Cell, Vol. 69, 217-220,
April 17, 1992, Copyright
0 1992 by Cell Press
Tackling a Weighty Problem Jeffrey M. Friedman* and Rudolph L. Leibelt *Howard Hughes Medical Institute fLaboratory of Human Behavior and Metabolism The Rockefeller University New York. New York 10021
Obesity, defined as an excess of body fat relative to lean body mass, is associated with important psychological and medical morbidities, the latter including hypertension, elevated blood lipids, and Type II or non-insulin-dependent diabetes mellitus (NIDDM). There are 6-10 million individuals with NIDDM in the U. S., including 18% of the population over 65 years of age (Harris et al., 1987). Approximately 45% of males and 70% of females with NIDDM are obese, and their diabetes is substantially improved or eliminated by weight reduction (Harris, 1991). As described below, both obesity and NIDDM are strongly heritable, though the predisposing genes have not been identified. The molecular genetic basis of these metabolically related disorders is an important, poorly understood problem. The Set Point Hypothesis The assimilation, storage, and utilization of nutrient energy constitute a complex homeostatic system central to survival of metazoa. Among land-dwelling mammals, storage in adipose tissue of large quantities of metabolic fuel as triglycerides is crucial for surviving periods of food deprivation. The need to maintain a fixed level of energy stores without continual alterations in the size and shape of the organism requires the achievement of a balance between energy intake and expenditure. However, the molecular mechanisms that regulate energy balance remain to be elucidated. The isolation of molecules that transduce nutntional information and control energy balance will be critical to an understanding of the regulation of body weight in health and disease. The “set point” hypothesis, a useful model for the longterm regulation of energy balance, proposes that food intake and energy expenditure are coordinately regulated by vegetative centers in the central nervous system, so as to maintain a relatively constant level of energy stores and, hence, body weight (Keesey, 1980; Harris, 1990). This hypothesis suggests that the status of energy stores is sensed by the central nervous system, and deviations in the intensity of a putative signal(s) results in adjustments in food intake and energy expenditure. Theset point model implies the existence of four major components of an energy homeostasis system: afferent signals indicating the quantity/composition of energy stores, efferent processes regulating energy storage and expenditure (e.g., autonomic nervous system), efferent mechanisms controlling ingestive behavior (e.g., neuropeptides, other neurotransmitters), and integration of the first three components in the central nervous system. The hypothalamus plays an important role in regulating body weight and may be the site at which the set point
Minireview
is regulated. Lesions of the ventromedial hypothalamus result in an obese phenotype with an apparent elevation of the set point for body fat. Conversely, lesions of the lateral hypothalamus cause a decrease in food intake, an increase in energy expenditure, and a resulting decline in body fat (Keesey and Pawley, 1986). The magnitude of these apparent changes in set point is proportional to the amount of injury sustained (Bray et al., 1989). Other hypothalamic regions also appear to play a role in the control of food intake and/or energy expenditure, asdoother brain regions, such as the area postrema and dorsal vagal complex in the brain stem. In general, central nervous system lesions that increase body weight are accompanied by an increase in the activity of the parasympathetic nervous system and a concomitant decrease in sympathetic tone. These alterations in the activity of the autonomic nervous system result in diminished energy expenditure and increased pancreatic insulin release, which, when accompanied by increased food intake, result in rapid weight gain (Bray, 1989). A number of neuropeptides, neurotransmitters, and drugs alter food intake when administered peripherally or into the hypothalamus. In general, the specificity of biologic effects is easier to attribute to agents that increase rather than decrease food intake, because it is often difficult to distinguish a physiologic effect on satiety from nonspecific effects that result in aversive behaviors. Nevertheless, cholecystokinin, insulin, corticotrophin-releasing hormone, and bombesin can cause satiety, while neuropeptide Y, peptide YY, and galinin increase food intake. Among classical neurotransmitters, norepinephrine acting via a2 receptors and GABA acting via the GABAa receptor tend to increase food intake, while serotonin tends to preferentially decrease carbohydrate intake after introduction into the hypothalamus (Woods and Gibbs, 1989). It is not clear in most cases that such effects are physiologic, nor are the precise mechanism and site of action known. The effects of cholecystokinin do appear to be