Sex-specific effects of dual ET-1/ANG II receptor (Dear) variants in Dahl salt-sensitive/resistant hypertension rat model
Yuji Kaneko,
Victoria L. M. Herrera,
Tamara Didishvili and
Nelson Ruiz-Opazo
Section Molecular Medicine, Department of Medicine, Boston University School of Medicine, Boston, Massachusetts
 |
ABSTRACT
|
---|
Essential (polygenic) hypertension is a complex genetic disorder that remains a major risk factor for cardiovascular disease despite clinical advances, reiterating the need to elucidate molecular genetic mechanisms. Elucidation of susceptibility genes remains a challenge, however. Blood pressure (BP) regulatory pathways through angiotensin II (ANG II) and endothelin-1 (ET-1) receptor systems comprise a priori candidate susceptibility pathways. Here we report that the dual ET-1/ANG II receptor gene (Dear) is structurally and functionally distinct between Dahl salt-sensitive, hypertensive (S) and salt-resistant, normotensive (R) rats. The Dahl S S44/M74 variant is identical to the previously reported Dear cDNA with equivalent affinities for both ET-1 and ANG II, in contrast to Dahl R S44P/M74T variant, which exhibits absent ANG II binding but effective ET-1 binding. The S44P substitution localizes to the ANG II-binding domain predicted by the molecular recognition theory, providing compelling support of this theory. The Dear gene maps to rat chromosome 2 and cosegregates with BP in female F2(RxS) intercross rats with highly significant linkage (LOD 3.61) accounting for 14% of BP variance, but not in male F2(RxS) intercross rats. Altogether, the data suggest the hypothesis that modification of the critical balance between ANG II and ET-1 systems through variant Dear contributes to hypertension susceptibility in female F2(RxS) intercross rats. Further investigations are necessary to corroborate genetic linkage through congenic rat studies, to investigate putative gene interactions, and to show causality by transgenesis and/or intervention. More importantly, the data reiterate the importance of sex-specific factors in hypertension susceptibility.
genetics; angiotensin II; endothelin-1; receptor; hypertension; Dahl rats; Dear
 |
INTRODUCTION
|
---|
THE RENIN-ANGIOTENSIN SYSTEM plays a critical role in blood pressure (BP) homeostasis (29, 39). This system is complex since there are several ANG II receptors (20, 21, 30, 31), with two showing linkage to BP variation (31, 37). Likewise, the endothelin-1 (ET-1) receptor system also plays significant roles in blood pressure and vascular tone regulation and has been implicated in salt-sensitive hypertension, heart failure, atherosclerosis, and pulmonary hypertension (1, 6, 36). Although ANG II and ET-1 are usually investigated separately as two potent BP regulatory systems, the pathogenic significance of their interaction and balance in hypertension is emerging (1, 13, 18). Dear responds equivalently to both ANG II and ET-1 and is coupled to a Ca2+ mobilizing transduction system (31). This receptor is expressed in several organs relevant to the cardiovascular system, including kidney and blood vessels (31), making it a logical candidate for playing a role in hypertension pathogenesis especially since it represents, in a single receptor system, a putative focal point for crosstalk and/or interaction between the ET-1 and ANG II receptor systems upstream to second messenger-mediated crosstalk.
Here we report the identification of functionally distinct Dear gene variants between Dahl S and Dahl R rats, with the S variant cosegregating with high BP in female but not in male F2(RxS) intercross rats.
 |
MATERIALS AND METHODS
|
---|
Characterization of Dahl S and Dahl R Dear cDNAs.
Dahl S and Dahl R Dear cDNAs were obtained by RT-PCR from Dahl S/JRHsd and Dahl R/JRHsd rat kidney poly(A)+ RNAs, respectively (forward primer, 5'-AAG-AAA-GCA-GCA-CCT-TGG-T-3'; reverse primer, 5'-CGT-GGA-CAG-AGA-CCT-TGT-CT-3') and subsequently subcloned into the PT-vector system (Clontech, Palo Alto, CA). Primer sequences were obtained from the previously reported Sprague-Dawley Dear cDNA (GenBank accession no. AY664492). The cDNAs (432 bp) encompassing the entire Dear amino acid coding region were then sequenced on both strands. Six Dahl S and six Dahl R rats were sequenced showing no intra-strain sequence heterogeneity.
Detection of the Dear gene S44P/M74T variant by single-strand conformation polymorphism (SSCP) analysis.
