhelper ME, 1993, Structure, expression, and genomic mapping of the mouse natriuretic peptide type-B gene. Circ Res 72: 984-992). In the present study the spatial relationship and orientation of the mouse atrial natriuretic peptide and B-type natriuretic peptide transcription units were identified and a physical map of the mouse cardiac natriuretic peptide locus was obtained. To this end, genomic clones encoding atrial natriuretic peptide and B-type natriuretic peptide were isolated from a mouse genomic library in bacteriophage P1. Three independent clones encoding atrial natriuretic peptide were isolated and two of these also encode B-type natriuretic peptide. Both transcripts were shown to arise from the same DNA strand, with B-type natriuretic peptide encoded approximately 15 kb 5'-of atrial natriuretic peptide based on field inversion gel electrophoresis of fragments amplified with specific oligonucleotides. This finding was confirmed by isolation of subclones comprising the entire locus and by blot hybridization analysis of mouse genomic DNA. The results show that the genes encoding the two natriuretic peptides expressed predominantly in mammalian cardiac myocytes are organized in tandem on mouse chromosome 4. This information provides a physical framework for investigating mechanisms that regulate transcription of the cardiac natriuretic peptide locus.
2. Two cardiac natriuretic peptide genes (atrial natriuretic peptide and brain natriuretic peptide) are organized in tandem in the mouse and human genomes
N Tamura, Y Ogawa, A Yasoda, H Itoh, Y Saito, K Nakao J Mol Cell Cardiol. 1996 Aug;28(8):1811-5. doi: 10.1006/jmcc.1996.0170.
Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), which act as cardiac hormones, are produced mainly by the atrium and ventricle, respectively, and are involved in body fluid homeostasis and blood pressure control. The ANP and BNP gene expressions are markedly augmented in ventricles of patients with a wide variety of cardiovascular diseases. It has been demonstrated that the ANP and BNP genes are tightly linked on mouse chromosome 4 and on the distal short arm of human chromosome 1. However, the precise physical map of the ANP and BNP genes has never been elucidated. In the present study, we characterized the genomic DNA fragment containing the ANP and BNP genes in mice and humans. Three genomic DNA clones harboring the entire mouse BNP gene were isolated from a 129/Sv mouse genomic DNA library. Nucleotide sequence analysis revealed that a phage clone (lambda mBNP #3) contains at its 3'-end the 5'-flanking region and the first 209-bp sequence of the first exon of the mouse ANP gene. In mice, the BNP gene was located about 12 kb upstream of the ANP gene. By polymerase chain reaction, we isolated an approximately 11-kb human genomic DNA fragment containing the third exon of the BNP gene and the first and second exons of the ANP gene. In humans, the BNP gene was located upstream of the ANP gene, approximately 8 kb apart. The present study provides the direct evidence that the ANP and BNP genes are organized in tandem in the mouse and human genomes.
3. Genes encoding atrial and brain natriuretic peptides as candidates for sensitivity to brain ischemia in stroke-prone hypertensive rats
M J Brosnan, J S Clark, B Jeffs, C D Negrin, P Van Vooren, S M Arribas, H Carswell, T J Aitman, C Szpirer, I M Macrae, A F Dominiczak Hypertension. 1999 Jan;33(1 Pt 2):290-7. doi: 10.1161/01.hyp.33.1.290.
-Previous studies suggested that atrial natriuretic peptide gene (Anp) and brain natriuretic peptide gene (Bnp) are plausible candidate genes for susceptibility to stroke and for sensitivity to brain ischemia in the stroke-prone spontaneously hypertensive rat (SHRSP). We performed structural and functional analyses of these 2 genes in SHRSP from Glasgow colonies (SHRSPGla) and Wistar-Kyoto rats from Glasgow colonies (WKYGla) and developed a radiation hybrid map of the relevant region of rat chromosome 5. Sequencing of the coding regions of the Anp and Bnp genes revealed no difference between the 2 strains. Expression studies in brain tissue showed no differences at baseline and at 24 hours after middle cerebral artery occlusion. Plasma concentrations of atrial natriuretic peptide (ANP) did not differ between the SHRSPGla and WKYGla, whereas concentrations of brain natriuretic peptide were significantly higher in the SHRSPGla as compared with the WKYGla (n=11 to 14; 163+/-21 pg/mL and 78+/-14 pg/mL; 95% confidence interval 31 to 138, P=0.003). We did not detect any attenuation of endothelium-dependent relaxations to bradykinin or ANP in middle cerebral arteries from the SHRSPGla; indeed the sensitivity to ANP was significantly increased in arteries harvested from this strain (WKYGla: n=8; pD2=7. 3+/-0.2 and SHRSPGla: n=8; pD2=8.2+/-0.15; P<0.01). Moreover, radiation hybrid mapping and fluorescence in situ hybridization allowed us to map the Anf marker in the telomeric position of rat chromosome 5 in close proximity to D5Rat48, D5Rat47, D5Mgh15, and D5Mgh16. These results exclude Anp and Bnp as candidate genes for the sensitivity to brain ischemia and pave the way to further congenic and physical mapping strategies.