physiologic, because nonpeptide antagonists of cholecystokinin transiently increase food intake (Smith et al., 1991). Administration of cholecystokinin intraperitoneally suppresses food intake via interaction with cholecystokinin-A receptors, which stimulate afferent fibers of the vagus nerve that connect with the hindbrain. However, none of these molecules appear to play a central role in regulating body weight. The identification of novel molecules that are conclusively involved in the regulation of energy balance has proven difficult, because of the lack of simple, rapid bioassays that reflect changes in the control of energy homeostasis in vivo. The cloning of mutant mammalian genes that affect the regulation of body weight represents an alternative means of identifying molecules that play a central role in this complex system. Heritable Forms of Obesity An individual‘s level of adiposity is, to a large extent, genetically determined. Examination of the concordance rates of body weight and adiposity amongst mono- and dizygous
Cell 218
twins or adoptees and their biological parents have suggested that the heritability of obesity (0.4-0.8) exceeds that of many other traits commonly thought to have a substantial genetic component, such as schizophrenia, alcoholism, and atherosclerosis (Stunkard et al., 1990). Familial similarities in rates of energy expenditure have also been reported (Bogardus et al., 1986). Genetic analysis in geographically delimited populations has suggested that a relatively small number of genes may account for the 300/o-50% of variance in body composition (Mall et al., 1991). However, none of the genes responsible for obesity in the general population have been genetically mapped. Several uncommon genetic syndromes that include obesity as a salient feature have been characterized. Prader-Willi syndrome, the most common form of dysmorphic genetic obesity, is associated with mental retardation, hypotonia, small hands and feet, short stature, and hypogonadotrophic hypogonadism. This syndrome is often associated with deletions of paternal chromosome 15ql l13. Prader-Willi syndrome can also be a result of maternal isodisomy for the same subgenomic region, suggesting that the gene(s) responsible for this syndrome are imprinted (Nicholls et al., 1989). Individuals with BardetBiedl syndrome, Alstrom syndrome, or Cohen (Pepper) syndrome also manifest an obese phenotype associated with a variety of dysmorphic features (Norioa et al., 1984, and references therein). Each of these uncommon disorders appears to be inherited as an autosomal recessive, although their map positions remain to be determined. Certain relatively inbred populations, such as the Pima Indians and some Micronesian islanders, manifest a high
Table
1. Obesity
Syndromes
Disorder
Single-Gene Humans
Mice
Prader-Willi syndrome Alstrom syndrome Bardet-Biedl syndrome Cohen syndrome
Mice
Chromosome
Homologous in Mouse
Dominant Autosomal Autosomal Autosomal
(imprinted) recessive recessive recessive
15q11-13 ? ? ?
7 ? ? ?
(?) (?)
Chromosome
Homologous in Human
Name
Allele
Inheritance
Yellow
AY A”” A’” ob
Autosomal
dominant
2
2oq13
Autosomal
recessive
6
7q31
db db” db” dM”
Autosomal
recessive
4
lp31
Diabetes
Inbred
Inheritance
Region
Mutations
Obese
Rats
incidence of obesity and Type II diabetes, although the precise mode of inheritance in these populations has not been ascertained. Rodent models of obesity include seven apparently single-gene mutations and several inbred lines that are obese as a result of polygenic inheritance (Table 1). The use of such animal models of human disease-particularly mouse-offers important experimental advantages that allow genetic dissection of heterogeneous and polygenic traits. Single-gene mutations can be segregated in genetic crosses and analyzed in a fashion similar to that implemented for positional cloning of human disease genes (Friedman et al., 1991a). Polygenes can now be mapped by segregating traits in genetic crosses and haplotyping the progeny at the phenotypic extremes with molecular markers that span the genome (Lander and Botstein, 1989). This approach has been employed to map rodent genes involved in the inheritance of Type I (insulindependent) and hypertension and will also make it possible to map genetic loci that modify the phenotype produced by mutations in single genes (Jacob et al., 1991; Todd et al., 1991). The most intensively studied mouse obesity mutations are the ob (obese) and db (diabetes) genes. When present on the same genetic strain background, ob and db result in indistinguishable metabolic and behaviorial phenotypes, suggesting that these genes may function in the same physiologic pathway (Coleman, 1978). Mice homozygous for either mutation are hyperphagic and hypometabolic, leading to an obese phenotype that is notable at one month of age. The weight of these animals tends to stabilize at
Fat
fat
Autosomal
recessive
8
16q?
Tubby
tub
Autosomal
recessive
7
11p?
Adipose
Ad
Autosomal
dominant
7
lip?
Fatty (corpulent)
fa fa”
Autosomal
recessive
5
1p31
? ?
? ?
