Nescient helix-loop-helix 2 interacts with signal transducer and activator of transcription 3 to regulate transcription of prohormone convertase 1/3.

Mechanisms controlling body weight involve gene regulation through the activation of signal transduction pathways. The Janus kinase/signal transducer and activator of transcription (STAT) signal transduction pathway is the mechanism primarily used by leptin in the hypothalamus. The transcription factor nescient helix-loop-helix 2 (Nhlh2) is a downstream target of leptin signaling and is expressed in proopiomelanocortin arcuate neurons. Proopiomelanocortin is cleaved by prohormone convertase 1/3 (PC1/3) to produce peptides that regulate the body's response to energy availability. Previous studies show that the PC1/3 promoter contains STAT3 sites mediating leptin-induced PC1/3 expression, and that Nhlh2 is required for hypothalamic PC1/3 expression because Nhlh2 knockout mice have reduced PC1/3 mRNA levels. Studies herein reveal that leptin-induced PC1/3 gene expression is abrogated in N2KO mice, and that in a hypothalamic cell line both STAT3 and Nhlh2 are required for the full transcriptional response of a PC1/3 reporter gene after leptin stimulation. Furthermore, it is shown that Nhlh2 binds to E-box motifs found adjacent to STAT3 sites in the PC1/3 promoter both in vitro and in chromatin immunoprecipitation assays. Finally, two different protein-protein interaction assays confirm the presence of a STAT3:Nhlh2 heterodimer on the PC1/3 promoter. The Nhlh2:STAT3 heterodimer may be an important transcriptional regulator of other hypothalamic genes in the leptin signaling pathway. These data confirm Nhlh2 as an integral element of the Janus kinase/STAT signaling pathway and are the first to demonstrate coordinated control of PC1/3 transcription by Nhlh2 and STAT3 after leptin stimulation.

Nhlh2: a basic helix-loop-helix transcription factor controlling physical activity.

In mice, targeted deletion of the basic helix-loop-helix transcription factor, nescient helix-loop-helix 2 (Nhlh2), leads to adult-onset obesity and reduced physical activity. We propose the novel hypothesis that transcriptional activity by Nhlh2 (NHLH2 in humans) controls either the ability or the motivation for exercise.

Energy balance pathways converging on the Nhlh2 transcription factor.

Multiple regulatory pathways exist to control the expression levels of neuropeptides in response to body weight and energy availability changes. Since many neuropeptides are first synthesized in a pro-neuropeptide form, the availability of processing enzymes in a neuron can control the amount of active mature neuropeptide produced at any given time. In this review, we will focus on the regulation of prohormone convertase 1 (PC1) and prohormone convertase 2 (PC2), as well as downstream neuropeptide genes. Evidence from our laboratory suggests that Nescient helix-loop-helix 2 (Nhlh2) regulates the transcription of PC1 and PC2, possibly in conjunction with the leptin-stimulated transcription factor, STAT3. Furthermore, Nhlh2 itself is a target of leptin and other energy availability signals, with high levels of expression during energy surplus, and low levels of expression in conditions of reduced energy availability such as food deprivation or cold exposure. Overall, coordinate regulation of Nhlh2, PC1, PC2 and downstream hypothalamic neuropeptides such as thyrotropin releasing hormone (TRH) and pro-opiomelanocortin (POMC) does lead to energy balance modulation and ensuing long-term changes in body weight.

Deletion of Nhlh2 results in a defective torpor response and reduced Beta adrenergic receptor expression in adipose tissue.

BACKGROUND: Mice with a targeted deletion of the basic helix-loop-helix transcription factor, Nescient Helix-Loop-Helix 2 (Nhlh2), display adult-onset obesity with significant increases in their fat depots, abnormal responses to cold exposure, and reduced spontaneous physical activity levels. These phenotypes, accompanied by the hypothalamic expression of Nhlh2, make the Nhlh2 knockout (N2KO) mouse a useful model to study the role of central nervous system (CNS) control on peripheral tissue such as adipose tissue. METHODOLOGY: Differences in body temperature and serum analysis of leptin were performed in fasted and ad lib fed wild-type (WT) and N2KO mice. Histological analysis of white (WAT) and brown adipose tissue (BAT) was performed. Gene and protein level expression of inflammatory and metabolic markers were compared between the two genotypes. PRINCIPAL FINDINGS: We report significant differences in serum leptin levels and body temperature in N2KO mice compared with WT mice exposed to a 24-hour fast, suggestive of a defect in both white (WAT) and brown adipose tissue (BAT) function. As compared to WT mice, N2KO mice showed increased serum IL-6 protein and WAT IL-6 mRNA levels. This was accompanied by slight elevations of mRNA for several macrophage markers, including expression of macrophage specific protein F4/80 in adipose, suggestive of macrophage infiltration of WAT in the mutant animals. The mRNAs for beta3-adrenergic receptors (beta3-AR), beta2-AR and uncoupling proteins were significantly reduced in WAT and BAT from N2KO mice compared with WT mice. CONCLUSIONS: These studies implicate Nhlh2 in the central control of WAT and BAT function, with lack of Nhlh2 leading to adipose inflammation and altered gene expression, impaired leptin response to fasting, all suggestive of a deficient torpor response in mutant animals.