Lack of association between polymorphism of the
human cyclic GMP-dependent protein kinase gene and
SO Zakharkin1, AT Belay2, JR Fernandez1,3, V De Luca4, JL Kennedy4, MB Sokolowski2and
1Section on Statistical Genetics, Department of Biostatistics, University of Alabama at Birmingham, Birmingham, AL, USA;
2Department of Biology, University of Toronto at Mississauga, Mississauga, Ontario, Canada;3Clinical Nutrition Research
Center, Department of Nutrition Sciences, University of Alabama at Birmingham, Birmingham, AL, USA; and
4Neurogenetics Section, Centre for Addiction and Mental Health, Department of Psychiatry, University of Toronto, Toronto,
OBJECTIVE: To investigate whether genetic variation in the cyclic GMP-dependent protein kinase gene (PRKG1) is associated
METHODS: The study included 143 individuals from New York City area, NY, USA. The subjects were sampled on the basis of
body mass index (BMI): obese (BMI ranging from 33.8 to 89.5kg/m2), and nonobese (BMI ranging from 16.0 to 29.4kg/m2).
The association between C2276T polymorphism in PRKG1 gene and obesity was tested using linear regression analysis.
RESULTS: BMI levels were predicted by linear regression models adjusted for demographic factors. An analysis was performed
twice: in individuals of all ethnical backgrounds and in European-Americans only. In both cases, genotype did not have a
CONCLUSION: We found no evidence that the C2276T polymorphism in the PKRG1 gene is associated with obesity.
International Journal of Obesity (2005) 29, 872–874. doi:10.1038/sj.ijo.0802973
Published online 10 May 2005
Keywords: association study; polymorphism; cGMP-dependent protein kinase; PRKG1
Obesity is a major health problem worldwide1influenced by
both genetic and environmental factors. Association studies
represent an important paradigm for investigation of com-
plex traits, such as obesity. Using this approach, many
candidate genes including b2- and b3-adrenergic receptors,
leptin receptor, glucocorticoid receptor, and tumor necrosis
factor have been assessed for association with obesity.2The
cyclic GMP-dependent protein kinase (PKG) gene could be
another candidate. PKG is a transduction pathway enzyme
and operates in a variety of cell responses.3Allelic variation
in the homologs of this gene has been linked to differences
in food-related behaviors in fruitflies,4honey bees5and
nematodes.6In Drosophila melanogaster, the product of the
for gene, encoding PKG, might be a component of the
Neuropeptide F (a homolog of human Neuropeptide Y
(NPY)) signaling pathway, which is involved in regulation
of food-related behaviors.7Moreover, PKG proteins are
highly phylogenetically conserved.8Here, we tested relation-
ships between a polymorphism of the PRKG1 gene, encoding
cyclic GMP-dependent protein kinase, and obesity.
The genetic polymorphism in PRKG1 gene was assayed in
143 individuals from the New York City area. The study was
approved by the IRB at the St Luke’s-Roosevelt Institute for
Health Sciences. Written informed consent was obtained
from all subjects. Genomic DNA was extracted as previously
described.9The C2276T polymorphism is located in the
30UTR of the human PRKG1 gene (GenBank accession
number NM_006258). A 236bp fragment (positions 2129–
2364) encompassing polymorphic position 2276 was ampli-
fied using PCR as previously described9with the following
Received 22 June 2004; revised 7 January 2005; accepted 25 February
2005; published online 10 May 2005
*Correspondence: Dr DB Allison, Department of Biostatistics, 327 Ryals
Public Health Building, 1665 University Boulevard, University of Alabama
at Birmingham, Birmingham, AL 35294-0022, USA.
International Journal of Obesity (2005) 29, 872–874
& 2005 Nature Publishing Group All rights reserved 0307-0565/05 $30.00
CAGG-30and annealing temperature of 641C. In all, 10ml
of the PCR product were digested at 371C for 16h with 10u
of the restriction enzyme AciI, specific for the sequence
CCGC. The polymorphisms were detected after separation
by electrophoresis on a 2% agarose gel in TEA at 100V for
1.5h. Gels were stained with ethidium bromide followed
by visualization of separated DNA fragments under UV
light. The T allele is not cut with AciI and is seen as 236bp
band, whereas allele C is cut into two fragments of 148
and 88bp. An association between the genetic poly-
morphism and body mass index (BMI) was tested using
linear regression. Models were adjusted for age and gender.