Strains NZO KK
Polygenic Polygenic
Region
Minireview 219
60-70 g (compared with 30-35 g in control mice). ob and animals manifest a myriad of other hormonal and metabolic changes that have made it difficult to identify the primary defect attributable to the mutation (Bray et al., 1989). These include abnormalities in several hypothalamic functions, such as thermoregulation and gonadotrophin secretion (resulting in infertility). While many of these features are manifest in humans with Prader-Willi syndrome, comparative genetic mapping between man and mouse suggests that the mouse homolog of Prader-Willi syndrome is on proximal chromosome 7 and is unlikely to be allelic with any of the rodent obesity genes (Chaillet et al., 1991). The AY (lethal yellow, yellow agouti) gene is a dominant mutation that is very tightly linked to (and possibly allelic with) the agouti coat color locus (A). AY is pleiotypic, with phenotypic changes that include a yellow coat color, mild obesity, and heightened susceptibility to mammary, hepatic, and bladder tumors (Wolff et al., 1986). AY is the only obesity mutation in which the linear (skeletal) growth of the animal is increased. In AY and several other dominant yellow mutations (A”“, etc.) the intensity of the yellow pigmentation correlates positively with the extent of the obesity. Transplantation of hair follicles and fat cells from mutant into wild-type mice suggests that the mutant phenotype is a result of increased activity of a non-cellautonomous molecule. This mutation has been mapped to a short interval that is syntenic between distal chromosome 2 in mouse and 20q in man (Siracusa et al., 1987). The recent cloning of DNA from an inversion breakpoint between Id (limb deformity) and A (agouti) makes it likely that AY will be the first obesity gene to be cloned (Bultman et al., 1991). Each of the rodent obesity models is accompanied by alterations in carbohydrate metabolism resembling those in Type II diabetes in man. In some cases, the severity of the diabetes depends in part on the background mouse strain (Leiter, 1989). For both ob and db, congenic C57BL/ KS mice develop a severe diabetes with ultimate 6 cell necrosis and islet atrophy, resulting in a relative insulinopenia. Conversely, congenic C57BL16J ob and db mice develop a transient insulin-resistant diabetes that is eventually compensated by 6 cell hypertrophy resembling human Type II diabetes. Moreover, in several different strains, there is a higher incidence in males carrying the db mutation of the development of insulinopenic diabetes. Phenotypic characterization of different congenic db mouse strains has suggested that genetic differences in the rates of secretion and metabolism of androgens and estrogens account for some of the variability in the incidence of pancreatic decompensation in mutant animals (Leiter, 1989). Estrogens protect against and androgens predispose to the adverse consequences of these obesity genes, with regard to decompensation of pancreatic f3 cells. Accordingly, genetically influenced factors such as hepatic steroid sulfuration may influence the severity of diabetes occurring in genetically obese animals. The phenotype of ob and db mice resembles human obesity in ways other than the development of diabetesthe mutant mice eat more and expend less energy than do
db
lean controls (as do obese humans). This phenotype is also quite similar to that seen in animals with lesions of the ventromedial hypothalamus, which suggests that both mutations may interfere with the ability to properly integrate or respond to nutritional information within the central nervous system. Support for this hypothesis comes from the results of parabiosis experiments (Coleman, 1973) that suggest ob mice are deficient in a circulating satiety factor and that db mice are resistant to the effects of the ob factor (possibly due to an ob receptor defect). These experiments have led to the conclusion that obesity in these mutant mice may result from different defects in an afferent loop and/or integrative center of the postulated feedback mechanism that controls body composition. Adrenal steroids may also play a permissive role in the development of the obese phenotype in these animals, since adrenalectomy ameliorates many aspects of the phenotype (Bray, 1989). Glucocorticoids might be required in this context to suppress corticotrophin-releasing hormone levels in the hypothalamus, since hypothalamic administration of corticotrophin-releasing hormone into fa rats appears to reduce body weight (Rohner-Jeanrenaud et al., 1989). Corticotrophin-releasing hormone is known to affect sympathetic output from the hypothalamus. Using molecular and classical genetic markers, the ob and db genes have been mapped to proximal chromosome 6 and midchromosome 4, respectively (Baharyet al., 1990; Friedman et al., 1991 b). In both cases, the mutations map to regions of the mouse genome that are syntenic with human, suggesting that, if there are human homologs of ob and db, they are likely to map, respectively, to human chromosomes 7q and lp. Defects in the db gene may result in obesity in other mammalian species: in genetic crosses between Zucker falfa rats and Brown Norway +/+ rats, the fa mutation (rat chromosome 5) is flanked by the same loci that flank db in mouse (Truett et al., 1991). A second allele of fat, corpulent (fa”p), is also available for study. Thus, the mouse db and rat fa mutations are likely to be defects in homologous genes. Using markers from 7q and 1p, genetic analysis of