Fat Mass and Obesity Associated Gene (FTO)
Allen W. Root, MD
Several genome wide association (GWA) studies have linked FTO (Fat mass and obesity-associated gene - OMIM 610966, chromosome 16q12.2) to weight and obesity risk in children and adults of diverse ethnic origin.1 Several single nucleotide polymorphisms (SNPs) in intron 1 of FTO predispose to obesity while others seem to protect the carrier from this trait. FTO encodes a nuclear non-heme iron- and 2-oxoglutarate-dependent dioxygenase that catalyzes the conversion of 2-oxoglutarate to succinate and the demethylation of 3-methylthymine and 3-methyluracil in DNA and RNA, respectively.2,3 Oxidative demethylation of alkylated nucleic acids is essential for maintenance of an intact genome. FTO is expressed ubiquitously in all fetal and adult tissues - particularly in the hypothalamic arcuate nucleus, pituitary, heart, and liver. The arcuate nucleus is the site of synthesis of proopiomelanocortin (POMC) and its anorexigenic product α-melanocyte stimulating hormone and of orexigenic agouti-related peptide (AGRP) and neuropeptide Y (NPY) - essential components of the appetite regulating system. Within the arcuate nucleus, Fto is expressed in Pomc synthesizing as well as other neurons. Arcuate nucleus expression of Fto is attenuated by fasting and amplified by feeding - particularly of a high fat diet. 2,4
Boissel et al have identified a consanguineous Palestinian family in which many third generation members displayed impairment of postnatal growth, developmental delay, and death within the first three years of life.5 They presented malformations involving the CNS (microcephaly, lissencephaly, brain atrophy, neurosensory deafness), heart (ventricular septal and atrioventricular defects, hypertrophic cardiomyopathy), face (anteverted nostrils, thin vermillion borders, retrognathia, cleft palate) and other regions (short neck, brachydactyly, hypoplasia of toenails, ambiguous genitalia). The investigators linked this malformative syndrome to an autosomal recessive, homozygous, loss-of-function mutation in FTO. A homozygous guanine to adenine transition at nucleotide position 947 (c.947G⇒A) resulted in substitution of glutamine for arginine at codon 316 (Arg316Gln = p.R316Q), an absolutely conserved position in related orthologous genes in many species. The p.R316Q substitution significantly impaired the function of the enzyme. In addition, in vitro the rate of proliferation and the life span of cultured skin fibroblasts from one of these patients were significantly reduced indicating that these cells aged quickly.
FTO in Experimental Animals
While loss of FTO in humans results in a devastating and lethal complex of anomalies, "knock out" of the murine homolog Fto leads to a less severe outcome. Fischer et al developed Fto-/- mice by replacing exons 2 and 3 with a neomycin resistant STOP cassette leading to diffuse, germline loss of expression of Fto.6Â "Knock out" of Fto did not increase fetal wastage; Fto-/-fetuses had normal embryogenesis and organogenesis. Although of normal size at birth, weight gain and linear growth of male and female Fto-/- neonates faltered within the first week after birth. The growth of heterozygous Fto+/- mice was similar to that of wild-type (WT) animals (Figure).
Decreased weight of Fto-/- mice was due primarily to lower white fat mass compared to WT animals. Interestingly, brown fat mass was similar in WT and Fto-/- mice. White fat accumulates and stores fat and energy, while brown fat metabolizes and expends energy by uncoupling the processes of heat production and ATP generation by generation of uncoupling proteins encoded by Ucp1, Ucp2, and Ucp3. Further studies revealed that the food intake of the WT and Fto-/- mice was comparable indicating that relative to body weight the Fto-/- animals were actually hyperphagic. The expression of Pomc and Npy in the arcuate nucleus was similar in Fto-/-and WT mice. Energy expenditure in Fto-/- mice as assessed by oxygen consumption, carbon dioxide production, and heat generation was significantly greater than in WT mice despite their relative physical inactivity as assessed by determination of spontaneous locomotion. However, increased energy expenditure was not due to greater expression of mitochondrial Ucp1 in brown adipose tissue or to increased thyroid hormone generation but rather to enhanced sympathetic activity as suggested by higher plasma concentrations of norepinephrine and epinephrine in Fto-/- than WT mice. The authors concluded that, in mice, loss of Fto increases energy expenditure by enhancing sympathetic activity resulting in futile (ie, non-energy producing) metabolism of triglycerides and fatty acids perhaps in skeletal or cardiac muscle or liver.7 It is also possible that the expression of Ucp2 and/or Ucp3 was increased in brown adipose tissue thus dissipating energy through non-shivering thermogenesis.