Dahl S/jrHsd, Dahl R/jrHsd, LEW/SsNHsd, WKY/NHsd, SHR/NHsd, and BN/Hsd rats were purchased from Harlan (Indianapolis, IN). SSCP analysis was performed on genomic DNA isolated from the different rat strains essentially as described (34). The SSCP marker was based on a PCR product encompassing nucleotides 27742911 (spanning the S44P substitution) within the amino acid encoding region of the Dear cDNA (forward primer, 5'-GCT-ATG-TAT-CTG-GAC-AGC-AGC-3'; reverse primer, 5'-AGT-GAA-GCA-CAT-GAT-GCA-AGT-3'; product, 137 bp) (31). The SSCP marker was detected by 6% nondenaturing polyacrylamide gel electrophoresis.
Receptor expression and membrane preparation.
The Dahl S and Dahl R Dear cDNAs were subcloned directionally (5' to 3') into the pcDNA (+) expression vector (Invitrogen, Carlsbad, CA) and transiently expressed in COS-1 cells (ATCC). COS-1 cells were transfected with the expression vectors via Lipofectin-mediated gene transfer, and cell membranes were isolated 72 h posttransfection for hormone binding. Rat kidney membranes were prepared essentially as described (15). COS-1 cell membranes were isolated as described (9). Briefly, cells were washed twice in phosphate-buffered saline and homogenized in 10-fold ice-cold buffer (0.25 M sucrose, 1 mM EDTA, 50 µg/ml aprotinin, 10 µg/ml leupeptin, 100 µM phenylmethylsulfonyl fluoride, and 25 mM imidazole-HCl, pH 7.4). The homogenate was centrifuged at 5,000 g for 15 min, and the pellet was discarded. The supernatant was then centrifuged at 27,500 g for 30 min, and the resulting pellet was washed twice in ice-cold suspension buffer (5 mM MgCl2, 0.2 mM EDTA, 50 mM HEPES, pH 7.4). The final pellet was resuspended into the appropriate assay buffer and quickly frozen in liquid nitrogen. The membrane preparations were store at 80°C until use. Protein concentrations of the membranes were determined by BCA protein assay kit (Pierce).
Radioligand binding assays.
Binding of 125I-labeled [Tyr4]angiotensin II and 125I-[Tyr13]endothelin-1 to COS-1 membranes was performed by a rapid filtration method (5, 23). Briefly, 125I-[Tyr4]angiotensin II (0.256.5 nM) or 125I-[Tyr13]endothelin-1 (0.0451.46 nM) was incubated with membranes (100 µg) for 20 min at 37°C in 100 µl buffer A (5 mM MgCl2, 0.2 mM EDTA, 10 mg/ml BSA, and 10 mM HEPES, pH 7.4). Binding reactions were terminated by the addition of 1 ml ice-cold buffer A and immediately filtered through a Whatman GF/C filter (presoaked overnight at 4°C in 10 mg/ml BSA) and subsequently washed with 15 ml ice-cold buffer A. Specific binding was determined as the difference between the total radioactivity bound to membranes and the radioactivity bound to blanks containing 1 µM ANG II or 1 µM ET-1. The dissociation constant (Kd) and maximum ligand-binding sites (Bmax) were determined using Hill plot analysis (28). Hill coefficient values (h) were calculated from the relationship ln[B/(Bmax B)] = hln[free radioligand] lnKd. An F test (P < 0.05) was used to determine whether the saturation binding curves best fitted one or two independent binding sites. The data were best fit by two affinity states determined by Scatchard plot analysis; KH and KL designate the Kd for high- and low-affinity states of the receptor, respectively (17). Most results are expressed as the means ± SE (standard error) from three to five independent experiments.
Western and West-Western blotting analysis.
A polyclonal rabbit antipeptide antibody raised against the synthetic peptide P51LLTSLGSKE60 was utilized for Western blot analysis (31). Plasma membranes (40 µg protein/lane) were subjected to 12.5% SDS-PAGE, and the separated proteins were electro-transferred onto PVDF membranes, which were incubated with blocking buffer (0.3% Tween-20, 5% nonfat milk, 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, and 1.5 mM KH2PO4, pH 7.4) for 2 h at room temperature, and then incubated with primary antibody (1:500) for 16 h at 4°C. The PVDF membranes were then sequentially incubated with biotinylated goat anti-rabbit IgG followed by immunostaining with horseradish peroxidase-linked streptavidin. To confirm the interaction between Dear and ligands, we performed West-Western blot analysis (37). Briefly, protein blots of kidney and COS-1 cell membranes were incubated with radioligands (0.5 µCi in 10 ml) in buffer A at 37°C for 16 h. PVDF membranes were then washed three times for 15 min with buffer A at 37°C and exposed to X-ray film at 80°C for 13 days.
Genetic crosses.