human pedigrees segregating an obese phenotype may suggest whether haplotypes at the putative human homologs of ob and db are etiologic in the human inheritance of obesity. While the tubby (tub) mutation has been mapped to mouse chromosome 7 and fat has been mapped to mouse chromosome 8, the genetic position of the putative human homologs of these mutations is less certain. The relevance of these rodent genes in human obesity should be discernible when the genes are cloned. Conventional mouse crosses that segregate mutant phenotypes can be used to map candidate genes and to narrow the search for the mutant genes to genetic intervals of well under 1 CM, even in instances when the incidence of the mutation is low (Friedman et al., 1991a). In the case of obesity genes, efforts to identify the mutant genes from genetically defined intervals are likely to be confounded by uncertainty regarding the site of synthesis of the mutant genes. In principle, allelic variation between the mutant and wild-type loci can be used to identify the mutant gene, particularly when the mutant strain is coisogenic with a
Cell 220
wild-type strain. This task will be simplified by the existence of several mutant alleles, as in the case of db. A major advantage of mouse for identifying mutant genes is that mutant phenotypes can be complemented either in transgenic mice, as was done for the shiverer mutation (Readhead et al., 1987), or by using ES cell lines. The latter technique will be particularly useful if it becomes possible to introduce yeast artificial chromosomes into totipotent ES cells to generate chimeric mice (Strauss and Jaenisch, 1991). Yeast artificial chromosomes that retain enzymatic activity or complement a mutation in the collagen gene (Strauss and Jaenisch, 1992) have been successfully introduced into cultured cells (D’Urso et al., 1990). In the long term, genes may also be mapped that modify the obese phenotypes or influence the development of Type II diabetes. Phenotypic analysis of progeny of interspecific genetic crosses segregating ob and db has suggested that unlinked polygenes inherited from the Spanish strain, Mus spretus, and the DBA/2J strain can influence insulin secretion rates, insulin sensitivity, and degree of obesity of mutant animals (Friedman et al., 1991 b). Future Prospects Molecular techniques that allow the cloning of single-gene mutations and make possible the dissection of polygenic traits are currently applicable to genetic studies of obese mammals. The cloning of many mammalian disease genes attests to the former capability, while the mapping of genetic loci conferring Type I diabetes susceptiblity in mice and hypertension in rats documents the feasibility of the latter approach (Todd et al., 1991; Jacob et al., 1991). The cloning of obesity genes could in principle provide considerable insight into the molecular mechanisms that control body composition by effects on food intake and energy expenditure. Eventually, it should be possible to clone all the singlegene rodent obesity mutations, as well as the Prader-Willi gene(s). The cloning of the genes for other human obesity syndromes is somewhat more problematic, owing to their infrequent occurrence. The use of appropriate genetic crosses in mice should allow the localization of genes that modify the severity of the obese phenotype and the Type II diabetes that accompanies obesity. Similarly, polygenes responsible for obesity in polygenic obese strains, such as NZO, can be mapped. Such loci can be identified by phenotyping and genotyping the progeny of F2 offspring in crosses between inbred mouse lines that differ widely in the amount of body fat. Localization of polygenes that determine body composition will also clarify whether allelic variation of the single genes that influence body composition (i.e., ob, db, fat, and tub) plays a role in regulating body weight in polygenic obese mice. The successful application of modern molecular genetic techniques to this area of nutrition research could have a profound impact on our understanding of the mechanisms that control body weight in health and disease. References Bahary, N., Leibel, R. L., Joseph, L., and Friedman, Natl. Acad. Sci. USA 87, 8642-6646.
J. M. (1990).
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Bogardus, C., Ravussin, E., Abbott, W., Zawakzku, J. K., Young, A., Knowler, W. C., Jacobowitz, R., and Mall, P. 0. (1986). N. Engl. J. Med. 375, 96-100. Bray,
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Bultman, S. J., Russell, L. B., Gutierrez-Espeleta, G. A., and Woychik, Ft. P. (1991). Proc. Natl. Acad. Sci. USA 88, 6062-6066. Chaillet, J. R., Knoll, J. H. M., Horsthemke, Genomics 11, 773-776. Coleman,
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B., and Lalande,
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P. H. (1967).
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Jacob, H. J., Lindpainter, K., Lincoln, S. E., Kusumi, K., Bunker, R. K., Mao, Y.-P., Ganten, D., Dzau, V. J., and Lander, E. S. (1991). Cell 67, 213-224. Keesey, R. E. (1980). In Obesity, W. B. Saunders Co.), pp. 144-166. Keesey, 133. Lander, Leiter,
R. E., and Pawley,
Mall, P. P., Burns, 49, 1243-1255.
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R., Raitta,
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eds.
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Readhead, C., Popko, B., Takahashi, N., Shine, H. D., Saavedra, P. A., Sidman, Ft. L., and Hood, L. (1987). Cell 48, 703-712. Rohner-Jeanrenaud, renaud, B. (1989).