Church et al developed a mouse model with a missense mutation (A⇒T) in exon 6 of Fto that resulted in replacement of isoleucine by phenylalanine in codon 367 (Ile367Phe = I367F) in the carboxyl terminal region of Fto.8 This site is not within the catalytic core of Fto but rather in a highly conserved sequence of ~20 amino acids that is required for dimerization of Fto protein and for its optimal catalytic activity. Although FtoI367F localized to the cell nucleus, its expression was reduced and its catalytic activity attenuated but not completely absent. Normal at birth, both homozygous FtoI367F and heterozygous FtoI367F/I367 male (but not female) mice gained fat mass less rapidly than WT mice after 12 weeks of age; nevertheless, linear growth of mutant mice was comparable to that of WT animals. (The heterozygous FtoI367F/I367 mutation may exert a dominant-negative effect on the WT protein.) Relative to WT animals, metabolic rate was higher in both homozygous and heterozygous FtoI367F male mice as estimated by oxygen consumption and carbon dioxide production despite similar levels of physical exertion and brown adipose tissue thermogenic activity. Urinary excretion of catecholamines was greater in mutant than WT animals. In skeletal muscle, expression of the genes encoding the β3-adrenergic receptor, uncoupling protein-2, and catechol-O-methyl transferase was increased. Microarray analyses in white adipose tissue, skeletal muscle , and liver revealed that in FtoI367F mice expression of genes associated with inflammation were decreased and those related to both fatty acid catabolism and synthesis were increased. Hypothalamic expression of Pomc, Agrp, and Npy was not altered in FtoI367F mice. Thus, in a mouse model with less complete loss of Fto activity than in the Fto "knock-out" model, similar manifestations of Fto deficiency were noted but to a lesser extent. Interestingly, the effect of attenuation of Fto activity was not observed in female mice; a somewhat similar observation has been made in humans with a common variant of FTO.9
The impairment in weight gain and linear growth due to inactivating mutations of Fto in mice as demonstrated by Fischer et al6 and Church et al8 is primarily due to increased energy expenditure possibly due to augmented adrenergic activity. The more extensive is the loss of Fto function in mice, the more dramatic is the effect. The mechanisms by which FTO regulates energy intake and utilization are unknown. Inasmuch as FTO is a nucleotide demethylase, it is likely that its effects are mediated by differential expression of target genes that are beginning to be identified. Utilizing the male rat as a model, Tung and colleagues4 stereotactically injected Fto cDNA into the arcuate or paraventricular nuclei in order to increase Fto expression or shRNA in order to decrease synthesis of endogenous Fto. Increased expression of arcuate nucleus Fto lowered spontaneous food intake while impaired generation of Fto enhanced caloric ingestion. Overexpression of Fto in the paraventricular nucleus also impaired food intake in the rat model. They further demonstrated (as did Church et al8) that alteration in Fto expression did not affect arcuate nucleus expression of Agrp, Npy, and Pomc but enhanced Fto expression increased that of Stat3 and lowered that of Th (encoding tyrosine hydroxylase). Tyrosine hydroxylase is necessary for catecholamine synthesis. Decline in adrenergic hormone synthesis might substantially reduce catecholamine mediated-energy expenditure and thus contribute to obesity in subjects carrying the intron 1 polymorphic variant of FTO associated with obesity. Church et al extended these studies to identify possible target genes of FTO that regulate fatty acid synthesis and degradation and energy metabolism.8 Future studies will be directed to deciphering the cellular mechanisms by which FTO regulates energy metabolism and body weight.
In humans, the increased adiposity of patients with polymorphic variants in intron 1 of FTO associated with obesity has been ascribed to increased appetite (decreased satiety) and caloric intake rather than to reduced energy utilization.10 The experimental studies demonstrate that polymorphic variants of FTO associated with obesity likely reflect increased FTO activity, while those linked to resistance to weight gain probably attenuate FTO expression.
The FTO was identified as a new obesity candidate by a GWA study by Frayling et al11 in 2007. They found a strong association between SNPs (eg, rs9939609) and adiposity in the first intron of FTO. The predisposition to obesity conferred by this gene was not related to the regulation of energy expenditure, but was mainly accounted for the control of intake of food of high caloric density.12 The FTO gene rs9939609 obesity-risk allele has also been found to be associated with the loss of control over eating.13 Given the findings of these and other studies of the molecular physiology of weight regulation (some described by Allen Root above), excess food intake (rather than reduced basal energy expenditure) seems to be the major mechanism for obesity in humans. However, reduced energy expenditure in the pathogenesis of obesity should not be underestimated. In an experimental setup we showed that non-human primates (Bonnet Macaque) who spontaneously developed obesity had reduced energy expenditure compared with their non-obese controls.14
GWA studies, in which hundreds of thousands of SNPs are tested for association with a disease in hundreds or thousands of persons, have revolutionized the search for genetic influences on complex traits. 15,16 The importance for medicine of GWAs were highlighted in the paper by Christensen and Murray.17 In the past 5 years GWA studies have identified SNPs implicating hundreds of robustly replicated loci (ie, specific genomic locations) for common traits. Nearly 600 GWA studies covering 150 distinct diseases and traits have been published, with nearly 800 SNP-trait associations reported as significant. The GWAs reported through March 2010 are available within the full text of the article by Manolio and colleagues.18 The reader is encouraged to review the paper in relation to the assessment of risk of disease19 as well as the series of 3 articles by Attia and colleagues regarding the basic concepts of genetic associations.20-22
Fima Lifshitz, MD
References - (linked to )