Dahl S/jrHsd and Dahl R/jrHsd rat strains (Harlan, Indianapolis, IN) were used to develop the F2 cohort. The F2 cohort was derived from brother-to-sister mating of F1 (R female x S male) hybrids to produce the F2 male (n = 106, carrying exclusively Y chromosomes from the Dahl S genetic background) and F2 female (n = 102) segregating populations.
Phenotypic characterization of F2 cohorts.
All animal procedures were performed in accordance with institutional guidelines. Animals were maintained on a LabDiet 5001 rodent chow (Harlan Teklad, Madison, WI) containing 0.4% NaCl from weaning until the high-salt diet begun at 12 wk of age. The food pellets and water were made available ad libitum. BP was measured essentially as described (11) using intra-aortic abdominal radiotelemetric implants (Data Science International) obtaining nonstressed BP measurements taking the average over 10 s every 5 min for 24 h (11). Systolic (SBP), diastolic (DBP), and mean arterial pressures (MAP) were obtained along with heart rate and activity. The protocol for the F2 rats was as follows: implant surgery at 10 wk of age; only rats with no postoperative complications were used; after 12 days, baseline BP levels were obtained. High-salt (8% NaCl) challenge was begun at 12 wk of age and maintained for 8 wk for male and 12 wk for female F2 intercross rats; a longer high-salt challenge was necessary for females to attain a similar F2-mean BP, since BP in females is lower. BP values used for phenotype are the averages obtained in the final week of the salt loading from a 24-h recording during a no-entry day ascertaining nonstress BP. We note that baseline BP means for SBP, DBP, and MAP were equivalent, ±1 mmHg range for all three BP parameters among the different Dear genotypes (P
0.5).
Intercross linkage analysis.
Genotyping was done with 10 chromosome 2 microsatellite markers informative for our Dahl (RxS) intercross and one SSCP-based Dear marker (described above). Marker regression and QTL analyses was performed with the Map Manager QTXb17 (MMQTXb17) program (19) using MAP as quantitative trait. MMQTXb17 generates a likelihood ratio statistic (LRS) as a measure of the significance of a possible QTL. Genetic distances were calculated using Kosambi mapping function (genetic distances are expressed in cM). Critical significance values (LRS values) for linkage were determined by a permutation test (2,000 permutations at 10-cM interval) on our male and female progenies using Kosambi mapping function and a free regression model. Thus the minimum LRS values for the F2 male cohort were as follows: for "suggestive linkage" = 4.1 (LOD = 0.89), for "significant linkage" = 10.6 (LOD = 2.30), for "highly significant linkage" = 18.4 (LOD = 4.00). For the F2 female cohort, these were as follows: for suggestive linkage = 3.9 (LOD = 0.85); for significant linkage = 9.9 (LOD = 2.15); for highly significant linkage = 16.6 (LOD = 3.61). LRS 4.6 delineates LOD 1-support interval. Confidence interval for a QTL location was estimated by bootstrap resampling method wherein the histogram single peak delineates the QTL and peak widths define the confidence interval for the QTL. Histograms that show more than one peak warn that the position for the QTL is not well defined or that there may be multiple linked QTLs (QTX Map Manager) (19).
 |
RESULTS AND DISCUSSION
|
---|
The inbred Dahl/JR rat model is an established model of human essential hypertension comprised of a salt-sensitive, hypertensive strain (Dahl S) with its cognate salt-resistant, normotensive control strain (Dahl R) (25). To investigate the possible involvement of Dear in hypertension pathogenesis, we obtained cDNAs spanning the entire amino acid coding region for both Dahl S and Dahl R receptors. Two nucleotide differences were detected resulting in two nonconservative amino acid substitutions: T2814 (Dahl S)/C2814 (Dahl R) nucleotide transition, resulting in S44P substitution; and T2901 (Dahl S)/C2901 (Dahl R) nucleotide transition, resulting in M74T substitution (Fig. 1, A and B). The S44P substitution is located in the putative ANG II binding site in the extracellular domain, whereas the M74T substitution is located in the putative transmembrane domain (Fig. 1B). The Dahl S cDNA nucleotide sequence is identical to the previously reported Sprague-Dawley rat brain Dear cDNA (31). We note that both the Dahl S and Dahl R rat strains were derived from the Sprague-Dawley strain (25).