F., Walker, Endocrinology
C. L., Greco-Perotto, 124, 733-739.
R., and Jean-
Siracusa, L. D., Russell, L. B., Either, E. M., Corrow, D. J., Copeland, N. G., and Jenkins, N. A. (1987). Genetics 777, 93-100. Smith, G. P., Gibbs, J., Schneider, L., Greenberg, D., Murphy, R., Corp, E.. and Corwin, Ft. (1991). In Progress in Obesity Research, Y. Oomura, S. Tarui, S. Inoue, and T. Shimayu, eds. (London: John Libbey & Co. Ltd.), pp. 25-28. Strauss, W. M., and Jaenisch, R. (1991). In Mouse Molecular Genetics, E. Wagner, U. Ruther, R. Palmiter, and D. Hanahan, eds. (Heidelberg: European Molecular Biology Laboratory), p. 68. Strauss,
W. M., and Jaenisch,
R. (1992).
Stunkard, A. J., Harris, J. R., Pedersen, (1990). N. Engl. J. Med. 322, 1463-1487.
EMBO
J. 77, 417-422.
N. L., and McClearn,
G. E.
Todd, J. A., Aitman, T. J., Love, R. J., McAleer, M. A., Prins, J.-B., Rodrigues, N., Lathrop, N.. Pressey, A., DeLarato, N. H., Peterson, L. B., and Wicker, L. S. (1991). Nature 357, 542-547. Truett, G. E., Bahary, N., Friedman, J. M., and Leibel, Proc. Natl. Acad. Sci. USA 88, 7806-7809.
R. L. (1991).
Wolff, G. L., Roberts, 151-158.
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D. B. (1986).
J. (1989). Ann. NY Acad. Sci. 575,236-243.
77,
April 17, 1992, Copyright
0 1992 by Cell Press
Tackling a Weighty Problem Jeffrey M. Friedman* and Rudolph L. Leibelt *Howard Hughes Medical Institute fLaboratory of Human Behavior and Metabolism The Rockefeller University New York. New York 10021
Obesity, defined as an excess of body fat relative to lean body mass, is associated with important psychological and medical morbidities, the latter including hypertension, elevated blood lipids, and Type II or non-insulin-dependent diabetes mellitus (NIDDM). There are 6-10 million individuals with NIDDM in the U. S., including 18% of the population over 65 years of age (Harris et al., 1987). Approximately 45% of males and 70% of females with NIDDM are obese, and their diabetes is substantially improved or eliminated by weight reduction (Harris, 1991). As described below, both obesity and NIDDM are strongly heritable, though the predisposing genes have not been identified. The molecular genetic basis of these metabolically related disorders is an important, poorly understood problem. The Set Point Hypothesis The assimilation, storage, and utilization of nutrient energy constitute a complex homeostatic system central to survival of metazoa. Among land-dwelling mammals, storage in adipose tissue of large quantities of metabolic fuel as triglycerides is crucial for surviving periods of food deprivation. The need to maintain a fixed level of energy stores without continual alterations in the size and shape of the organism requires the achievement of a balance between energy intake and expenditure. However, the molecular mechanisms that regulate energy balance remain to be elucidated. The isolation of molecules that transduce nutntional information and control energy balance will be critical to an understanding of the regulation of body weight in health and disease. The “set point” hypothesis, a useful model for the longterm regulation of energy balance, proposes that food intake and energy expenditure are coordinately regulated by vegetative centers in the central nervous system, so as to maintain a relatively constant level of energy stores and, hence, body weight (Keesey, 1980; Harris, 1990). This hypothesis suggests that the status of energy stores is sensed by the central nervous system, and deviations in the intensity of a putative signal(s) results in adjustments in food intake and energy expenditure. Theset point model implies the existence of four major components of an energy homeostasis system: afferent signals indicating the quantity/composition of energy stores, efferent processes regulating energy storage and expenditure (e.g., autonomic nervous system), efferent mechanisms controlling ingestive behavior (e.g., neuropeptides, other neurotransmitters), and integration of the first three components in the central nervous system. The hypothalamus plays an important role in regulating body weight and may be the site at which the set point
Minireview
is regulated. Lesions of the ventromedial hypothalamus result in an obese phenotype with an apparent elevation of the set point for body fat. Conversely, lesions of the lateral hypothalamus cause a decrease in food intake, an increase in energy expenditure, and a resulting decline in body fat (Keesey and Pawley, 1986). The magnitude of these apparent changes in set point is proportional to the amount of injury sustained (Bray et al., 1989). Other hypothalamic regions also appear to play a role in the control of food intake and/or energy expenditure, asdoother brain regions, such as the area postrema and dorsal vagal complex in the brain stem. In general, central nervous system lesions that increase body weight are accompanied by an increase in the activity of the parasympathetic nervous system and a concomitant decrease in sympathetic tone. These alterations in the activity of the autonomic nervous system result in diminished energy expenditure and increased pancreatic insulin release, which, when accompanied by increased food intake, result in rapid weight gain (Bray, 1989). A number of neuropeptides, neurotransmitters, and drugs alter food intake when administered peripherally or into the hypothalamus. In general, the specificity of biologic effects is easier to attribute to agents that increase rather than decrease food intake, because it is often difficult to distinguish a physiologic effect on satiety from