Functional Inactivation of the Genome-Wide Association Study Obesity Gene Neuronal Growth Regulator 1 in Mice Causes a Body Mass Phenotype

Abstract

To date, genome-wide association studies (GWAS) have identified at least 32 novel loci for obesity and body mass-related traits. However, the causal genetic variant and molecular mechanisms of specific susceptibility genes in relation to obesity are yet to be fully confirmed and characterised. Here, we examined whether the candidate gene NEGR1 encoding the neuronal growth regulator 1, also termed neurotractin or Kilon, accounts for the obesity association. To characterise the function of NEGR1 for body weight control in vivo, we generated two novel mutant mouse lines, including a constitutive NEGR1-deficient mouse line as well as an ENU-mutagenised line carrying a loss-of-function mutation (Negr1-I87N) and performed metabolic phenotypic analyses. Ablation of NEGR1 results in a small but steady reduction of body mass in both mutant lines, accompanied with a small reduction in body length in the Negr1-I87N mutants. Magnetic resonance scanning reveals that the reduction of body mass in Negr1-I87N mice is due to a reduced proportion of lean mass. Negr1-I87N mutants display reduced food intake and physical activity while normalised energy expenditure remains unchanged. Expression analyses confirmed the brain-specific distribution of NEGR1 including strong expression in the hypothalamus. In vitro assays show that NEGR1 promotes cell-cell adhesion and neurite growth of hypothalamic neurons. Our results indicate a role of NEGR1 in the control of body weight and food intake. This study provides evidence that supports the link of the GWAS candidate gene NEGR1 with body weight control.

Approach to assessing determinants of glucose homeostasis in the conscious mouse

“…ENU-induced mutagenesis is a strong method in that it can generate a wide array of subtle phenotypes and the resulting lines may more closely recapitulate the human phenotype compared to engineered knock-out models. Diet challenges are now being employed in this model to identify genetic and environmental interactions in the pathogenesis of metabolic disease (Lee et al 2012; Svenson et al 2008). …”

Functional Analysis of Seven Genes Linked to Body Mass Index and Adiposity by Genome-Wide Association Studies: A Review

“…The metabolic rate of the Kilon/NEGR1 KO mouse was lower than that of wild-type mice, but normalising for lean body mass completely removed this effect, implying no impact of the gene on energy expenditure [56] . Although division by lean mass was perhaps not the best way to normalise such data [7,119] , the fact that it removed the apparent impact of the gene on energy expenditure is striking ( fig.…”

“…Localised expression of Kilon/NEGR1 (and OBCAM), particularly in dendrites, might suggest that these molecules play an important role in rearranging dendritic connectivity of these neurons. Using an in vitro assay with cells derived from mice that had the Kilon/ NEGR1 gene knocked out, it has been shown that Kilon/ NEGR1 plays a key role in both cell-cell adhesion and neurite outgrowth in hypothalamic neurons [56] . AVP and OXT have both been implicated in the regulation of energy balance.…”

“…Lee et al [56] generated a constitutive Kilon/NEGR1-deficient mouse. Mice with the Kilon/NEGR1 gene ablated weaned (day 20) about 25% lighter than wild-type or heterozygous mice.…”

“…Interestingly, the difference in body composition between the wild-type and KO mice was solely due to the difference in lean mass rather than a difference in fat mass ( fig. 2 ) [56] . Because the mice had equal body fat contents but different lean masses, the percentage of body fatness was elevated in the mice with the loss of the Kilon/NEGR1 gene.…”