View larger version (34K):
[in this window]
[in a new window]
|
Fig. 1. Molecular characterization of Dahl salt-sensitive, hypertensive (S) and Dahl salt-resistant, normotensive (R) Dear variants. A: comparative nucleotide sequence of Dahl S and Dahl R cDNAs spanning the T2814 (Dahl S)/C2814 (Dahl R) and T2901 (Dahl S)/C2901 (Dahl R) nucleotide transitions. Amino acid substitutions resulting from the corresponding nucleotide transitions detected S44 substitution in Dahl R Dear for P44 and M74 substitution in Dahl R Dear for T74 [amino acid numbering per Ruiz-Opazo et al. 1998 (31)]. B: schematic structure of the Dahl R Dear. The following functional domains are highlighted: putative ANG II binding site (aa 4148); endothelin-1 (ET-1) binding site (aa 6067); amino acid S44 and M74 substituted in the Dahl S Dear by P44 and T74, respectively; potential cAMP-dependent protein kinase phosphorylation sites (S91, T108 in green); a potential internalization recognition sequence (IRS, in orange). C: Western blot analysis detects equivalent levels of Dahl S (S) and Dahl R (R) Dear variants in Dahl S and Dahl R rat kidney membranes isolated from male and female rats. MW, 14.4-kDa molecular weight marker.
|
|
The S44P substitution spans the predicted ANG II binding domain within the Dahl R Dear variant (31) (Fig. 1B), suggesting the hypothesis that ANG II binding will most likely be different between Dahl S and Dahl R receptors. To examine this hypothesis, we first determined that there is no significant difference in Dear expression levels between Dahl S and Dahl R rats at 12 wk of age in both male and female rats as detected by Western blot analysis comparing Dahl S and Dahl R kidney membranes (Fig. 1C). To examine hormone binding, both Dahl S and Dahl R receptors were transiently expressed in COS-1 cells, respectively, and tested for both ANG II and ET-1 binding. Dahl R Dear do not exhibit ANG II binding but exhibit normal ET-1 binding as shown by direct radioligand binding (Fig. 2 and Table 1) and by West-Western blot analysis [i.e., labeled ligand (West) binding to receptor polypeptide on Western blot] of Dahl S and Dahl R rat kidney membranes and Dahl S and Dahl R Dear COS-1-transfectant cell membranes (Fig. 2C). These data demonstrate that the Dahl S Dear variant is a dual receptor binding both ET-1 and ANG II similar to the brain-derived clone first characterized (31) but that the Dahl R Dear variant responds solely to ET-1 stimulation. Two affinity binding sites for ET-1 are detected in Dahl S and Dahl R receptors (Fig. 3, Table 1), and two affinity binding sites for ANG II are detected in Dahl S receptors (Fig. 3, Table 1) consistent with previous characterization (31). Furthermore, compared with the Dahl R Dear S44P/M74S variant, the Dahl S Dear S44/M74 variant exhibits threefold increased affinity for ET-1 (for Dahl R, S44P/M74T KH ET-1 = 12.0 ± 1.12 pM; for Dahl S, S44/M74 KH ET-1 = 4.42 ± 0.89 pM, P < 0.001; Table 1), suggesting an enhanced response of the Dahl S receptor to ET-1 stimulation compared with the Dahl R receptor.

View larger version (23K):
[in this window]
[in a new window]
|
Fig. 2. Functional characterization of Dahl S and Dahl R Dear variants. Saturation binding curves of ligand binding studies of Dahl S ( ) and Dahl R () Dear expressed in COS-1 cells with radiolabeled 125I-ANG II (A) and 125I-ET-1 (B). Values are presented as mean ± SD from five independent experiments. C: detection of the 14-kDa Dear protein (arrowheads on right) by Western blot analysis (ab) of Dahl R (Kid-R) and Dahl S (Kid-S) kidney (Kid) membranes; control nontransfected COS-1 cell membranes (Cos1, c), COS-1 cell membranes expressing the Dahl R S44P/M74T variant (Cos1, R), and COS-1 cell membranes expressing the Dahl S S44/M74 variant (Cos1, S). 125I-ANG II West-Western blot analysis (*ANG II) detects binding only to Dahl S kidney membranes (Kid, S) and COS-1 cell membranes expressing the Dahl S S44/M74 variant (Cos1, S), whereas 125I-ET-1 West-Western blot analysis (*ET-1) reveals binding to both Dahl R (Kid, R) and Dahl S (Kid, S) kidney membranes as well as to COS-1 cell membranes expressing the Dahl R S44P/M74T (Cos1, R) and Dahl S S44/M74 (Cos1, S) molecular variants.
|
|

View larger version (16K):
[in this window]
[in a new window]
|
Fig. 3. Scatchard plots of saturation data for Dahl S and Dahl R Dear variants. Scatchard plots of 125I-ANG II (A) and 125I-ET-1 (B and C) saturation binding data of Dahl S ( ) and Dahl R () Dear expressed in COS-1 cells.
|
|
Based on its localization to the predicted ANG II-binding site, it is likely that the S44P substitution accounts for the observed absent ANG II binding in Dahl R Dear, although we cannot rule out the possibility that the M74T substitution could also affect receptor function as well. Interestingly, this S44P substitution and resultant differential ANG II binding elucidates for the first time a natural occurring mutation within a peptide-ligand binding domain predicted by the molecular recognition theory (31). This provides compelling validation of this still controversial theory in explaining the evolution of intermolecular and intramolecular interaction of peptides and proteins which are thought to have evolved from complementary strands of genomic DNA (3, 30).