nonspecific effects that result in aversive behaviors. Nevertheless, cholecystokinin, insulin, corticotrophin-releasing hormone, and bombesin can cause satiety, while neuropeptide Y, peptide YY, and galinin increase food intake. Among classical neurotransmitters, norepinephrine acting via a2 receptors and GABA acting via the GABAa receptor tend to increase food intake, while serotonin tends to preferentially decrease carbohydrate intake after introduction into the hypothalamus (Woods and Gibbs, 1989). It is not clear in most cases that such effects are physiologic, nor are the precise mechanism and site of action known. The effects of cholecystokinin do appear to be physiologic, because nonpeptide antagonists of cholecystokinin transiently increase food intake (Smith et al., 1991). Administration of cholecystokinin intraperitoneally suppresses food intake via interaction with cholecystokinin-A receptors, which stimulate afferent fibers of the vagus nerve that connect with the hindbrain. However, none of these molecules appear to play a central role in regulating body weight. The identification of novel molecules that are conclusively involved in the regulation of energy balance has proven difficult, because of the lack of simple, rapid bioassays that reflect changes in the control of energy homeostasis in vivo. The cloning of mutant mammalian genes that affect the regulation of body weight represents an alternative means of identifying molecules that play a central role in this complex system. Heritable Forms of Obesity An individual‘s level of adiposity is, to a large extent, genetically determined. Examination of the concordance rates of body weight and adiposity amongst mono- and dizygous
Cell 218
twins or adoptees and their biological parents have suggested that the heritability of obesity (0.4-0.8) exceeds that of many other traits commonly thought to have a substantial genetic component, such as schizophrenia, alcoholism, and atherosclerosis (Stunkard et al., 1990). Familial similarities in rates of energy expenditure have also been reported (Bogardus et al., 1986). Genetic analysis in geographically delimited populations has suggested that a relatively small number of genes may account for the 300/o-50% of variance in body composition (Mall et al., 1991). However, none of the genes responsible for obesity in the general population have been genetically mapped. Several uncommon genetic syndromes that include obesity as a salient feature have been characterized. Prader-Willi syndrome, the most common form of dysmorphic genetic obesity, is associated with mental retardation, hypotonia, small hands and feet, short stature, and hypogonadotrophic hypogonadism. This syndrome is often associated with deletions of paternal chromosome 15ql l13. Prader-Willi syndrome can also be a result of maternal isodisomy for the same subgenomic region, suggesting that the gene(s) responsible for this syndrome are imprinted (Nicholls et al., 1989). Individuals with BardetBiedl syndrome, Alstrom syndrome, or Cohen (Pepper) syndrome also manifest an obese phenotype associated with a variety of dysmorphic features (Norioa et al., 1984, and references therein). Each of these uncommon disorders appears to be inherited as an autosomal recessive, although their map positions remain to be determined. Certain relatively inbred populations, such as the Pima Indians and some Micronesian islanders, manifest a high
Table
1. Obesity
Syndromes
Disorder
Single-Gene Humans
Mice
Prader-Willi syndrome Alstrom syndrome Bardet-Biedl syndrome Cohen syndrome
Mice
Chromosome
Homologous in Mouse
Dominant Autosomal Autosomal Autosomal
(imprinted) recessive recessive recessive
15q11-13 ? ? ?
7 ? ? ?
(?) (?)
Chromosome
Homologous in Human
Name
Allele
Inheritance
Yellow
AY A”” A’” ob
Autosomal
dominant
2
2oq13
Autosomal
recessive
6
7q31
db db” db” dM”
Autosomal
recessive
4
lp31
Diabetes
Inbred
Inheritance
Region
Mutations
Obese
Rats
incidence of obesity and Type II diabetes, although the precise mode of inheritance in these populations has not been ascertained. Rodent models of obesity include seven apparently single-gene mutations and several inbred lines that are obese as a result of polygenic inheritance (Table 1). The use of such animal models of human disease-particularly mouse-offers important experimental advantages that allow genetic dissection of heterogeneous and polygenic traits. Single-gene mutations can be segregated in genetic crosses and analyzed in a fashion similar to that implemented for positional cloning of human disease genes (Friedman et al., 1991a). Polygenes can now be mapped by segregating traits in genetic crosses and haplotyping the progeny at the phenotypic extremes with molecular markers that span the genome (Lander and Botstein, 1989). This approach has been employed to map rodent genes involved in the inheritance of Type I (insulindependent) and hypertension and will also make it possible to map genetic loci that modify the phenotype produced by mutations in single genes (Jacob et al., 1991; Todd et al., 1991). The most intensively studied mouse obesity mutations are the ob (obese) and db (diabetes) genes. When present on the same genetic strain background, ob and db result in indistinguishable metabolic and behaviorial phenotypes, suggesting that these genes may function in the same physiologic pathway (Coleman, 1978). Mice homozygous for either mutation are hyperphagic and hypometabolic, leading to an obese phenotype that is notable at one month of age. The weight of these animals tends to stabilize at
Fat
fat
Autosomal
recessive
8
16q?
