From the Department of Physiology and Molecular Medicine, Medical College
of Ohio, Toledo, Ohio.
Correspondence to John P. Rapp, DVM, PhD, Department of Physiology and Molecular Medicine, Medical College of Ohio, Block Health Science Bldg, 3035 Arlington Ave, Toledo, OH 43614-5804.
Iwai and Inagami1 confirmed that the
Sa gene was differentially expressed in SHR and WKY kidneys
by Northern blot analysis, with SHR expressing markedly more
than WKY. They also showed that Dahl salt-sensitive (S) rats expressed
the Sa gene more than Dahl salt-resistant (R)
rats.1
Although the technique of differential hybridization is a standard one,
its application to hypertensive rat models was innovative, and the
Sa gene has generated a lot of interest. Important
subsequent results were that (1) the Sa gene is expressed in
the kidney proximal tubule,2 3 (2) the
Sa gene is located on rat chromosome 1 and cosegregates with
blood pressure (BP) in both the SHR/WKY4 5 6 7 and
Dahl rat models,8 9 10 and (3) polymorphisms
in the Sa gene and differential expression of the
Sa gene cosegregate.7
The linkage of a candidate gene such as the Sa gene to BP
certainly does not prove that the candidate is the causative factor.
Such linkage is necessary but not sufficient. Linkage analysis
for a quantitative trait such as BP in
theory11 12 and in practice yields only a very
broad segment of chromosome that is likely to include the causative
gene (or genes). This region is typically 20 to 40 centimorgans and
contains hundreds of genes. To be sure that in fact a BP quantitative
trait locus (QTL) exists in the region of interest, the next logical
step is the construction of congenic
strains.13 14 The congenic strain also provides a
vehicle for subsequent fine genetic mapping of the QTL.
In forming a congenic strain, a segment of chromosome is moved from one
inbred rat strain (donor) to another strain (recipient) by a standard
protocol involving a series of back-cross breeding to the recipient
strain with selection for donor alleles at marker loci along the
chromosomal segment to be moved.13 This results
in a pair of strains (the congenic and the recipient strain) that are
(essentially) genetically identical except that the congenic strain has
a defined chromosomal segment from the donor. If the congenic region in
fact contains a variant BP QTL allele compared with the recipient
strain, then the 2 strains will have different BP, demonstrating the
existence of a BP effect of the chromosomal region that was moved.
In this issue of Hypertension, Iwai et
al15 and Frantz et al16
report the construction of congenic strains involving SHR and WKY
chromosomal regions containing the Sa gene. In both cases,
the congenic strains demonstrated the existence of a BP QTL in the
general vicinity of the Sa gene. This work confirms and
extends the results from a previous congenic strain on rat chromosome 1
using SHR and WKY17 ; an additional report on a
chromosome 1 congenic strain also demonstrates a BP QTL using Dahl S
and Lewis rats.10 Thus, there is an impressive
consistency in these results.
Although the function of the Sa gene product is unknown,
the Sa gene has passed several important tests as a
hypertension candidate gene: (1) it has an interesting intermediate
phenotype (differential expression between strains in the
proximal tubule); (2) it cosegregates with BP; and (3) it is contained
in a congenic region influencing BP. An important remaining step is to
make congenic strains in which the alleles at the Sa
locus are moved with reduced (preferably minimal) flanking DNA to
reduce the chance that the real QTL is a locus linked to the
Sa locus. This procedure has been applied to the loci for
renin and the inducible form of nitric oxide synthase; in both cases,
the candidate gene was eliminated as causative for the BP difference
between the specific strains used.18 19 Thus,
some caution is appropriate until fine genetic mapping of the QTL in
the region of the Sa gene is accomplished by construction of
congenic substrains.
Other data suggest that the Sa gene is not responsible for
BP differences. The Milan hypertensive strain (MHS) carries the same
Sa allele as the normotensive WKY, and the Milan
normotensive strain (MNS) carries the same allele as
SHR.20 This conclusion was based on the
restriction fragment length patterns of 15 enzymes that yield a
polymorphic pattern between the 2 alleles. Also, MNS express a
high level of renal Sa mRNA compared with MHS, as would be
expected based on the allele in each strain. This reversal of
alleles compared with what might be anticipated based on the
SHR/WKY and Dahl S/Dahl R comparison in itself has no bearing on
interpretation of the linkage and congenic strain data. It is expected
theoretically, and found experimentally, that some QTL alleles for
increase of a quantitative trait will be carried by a low strain and
visa versa; some examples are found in work by Garrett et
al.10 A high or low strain is determined by the
net effect over multiple loci, so alleles at any single locus do
not determine the ultimate phenotype. Much more insightful is
the observation20 in an F2
population derived from MHS and MNS that alleles at the
Sa locus did not cosegregate with BP. This does imply that
the Sa alleles that control the large difference in
renal expression have nothing to do with BP and that the QTL on
chromosome 1 does not have functionally variant alleles in the MHS
versus MNS comparison. One could also argue (less convincingly in my
view) that ancillary factors (epistatic interaction with the genetic
background, environmental factors such as normal dietary NaCl) blunted
the response of BP to the Sa alleles in the
F2 (MHSxMNS) study.