Having found functionally significant variants between Dahl S and Dahl R Dear genes, we then investigated the potential genetic contribution to hypertension susceptibility by performing independent QTL analysis on both male (n = 106) and female (n = 102) F2(RxS) intercross rats phenotyped for BP by radiotelemetry after 8 and 12 wk of high-salt (8% NaCl) challenge, respectively (Table 2). The high-salt diet challenge was extended 4 wk longer for the F2(RxS) intercross female rats, since female BP phenotype was much lower than in male F2(RxS) intercross rats [average F2(RxS) intercross male SBP after 8 wk of high-salt diet = 157.2 ± 14.2 mmHg; average F2(RxS) intercross female SBP after 12 wk of high-salt diet = 145.0 ± 11.4 mmHg, Table 2]. Using an SSCP-based Dear gene-specific marker (Fig. 4A), we mapped the rat Dear to chromosome 2 (physical position in the current assembly of the rat genome: 2q34, 176.687 Mb), 4.5 cM centromeric to the
1-Na-K-ATPase locus, ATP1A1. A total chromosome 2 scan was then done with 11 informative markers that distinguish Dahl R and Dahl S strains. Marker regression and interval mapping analyses detect a single chromosomal region with suggestive linkage to MAP, SBP, and DBP, peaking at ATP1A1 + 2 cM (LOD = 1.70) in the F2(RxS) male cohort (Fig. 4B, Table 2). In contrast, chromosome 2 scan analysis of F2(RxS) intercross females revealed two QTLs on chromosome 2, one centered at D2Rat143 (LOD = 2.43, significant linkage) and the other centered at Dear 5 cM (LOD = 3.61, highly significant linkage) (Fig. 4C, Table 2). To assess pathophysiological relevance, analysis of Dear allele-specific contribution (Table 3) reveals that the Dahl S S44/M74 variant increases susceptibility to hypertension regardless of BP parameter: systolic, diastolic, or mean arterial pressure, with greatest changes in mean arterial pressure (ANOVA P < 103 to 104). Concordance of results in different BP parameters provide compelling evidence delineating the Dear locus as a candidate gene for hypertension susceptibility in F2(RxS) intercross female rats.

View larger version (24K):
[in this window]
[in a new window]
|
Fig. 4. Detection and genetic analysis of Dear variants. A: detection of the Dear gene variants by single-strand conformation polymorphism (SSCP) analysis in different rat strains. A 137-bp PCR product spanning the S44P substitution reveals the S44P/M74T variant in Dahl R (R) and LEW strains, whereas the S44/M74 variant is detected in Dahl S (S), BN, WKY, and SHR genomic DNAs. F1 denotes F1(RxS) subjects. Interval mapping with bootstrap analysis for chromosome 2 in male (B) and female (C) cohorts using Map Manager QTXb17 program. Horizontal lines mark likelihood ratio statistic (LRS) values for significance of linkage. For B (from top to bottom): line 1, LRS = 18.4 (LOD = 4.00) for highly significant; line 2, LRS = 10.6 (LOD = 2.30) for significant; and line 3, LRS = 4.1 (LOD = 0.89) for suggestive. For C (from top to bottom): line 1, LRS = 16.6 (LOD = 3.61) for highly significant; line 2, LRS = 9.9 (LOD = 2.15) for significant; and line 3, LRS = 3.9 (LOD = 0.85) for suggestive. Line a, regression coefficient for additive effect; line b, LRS; line c, regression coefficient for dominance effect. Histograms represent the bootstrap-based confidence intervals for the detected QTLs.
|
|
Although further QTL dissection by congenic and gene-interaction analyses remains to be done in F2(RxS) intercross female rats, we note that to date, only Dear (shown here) and ATP1A1 (11, 16) genes exhibit functionally significant variants between Dahl S and Dahl R rats with demonstrated pathogenic relevance to hypertension. These data implicate the above-mentioned two loci as candidate genes for the Dear 5 cM QTL region on chromosome 2 affecting BP in female F2(RxS) intercross rats based on a four-parameter analysis framework for hypertension genes (10, 27). Although causality needs to be demonstrated by transgenesis for Dear, the causal role of ATP1A1 in hypertension has been shown by transgenesis in both male and female Dahl S rats (11). Although putative gene interactions need to be investigated for Dear, ATP1A1-Na-K-2Cl cotransporter (NKCC2) gene interaction has been detected to increase susceptibility to high BP in cosegregation analysis of F2(RxS) intercross rats (12). This epistatic nature of the ATP1A1 effect on BP could account for the reduced statistical significance of linkage to BP detected at ATP1A1 when analyzed as a single locus as done in this chromosome 2 scan and in previous F2 intercross and congenic studies (26, 40).