Tubby
tub
Autosomal
recessive
7
11p?
Adipose
Ad
Autosomal
dominant
7
lip?
Fatty (corpulent)
fa fa”
Autosomal
recessive
5
1p31
? ?
? ?
Strains NZO KK
Polygenic Polygenic
Region
Minireview 219
60-70 g (compared with 30-35 g in control mice). ob and animals manifest a myriad of other hormonal and metabolic changes that have made it difficult to identify the primary defect attributable to the mutation (Bray et al., 1989). These include abnormalities in several hypothalamic functions, such as thermoregulation and gonadotrophin secretion (resulting in infertility). While many of these features are manifest in humans with Prader-Willi syndrome, comparative genetic mapping between man and mouse suggests that the mouse homolog of Prader-Willi syndrome is on proximal chromosome 7 and is unlikely to be allelic with any of the rodent obesity genes (Chaillet et al., 1991). The AY (lethal yellow, yellow agouti) gene is a dominant mutation that is very tightly linked to (and possibly allelic with) the agouti coat color locus (A). AY is pleiotypic, with phenotypic changes that include a yellow coat color, mild obesity, and heightened susceptibility to mammary, hepatic, and bladder tumors (Wolff et al., 1986). AY is the only obesity mutation in which the linear (skeletal) growth of the animal is increased. In AY and several other dominant yellow mutations (A”“, etc.) the intensity of the yellow pigmentation correlates positively with the extent of the obesity. Transplantation of hair follicles and fat cells from mutant into wild-type mice suggests that the mutant phenotype is a result of increased activity of a non-cellautonomous molecule. This mutation has been mapped to a short interval that is syntenic between distal chromosome 2 in mouse and 20q in man (Siracusa et al., 1987). The recent cloning of DNA from an inversion breakpoint between Id (limb deformity) and A (agouti) makes it likely that AY will be the first obesity gene to be cloned (Bultman et al., 1991). Each of the rodent obesity models is accompanied by alterations in carbohydrate metabolism resembling those in Type II diabetes in man. In some cases, the severity of the diabetes depends in part on the background mouse strain (Leiter, 1989). For both ob and db, congenic C57BL/ KS mice develop a severe diabetes with ultimate 6 cell necrosis and islet atrophy, resulting in a relative insulinopenia. Conversely, congenic C57BL16J ob and db mice develop a transient insulin-resistant diabetes that is eventually compensated by 6 cell hypertrophy resembling human Type II diabetes. Moreover, in several different strains, there is a higher incidence in males carrying the db mutation of the development of insulinopenic diabetes. Phenotypic characterization of different congenic db mouse strains has suggested that genetic differences in the rates of secretion and metabolism of androgens and estrogens account for some of the variability in the incidence of pancreatic decompensation in mutant animals (Leiter, 1989). Estrogens protect against and androgens predispose to the adverse consequences of these obesity genes, with regard to decompensation of pancreatic f3 cells. Accordingly, genetically influenced factors such as hepatic steroid sulfuration may influence the severity of diabetes occurring in genetically obese animals. The phenotype of ob and db mice resembles human obesity in ways other than the development of diabetesthe mutant mice eat more and expend less energy than do
db
lean controls (as do obese humans). This phenotype is also quite similar to that seen in animals with lesions of the ventromedial hypothalamus, which suggests that both mutations may interfere with the ability to properly integrate or respond to nutritional information within the central nervous system. Support for this hypothesis comes from the results of parabiosis experiments (Coleman, 1973) that suggest ob mice are deficient in a circulating satiety factor and that db mice are resistant to the effects of the ob factor (possibly due to an ob receptor defect). These experiments have led to the conclusion that obesity in these mutant mice may result from different defects in an afferent loop and/or integrative center of the postulated feedback mechanism that controls body composition. Adrenal steroids may also play a permissive role in the development of the obese phenotype in these animals, since adrenalectomy ameliorates many aspects of the phenotype (Bray, 1989). Glucocorticoids might be required in this context to suppress corticotrophin-releasing hormone levels in the hypothalamus, since hypothalamic administration of corticotrophin-releasing hormone into fa rats appears to reduce body weight (Rohner-Jeanrenaud et al., 1989). Corticotrophin-releasing hormone is known to affect sympathetic output from the hypothalamus. Using molecular and classical genetic markers, the ob and db genes have been mapped to proximal chromosome 6 and midchromosome 4, respectively (Baharyet al., 1990; Friedman et al., 1991 b). In