Animal work with the Sa gene has stimulated work in humans;
the Sa gene is located on human chromosome
16p13.11.21 22 In 1
study,23 there was a difference in allelic
frequency of Sa alleles between hypertensive and control
groups. In 3 other such association
studies,24 25 26 no relationship was found; linkage
analysis in humans was also negative.26
Thus, the studies on humans fail to provide convincing evidence for an
effect of the Sa locus on BP.
In summary, the work of Iwai and Inagami1 and the
subsequent work it generated is science at its best, regardless of the
ultimate status of the Sa gene as one of "the" genes
causing BP variation. At a very minimum, the Sa gene as a
candidate led to the detection and confirmation of a BP QTL on rat
chromosome 1. Fine genetic mapping in the animal models using
derivatives of the new congenic strains will certainly determine the
status of the Sa gene and provide a better QTL localization
on rat chromosome 1 for guiding analyses on human chromosome
16.
2.
Patel HR, Thiara AS, West KP, Lodwick D, Samani
NJ. Increased expression of the SA gene in
the kidney of the spontaneously hypertensive rat is localized to the
proximal tubule. J Hypertens. 1994;12:1347–1352.[Medline]
[Order article via Infotrieve]
3.
Yang T, Hassan SA, Singh I, Smart A, Brosius FC,
Holzman LB, Schnermann JB, Briggs JP. SA gene expression in
the proximal tubule of normotensive and hypertensive rats.
Hypertension. 1996;27(pt 2):541–545.
4.
Iwai N, Inagami T. Identification of a candidate gene
responsible for the high blood pressure of spontaneously hypertensive
rats. J Hypertens. 1992;10:1155–1157.[Medline]
[Order article via Infotrieve]
5.
Iwai N, Kurtz TW, Inagami T. Further evidence of the
SA gene as a candidate gene contributing to the hypertension in
spontaneously hypertensive rat. Biochem Biophys Res Commun. 1992;188:64–69.[Medline]
[Order article via Infotrieve]
6.
Lindpaintner K, Hilbert P, Ganten D, Nadal-Ginard B,
Inagami T, Iwai N. Molecular genetics of the
SA-gene: cosegregation with hypertension and
mapping to rat chromosome 1. J Hypertens. 1993;11:19–23.[Medline]
[Order article via Infotrieve]
7.
Samani NJ, Lodwick D, Vincent M, Dubay C, Kaiser MA,
Kelly MP, Lo M, Harris J, Sassard J, Lathrop M, Swales JD. A gene
differentially expressed in the kidney of the spontaneously
hypertensive rat cosegregates with increased blood pressure.
J Clin Invest. 1993;92:1099–1103.
8.
Harris EL, Dene H, Rapp JP. SA
gene and blood pressure cosegregation using Dahl salt-sensitive rats.
Am J Hypertens. 1993;6:330–334.[Medline]
[Order article via Infotrieve]
9.
Gu L, Dene H, Deng AY, Hoebee B, Bihoreau M-T, James
M, Rapp JP. Genetic mapping of two blood pressure quantitative trait
loci on rat chromosome 1. J Clin Invest. 1996;97:777–788.[Medline]
[Order article via Infotrieve]
10.
Garrett MR, Dene H, Walder R, Zhang Q-Y, Cicila GT,
Assadnia S, Deng AY, Rapp JP. Genome scan and congenic strains for
blood pressure QTL using Dahl salt-sensitive rats. Genome
Res.. 1998;8:711–723.
11.
van Ooijen JW. Accuracy of mapping quantitative trait
loci in autogamous species. Theor Appl Genet. 1992;84:803–811.
12.
Hyne V, Kearsey MJ, Pike DJ, Snape JW. QTL
analysis: unreliability and bias in estimation procedures.
Mol Breeding. 1995;1:273–282.
13.
Rapp JP, Deng AY. Detection and positional cloning of
blood pressure quantitative trait loci: is it possible?
Hypertension. 1995;25:1121–1128.
14.