To further analyze the Dear locus in the context of previous genetic rat model studies which report chromosome 2 QTLs for BP that span the Dear locus (4, 7, 14, 24, 33), we assessed Dear variants on WKY, SHR, BN, and LEW rat strains used in said studies by SSCP analysis. As shown in Fig. 4A, the Dahl R Dear S44P/M74T variant is detected in Dahl R and LEW strains, whereas the Dahl S Dear S44/M74 variant is detected in SHR, WKY, and BN strains. Detection of variant-specific alleles in the different strains was corroborated by direct nucleotide sequencing of two independent Dear cDNA clones spanning the entire amino acid coding region (data not shown). Since SHR, WKY, and BN rat strains have the S44/M74 Dahl S Dear allele, the Dear locus is a priori eliminated as a candidate gene for the chromosome 2 QTL for BP in F2 intercross studies derived from these strains (4, 14, 24, 33). The Dear locus is also eliminated as candidate gene for the reported chromosome 2 BP QTL in F2(Dahl S x LEW) intercross study, which investigated males only (7), since the Dear S variant cosegregates with high BP in females.
The detection of sex-specific QTLs for BP is not new. This has been reported in other studies demonstrating BP QTLs on chromosomes 2 and 3 in males but not in females in an F2(SHRSP x WKY) intercross (4). Altogether, these studies along with our current data reiterate the need for sex-specific analyses. Pathophysiological sex-specific effects of ET-1 have been reported (2, 8, 22, 35), consistent with our observations.
In summary, observations from genetic, molecular, and pathophysiological analyses suggest the hypothesis that modification in the balance of ANG II and ET-1 receptor systems through variant Dear contributes to hypertension susceptibility in female F2(RxS) intercross rats. However, further investigations are necessary to corroborate genetic linkage through congenic rat studies, to investigate putative gene interactions, and to show causality by transgenic and/or intervention approaches. Nevertheless, the data reiterate the importance of sex-specific factors in hypertension susceptibility and the role of Dear in ANG II-ET-1 response balance.
 |
GRANTS
|
---|
This work was supported by National Heart, Lung, and Blood Institute Grant HL-69937.
 |
ACKNOWLEDGMENTS
|
---|
The chromosome 2 scan was done through the Marshfield Genotyping Service.
 |
FOOTNOTES
|
---|
Article published online before print. See web site for date of publication (http://physiolgenomics.physiology.org).
Address for reprint requests and other correspondence: N. Ruiz-Opazo, Section Molecular Medicine, W609, Boston Univ. School of Medicine, 700 Albany St., Boston, MA 02118 (E-mail: nruizo{at}bu.edu).
10.1152/physiolgenomics.00108.2004.
 |
REFERENCES
|
---|
- Agapitov AV and Haynes WG. Role of endothelin in cardiovascular disease. J Renin Angiotensin Aldosterone Syst 3: 115, 2002.[ISI][Medline]
- Antoniucci D, Miller VM, Sieck GC, and Fitzpatrick LA. Gender-related differences in proliferative responses of vascular smooth muscle cells to endothelin-1. Endothelium 8: 137145, 2001.[Medline]
- Blalock JE. Genetic origins of protein shape and interaction rules. Nat Med 1: 876878, 1995.[ISI][Medline]
- Clark JS, Jeffs B, Davidson AO, Lee WK, Anderson NH, Bihoreau MT, Brosnan MJ, Devlin AM, Kelman AW, Lindpaintner K, and Dominiczak AF. Quantitative trait loci in genetically hypertensive rats, possible sex specificity. Hypertension 28: 898906, 1996.[Abstract/Free Full Text]
- Doi T, Hiroaki Y, Arimoto I, Fujiyoshi Y, Okamoto T, Satoh M, and Furuichi Y. Characterization of human endothelin B receptor and mutant receptors expressed in insect cells. Eur J Biochem 248: 139148, 1997.[Abstract]
- Elijovich F and Laffer CL. Participation of renal and circulating endothelin in salt-sensitive essential hypertension. J Hum Hypertens 16: 459467, 2002.[CrossRef][ISI][Medline]