All analyses were performed using SAS version 9.0 (SAS
Institute, Cary, NC, USA). A P-value o0.05 was considered
and hPRKG1-R 50-GTGAAAGGCTTTGCTT
The C2276T polymorphism in the PRKG1 gene was exam-
ined in 143 individuals from the New York City area. Among
them, 51 obese individuals were 15–63y old (mean age is
36.0y; s.d.¼10.9), with BMI ranging from 33.79 to 89.53kg/
m2. The 92 nonobese individuals were 18–54y old (mean age
is 29.4y; s.d.¼7.8), with BMI ranging from 16.02 to
29.46kg/m2. The ethnic makeup was 81% European-Amer-
ican, 14% African-American and 5% Hispanic-American. The
summary of phenotype and genotype information is pre-
sented in Table 1.
BMI levels were predicted by linear regression models
containing demographic factors (age and gender) and
genotype categories recoded as dummy variables. Race was
not included in the model due to the small number of
African-Americans and Hispanic-Americans in the study.
However, to assess if population stratification could affect
results, analysis was performed twice: for individuals of all
ethnical backgrounds (Table 2, model 1) and for European-
Americans only (Table 2, model 2). In Model 1, only age had
a significant effect (P-value¼0.0012). For model 2, the age
effect was not significant (P-value¼0.0933). In both cases,
neither gender nor genotype had significant effect. In
summary, we found no compelling evidence that the genetic
polymorphism C2276T in PRKG1 gene is associated with
Cyclic GMP is the intracellular molecule common to the
nitric oxide (NO) signaling pathway.3The cGMP-dependent
protein kinase is a major receptor for cGMP in a variety of
cells and has an evolutionary conserved structure.10Mam-
mals have two PKG genes, PRKG1 and PRKG2 (also known as
cGK I and cGK II). Two isoforms of the first type, a and b, are
produced via alternative splicing and are expressed in cardiac
and smooth muscle cells, platelets, cerebellum and other
tissues.10PRKG1 knockout mice have a decreased lifespan,
defects in relaxation of vascular, visceral, and penile smooth
muscle, disturbed platelet adhesion and activation, and
impaired guidance of sensory axons during embryogen-
esis.11,12The NO/cGMP pathway, where PRKG1 is one of the
downstream players, mediates regulatory effects on key
enzymes of fatty acid synthesis and oxidation and may be
involved in physiological control of fatty acid metabolism in
liver.13Leptin-deficient obese mice show significant changes
in NO/cGMP signaling pathway.14
The functions of PKG in neurons have been much less
clearly defined in mammals, but interesting observations
have been made in evolutionarily distant organisms.15,16
Allelic variation in the for gene encoding PRKG1 homolog in
Drosophila results in differences in food-related behaviors.4,16
The conserved role of PKG in food-related behavior has been
demonstrated in the honey bee Apis mellifera5and in the
nematode Caenorhabditis elegans.6Wu et al (2003)17specu-
lated that the product of the for gene might be a component
of the Drosophila Neuropeptide F (dNPF) signaling pathway,
A summary of the genotype and phenotype information
Parameter estimates and significance levels for linear regression
Model 1 was conducted using individuals of all ethnic backgrounds, and
model 2 using European-Americans only.
PKG polymorphism and obesity
SO Zakharkin et al
International Journal of Obesity
which regulates food-related behaviors.18 Download full-text
homolog of human NPY, is a conserved hypothalamic
neuromodulator that strongly increases food intake.7
Thus, we speculated that polymorphism in PRKG1 gene
could be associated with obesity via a number of different
mechanisms. The polymorphism found in the 30UTR of the
mature mRNA of PRKG1 at the position 2276 could be
important for the regulation of translation of the mature
mRNA or its stability.9We tested the relation between this
polymorphism and obesity but did not detect significant
This study has several limitations. We observe departure
from Hardy–Weinberg equilibrium in our sample, which is
not uncommon.19The PRKG1 gene may be a good candidate
from an evolutionary perspective, but its effect on obesity in
humans may be minimal. At the time of the study, there was
only one identified SNP in the coding region of the PRKG1
gene. Recently, more SNPs have been identified in this
locus (over 200 in dbSNP database at NCBI of which 80
were confirmed by the International HapMap project20).
Our results are limited in making a powerful conclusion
due to the fact that we have only used a single poly-
morphism. In future studies, it would be interesting to test
if any of the recently identified SNPs show association with
The dNPF, a
We acknowledge Ming D Yang and Hiwote T Belay for
techniqual support. This work was supported in part by NIH
Grants P30DK056336 and T32HL072757 and a research
grant from the Canada Research Chairs Program to MBS.
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PKG polymorphism and obesity
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International Journal of Obesity