both cases, the mutations map to regions of the mouse genome that are syntenic with human, suggesting that, if there are human homologs of ob and db, they are likely to map, respectively, to human chromosomes 7q and lp. Defects in the db gene may result in obesity in other mammalian species: in genetic crosses between Zucker falfa rats and Brown Norway +/+ rats, the fa mutation (rat chromosome 5) is flanked by the same loci that flank db in mouse (Truett et al., 1991). A second allele of fat, corpulent (fa”p), is also available for study. Thus, the mouse db and rat fa mutations are likely to be defects in homologous genes. Using markers from 7q and 1p, genetic analysis of human pedigrees segregating an obese phenotype may suggest whether haplotypes at the putative human homologs of ob and db are etiologic in the human inheritance of obesity. While the tubby (tub) mutation has been mapped to mouse chromosome 7 and fat has been mapped to mouse chromosome 8, the genetic position of the putative human homologs of these mutations is less certain. The relevance of these rodent genes in human obesity should be discernible when the genes are cloned. Conventional mouse crosses that segregate mutant phenotypes can be used to map candidate genes and to narrow the search for the mutant genes to genetic intervals of well under 1 CM, even in instances when the incidence of the mutation is low (Friedman et al., 1991a). In the case of obesity genes, efforts to identify the mutant genes from genetically defined intervals are likely to be confounded by uncertainty regarding the site of synthesis of the mutant genes. In principle, allelic variation between the mutant and wild-type loci can be used to identify the mutant gene, particularly when the mutant strain is coisogenic with a
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wild-type strain. This task will be simplified by the existence of several mutant alleles, as in the case of db. A major advantage of mouse for identifying mutant genes is that mutant phenotypes can be complemented either in transgenic mice, as was done for the shiverer mutation (Readhead et al., 1987), or by using ES cell lines. The latter technique will be particularly useful if it becomes possible to introduce yeast artificial chromosomes into totipotent ES cells to generate chimeric mice (Strauss and Jaenisch, 1991). Yeast artificial chromosomes that retain enzymatic activity or complement a mutation in the collagen gene (Strauss and Jaenisch, 1992) have been successfully introduced into cultured cells (D’Urso et al., 1990). In the long term, genes may also be mapped that modify the obese phenotypes or influence the development of Type II diabetes. Phenotypic analysis of progeny of interspecific genetic crosses segregating ob and db has suggested that unlinked polygenes inherited from the Spanish strain, Mus spretus, and the DBA/2J strain can influence insulin secretion rates, insulin sensitivity, and degree of obesity of mutant animals (Friedman et al., 1991 b). Future Prospects Molecular techniques that allow the cloning of single-gene mutations and make possible the dissection of polygenic traits are currently applicable to genetic studies of obese mammals. The cloning of many mammalian disease genes attests to the former capability, while the mapping of genetic loci conferring Type I diabetes susceptiblity in mice and hypertension in rats documents the feasibility of the latter approach (Todd et al., 1991; Jacob et al., 1991). The cloning of obesity genes could in principle provide considerable insight into the molecular mechanisms that control body composition by effects on food intake and energy expenditure. Eventually, it should be possible to clone all the singlegene rodent obesity mutations, as well as the Prader-Willi gene(s). The cloning of the genes for other human obesity syndromes is somewhat more problematic, owing to their infrequent occurrence. The use of appropriate genetic crosses in mice should allow the localization of genes that modify the severity of the obese phenotype and the Type II diabetes that accompanies obesity. Similarly, polygenes responsible for obesity in polygenic obese strains, such as NZO, can be mapped. Such loci can be identified by phenotyping and genotyping the progeny of F2 offspring in crosses between inbred mouse lines that differ widely in the amount of body fat. Localization of polygenes that determine body composition will also clarify whether allelic variation of the single genes that influence body composition (i.e., ob, db, fat, and tub) plays a role in regulating body weight in polygenic obese mice. The successful application of modern molecular genetic techniques to this area of nutrition research could have a profound impact on our understanding of the mechanisms that control body weight in health and disease. References Bahary, N., Leibel, R. L., Joseph, L., and Friedman, Natl. Acad. Sci. USA 87, 8642-6646.
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