- Garrett MR, Dene H, Walder R, Zhang QY, Cicila GT, Assadnia S, Deng AY, and Rapp JP. Genome scan and congenic strains for blood pressure QTL using Dahl Salt sensitive rats. Genome Res 8: 711723, 1998.[Abstract/Free Full Text]
- Hallberg P, Karlsson J, Lind L, Michaelsson K, Kurland L, Kahan T, Malmqvist K, Ohman KP, Nystrom F, Liljedahl U, Syvanen AC, and Melhus H. Gender-specific association between preproendothelin-1 genotype and reduction of systolic blood pressure during antihypertensive treatment-results from the Swedish Irbersartan Left Ventricular Hypertrophy Investigation versus Atenolol (SILVHIA). Clin Cardiol 27: 287290, 2004.[ISI][Medline]
- Hausdorff WP, Hnatowich M, ODowd BF, Caron MG, and Lefkowitz RJ. A mutation of b2-adrenergic receptor impairs agonist activation of adenylyl cyclase without affecting high affinity agonist binding. Distinct molecular determinants of the receptor are involved in physical coupling to and functional activation of GS. J Biol Chem 265: 13881393, 1990.[Abstract/Free Full Text]
- Herrera VLM and Ruiz-Opazo N. Genetics of hypertension: a multidisciplinary challenge. Trends Cardiovasc Med 1: 185189, 1991.[CrossRef][ISI]
- Herrera VLM, Xiang XH, Lopez LV, Schork NJ, and Ruiz-Opazo N. The
1 Na,K-ATPase gene is a susceptibility hypertension gene in the Dahl salt-sensitive rat. J Clin Invest 102: 11021111, 1998.[Abstract/Free Full Text]
- Herrera VLM, Lopez LV, and Ruiz-Opazo N.
1 Na,K-ATPase and Na,K,2Cl-cotransporter/D3Mit3 loci interact to increase susceptibility to salt-sensitive hypertension in Dahl SHSD rats. Mol Med 7: 125134, 2001.[ISI][Medline]
- Iwanaga Y, Kihara Y, Inagaki K, Onozawa Y, Yoneda T, Kataoka K, and Sasayama S. Differential effects of angiotensin II versus endothelin-1 inhibitions in hypertrophic left ventricular myocardium during transition to heart failure. Circulation 104: 606612, 2001.[Abstract/Free Full Text]
- Jeffs B, Negrin CD, Graham D, Clark JS, Anderson NH, Gauguier D, and Dominiczak AF. Applicability of a "speed" congenic strategy to dissect blood pressure quantitative trait loci on rat chromosome 2. Hypertension 35: 179187, 2000.[Abstract/Free Full Text]
- Jørgensen PL. Purification (Na+ plus K+)-ATPase: active site determinations and criteria of purity. Ann NY Acad Sci 242: 3652, 1974.[ISI][Medline]
- Kaneko Y, Cloix JF, Herrera VLM, and Ruiz-Opazo N. Corroboration of Dahl S Q276L alpha1-Na,K-ATPase protein sequence: impact on affinities for ligands and on E1 conformation. J Hypertens. In press.
- Kent RS, De Lean A, and Lefkowitz RJ. A quantitative analysis of beta-adrenergic receptor interactions: resolution of high and low affinity states of the receptor by computer modeling of ligand binding data. Mol Pharmacol 17: 1423, 1980.[Abstract]
- Lavallee M, Takamura M, Parent R, and Thorin E. Crosstalk between endothelin and nitric oxide in the control of vascular tone. Heart Fail Rev 6: 265276, 2001.[CrossRef][Medline]
- Manly KF, Cudmore RH Jr, and Meer JM. Map Manager QTX, cross-platform software for genetic mapping. Mamm Genome 12: 930932, 2001.[CrossRef][ISI][Medline]
- Mukoyama M, Nakajima M, Horiuchi M, Sasamura H, Pratt RE, and Dzau VJ. Expression cloning of type 2 angiotensin II receptor reveals a unique class of seven transmembrane receptors. J Biol Chem 268: 2453924542, 1993.[Abstract/Free Full Text]
- Murphy TJ, Alexander RW, Griendling KK, Runge MS, and Bernstein KE. Isolation of a cDNA encoding the vascular type-1 angiotensin II receptor. Nature 351: 233236, 1991.[CrossRef][ISI][Medline]
- Nuedling S, van Eickels M, Allera A, Doevendans P, Meyer R, Vetter H, and Grohe C. 17 Beta-estradiol regulates the expression of endothelin receptor type B in the heart. Br J Pharmacol 140: 195201, 2003.[Abstract/Free Full Text]
