Nescient Helix Loop Helix-2 (NHLH2) is a basic helix-loop-helix transcription factor, which has been implicated, using mouse knockouts, in adult body weight regulation and fertility. A scan of the known single nucleotide polymorphisms (SNPs) in the NHLH2 gene revealed one in the 3’ untranslated region (3’UTR), which lies within an AUUUA RNA stability motif. A second SNP is nonsynonymous within the coding region of NHLH2, and was found in a genome-wide association study for obesity. Both of these SNPs were examined for their effect on NLHL2 by creating mouse mimics and examining mRNA stability, and protein function in mouse hypothalamic cell lines. The 3’UTR SNP causes increased instability and, when the SNP-containing Nhlh2 3’UTR is attached to luciferase mRNA, reduced protein levels in cells. The nonsynonymous SNP at position 83 in the protein changes an alanine residue, conserved in NHLH2 orthologs through to Drosphilia sp. to a proline residue. This change affects migration of the protein on an SDS-PAGE gel, and appears to alter secondary structure of the protein, as predicted using in silico methods. These results provide functional information on two rare human SNPs in the NHLH2 gene. One of these has been linked to human obese phenotypes, while the other is present in a relatively high proportion of individuals. Given their effects on NHLH2 protein levels, both SNPs deserve further analysis in whether they are causative and/or additive for human body weight and fertility phenotypes.

Mice with a deletion of the hypothalamic basic helix-loop-helix transcription factor Nhlh2 display adult onset obesity. We have previously shown that Nhlh2 expression is induced by leptin. In this study, we identify a small proximal leptin-responsive promoter region in the Nhlh2 gene. This 163 bp promoter contains five putative binding sites for the leptin-activated Stat3 transcription factor, and two putative binding sites for the NFκB transcription factor. Results of mutagenesis studies reveal that deletion of the NFκB sites have little effect, mutagenesis of the third Stat3 site eliminates both leptin-induced and basal expression of Nhlh2. Mutagenesis of the 4th and 5th sites eliminates leptin-induced expression, and increases basal expression above the WT promoter. Stat3 can be preferentially pulled down from leptin-treated mouse hypothalamic chromatin extracts. This study identifies leptin-induced Stat3 transcription factor as the major transcriptional regulator of Nhlh2. As Nhlh2 transcriptionally regulates genes within the melanocortin pathway, these findings have implications for human body weight control.

This experiment was conducted to investigate the effect of the inclusion of saltbush (Atriplex sp.) on the productive performance and blood mineral profile of fattening lambs. Eighteen Awassi lambs, 70 +/- 3 days of age and 23 +/- 1.8 kg body weights, were randomly selected and divided into three equal groups. Lambs in group one (control) were offered 0.2 kg/day shredded straw of barley (tibin) as the sole source of roughage, while lambs in treatment groups 2 and 3 were provided either 0.2 kg/day Atriplex nummularia or Atriplex halimus as their sole roughage content for 10 weeks. All lambs were also provided 1.1 kg/day of concentrate ration. Although lambs fed A. halimus had significantly (P < 0.05) lower dry matter intake than the control group, the daily weight gain of the two groups of lambs were not significantly different (P > 0.05). Lambs fed either A. nummularia or A. halimus had significantly lower (P < 0.05) calcium and copper levels but significantly higher (P < 0.05) cobalt levels in their blood serum than the control group. Zinc blood serum level was not affected (P > 0.05) by the treatment diets. These results suggested that A. nummularia and A. halimus can substitute for barley straw in the ration of fattening Awassi lambs without compromising lambs weight gain; however, special attention should be given to the mineral status in lambs if saltbush is a major feed component for prolonged periods of time.

No abstract