Lander ES, Schork NJ. Genetic dissection of
complex traits. Science. 1994;265:2037–2048.
15.
Iwai N, Tsujita Y, Kinoshita M. Isolation of a
chromosome 1 region that contributes to high blood pressure and salt
sensitivity. Hypertension. 1998;32:636–638.
16.
Frantz SA, Kaiser M, Gardiner SM, Gauguier D, Vincent
M, Thompson JR, Bennett T, Samani NJ. Successful isolation of a rat
chromosome 1 blood pressure quantitative trait locus in reciprocal
congenic strains. Hypertension. 1998;32:639–646.
17.
St Lezin E, Liu W, Wang J-M, Wang N, Kren V, Krenova D,
Musilova A, Zdobinska M, Zidek V, Lau D, Pravenec M. Genetic isolation
of a chromosome 1 region affecting blood pressure in the spontaneously
hypertensive rat. Hypertension. 1997;30:854–859.
18.
Zhang QY, Dene H, Deng AY, Garrett MR, Jacob HJ, Rapp
JP. Interval mapping and congenic strains for a blood pressure QTL on
rat chromosome 13. Mamm Genome. 1997;8:636–641.[Medline]
[Order article via Infotrieve]
19.
Dukhanina OI, Dene H, Deng AY, Choi CR, Hoebee B, Rapp
JP. Linkage map and congenic strains to localize blood pressure QTL on
rat chromosome 10. Mamm Genome. 1997;8:229–235.[Medline]
[Order article via Infotrieve]
20.
Lodwick D, Zagato L, Kaiser MA, Torielli L, Casari G,
Bianchi G, Samani NJ. Genetic analysis of the
SA and
Na+/K+-ATPase
21.
Szpirer C, Riviere M, Szpirer J, Levan G, Guo DF, Iwai
N, Inagami T. Chromosomal assignment of human and rat hypertension
candidate genes: type 1 angiotensin II receptor genes and
the SA gene. J Hypertens. 1993;11:919–925.[Medline]
[Order article via Infotrieve]
22.
Samani NJ, Whitmore SA, Kaiser MA, Harris J, See CG,
Callen DF, Lodwick D. Chromosomal assignment of the human
SA gene to 16p13.11 and demonstration of its
expression in the kidney. Biochem Biophys Res Commun. 1994;199:862–868.[Medline]
[Order article via Infotrieve]
23.
Iwai N, Ohmichi N, Hanai K, Nakamura Y, Kinoshita M.
Human SA gene locus as a candidate locus for essential hypertension.
Hypertension. 1994;23:375–380.
24.
Harrap SB, Samani NJ, Lodwick D, Connor JM, Fraser R,
Davies DL, Lever AF, Foy CJW, Watt GCM. The SA
gene: predisposition to hypertension and renal function in man.
Clin Sci. 1995;88:665–670.[Medline]
[Order article via Infotrieve]
25.
Zee RY, Stephen AL, Iwai N, Morris BJ. Association
analyses of S(A) gene variant in essential hypertensives.
Am J Hypertens. 1997;10:235–242.[Medline]
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26.
Nabika T, Bonnardeaux A, James M, Julier C, Jeunemaitre
X, Corvol P, Lathrop M, Soubrier F. Evaluation of the SA locus in human
hypertension. Hypertension. 1995;25:6–13.
© 1998 American Heart Association, Inc.
Scientific Contributions
Editorial Commentary: The Sa Gene
What Does It Mean?
Key Words: Editorial • hypertension, genetic • genes • congenic strains • rats, inbred SHR
Introduction
Top
Introduction
References
The Sa gene was first described in 1991
by Iwai and Inagami1 in a study to identify genes
that were differentially expressed in the kidneys of spontaneously
hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). To find such genes
they first prepared a cDNA library from 16-week-old SHR. The clones
were grown on plates, and 4 replica filter lifts were prepared from
each plate. Two filters were hybridized to
32P-labeled single-strand antisense cDNA
fragments from kidneys of 16-week-old SHR, and 2 filters were
hybridized with a similar probe prepared from WKY rat kidneys. Clones
that gave a different intensity of autoradiographic signal
with the SHR and WKY probes were selected for study; one of these was
the Sa gene. The designation Sa is apparently
arbitrary.
Footnotes
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
References
Top
Introduction
References
1.
Iwai N, Inagami T. Isolation of preferentially
expressed genes in the kidneys of hypertensive rats.
Hypertension. 1991;17:161–169.
1 genes in the Milan hypertensive rat.
J Hypertens. 1998;16:139–144.[Medline]
[Order article via Infotrieve]
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