- Phalipou S, Seyer R, Cotte N, Breton C, Barberis C, Hibert M, and Mouillac B. Docking of linear peptide antagonists into the human V1a vasopressin receptor. J Biol Chem 274: 2331623327, 1999.[Abstract/Free Full Text]
- Pravenec M, Gauguier D, Schott JJ, Buard J, Kren V, Bila V, Szpirer C, Szpirer J, Wang JM, Huanng H, St Lezin E, Spence MA, Flodman P, Printz M, Lathrop GM, Vergnaud G, and Kurtz TW. Mapping of quantitative trait loci for blood pressure and cardiac mass in the rat by genome scanning of recombinant inbred strains. J Clin Invest 96: 19731978, 1995.[ISI][Medline]
- Rapp JP and Dene H. Development and characteristics of inbred strains of Dahl salt-sensitive and salt-resistant rats. Hypertension 7: 340349, 1985.[Abstract]
- Rapp JP and Dene H. Failure of alleles at the Na+,K+-ATPase
1 locus to cosegregate with blood pressure in Dahl rats. J Hypertens 8: 457462, 1990.[ISI][Medline]
- Rapp JP. Genetic analysis of inherited hypertension in the rat. Physiol Rev 80: 135172, 2000.[Abstract/Free Full Text]
- Rodbard D. Mathematics of hormone-receptor interaction. I. Basic principles. Adv Exp Med Biol 36: 289326, 1973.[Medline]
- Romero JC and Reckelhoff JF. Role of angiotensin and oxidative stress in essential hypertension. Hypertension 34: 943949, 1999.[Abstract/Free Full Text]
- Ruiz-Opazo N, Akimoto K, and Herrera VLM. Identification of a novel dual angiotensin II/vasopressin receptor on the basis of molecular recognition theory. Nat Med 1: 10741081, 1995.[ISI][Medline]
- Ruiz-Opazo N, Hirayama K, Akimoto K, and Herrera VLM. Molecular characterization of a dual endothelin-1/angiotensin II receptor. Mol Med 4: 96108, 1998.[ISI][Medline]
- Ruiz-Opazo N, Lopez LV, and Herrera VLM. The dual AngII/AVP receptor gene N119S/C163R variant exhibits sodium-induced dysfunction and cosegregates with salt-sensitive hypertension in the Dahl salt-sensitive hypertensive rat model. Mol Med 8: 2432, 2002.[CrossRef][ISI][Medline]
- Samani NJ, Gauguier D, Vincent M, Kaiser MA, Bihoreau MT, Lodwick D, Wallis R, Parent V, Kimber P, Rattray F, Thompson JR, Sassard J, and Lathrop M. Analysis of quantitative trait loci for blood pressure on rat chromosomes 2 and 13. Age-related differences in effect. Hypertension 28: 11181122, 1996.[Abstract/Free Full Text]
- Song Y, Herrera VLM, Filigheddu F, Troffa C, Lopez LV, Glorioso N, and Ruiz-Opazo N. Non-association of the thiazide-sensitive Na,Cl-cotransporter gene with polygenic hypertension in both rats and humans. J Hypertens 19: 15471551, 2001.[CrossRef][ISI][Medline]
- Tatchum-Talom R, Martel C, Labrie C, Labrie F, and Marette A. Gender differences in hemodynamic responses to endothelin-1. J Cardiovasc Pharmacol 36: S102S104, 2000.[ISI][Medline]
- Touyz RM and Schiffrin EL. Role of endothelin in human hypertension. Can J Physiol Pharmacol 81: 533541, 2003.[CrossRef][ISI][Medline]
- Tsukamoto T, Shibagaki Y, Imajoh-Ohmi S, Murakoshi T, Suzuki M, Nakamura A, Gotoh H, and Mizumoto K. Isolation and characterization of the yeast mRNA capping enzyme b subunit gene encoding RNA 5'-triphosphatase, which is essential for cell viability. Biochem Biophys Res Commun 239: 116122, 1997.[CrossRef][ISI][Medline]
- Wong C, Mahapatra NR, Chitbangonsyn S, Mahboubi P, Mahata M, Mahata SK, and OConnor DT. The angiotensin II receptor (Agtr1a): functional regulatory polymorphisms in a locus genetically linked to blood pressure variation in the mouse. Physiol Genomics 14: 8393, 2003. First published April 15, 2003; doi:10.1152/physiolgenomics.00162.2002.[Abstract/Free Full Text]
- Wright JW and Harding JW. Regulatory role of brain angiotensins in the control of physiological and behavioral responses. Brain Res Rev 17: 227262, 1992.[ISI][Medline]
- Zicha J, Negrin CD, Dobesova Z, Carr F, Vokurkova M, McBride MW, Kunes J, and Dominiczak AF. Altered Na+-K+ pump activity and plasma lipids in salt-hypertensive Dahl rats: relationship to Atp1a1 gene. Physiol Genomics 6: 99104, 2001.[Abstract/Free Full Text]