Transcriptional Control of Obesity

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Although over 20 neuronally-expressed genes have been implicated in the regulation of body weight, the transcriptional and post-transcriptional mechanisms through which these genes are expressed in response to changes in energy balance remain unclear. Recent evidence from my laboratory suggests that the NhIh2 transcription factor plays a key role in the regulation of hypothalamic genes controlling body weight. NhIh2 knockout mice (N2KO) display adult-onset obesity and NhIh2 is expressed in the hypothalamic regions that control appetite-including the arcuate ncuelus (ARC) of the hypothalamus. In dual-label in situ hybridization studies, we found that nearly all pro- opiomelanocortin (POMC) neurons in the ARC expressed NhIh2. In these neurons, the POMC precursor protein is processed to alpha- melanocyte stimulating hormone (alphaMSH), a peptide that regulates food intake and energy expenditure. In studies comparing POMC mRNA and peptide levels in N2KO and normal mice, we found that while POMC mRNA levels were identical between the two groups, levels of two processed forms of POMC, beta-endorphin (betaEND) and adrenocorticotropin hormone (ACTH) were dramatically reduced, while levels of alphaMSH were modestly reduced in the N2KO animals. These findings suggest that N2KO animals could have a defect that reduces processing of the POMC precursor. Indeed, mRNA levels for two of the processing enzymes necessary for cleaving POMC into bioactive peptides prohormone convertases I (PC1) and II (PC2) were reduced up to 80 percent in N2KO animals. Thus, it is likely that PC1 and PC2 are direct gene regulatory targets of the NhIh2 transcription factor and that a reduction in fully processed neuropeptides contributes to obesity in the N2KO animals. In addition, we predict that NhIh2 is a direct target of leptin-signaling pathways, as it is expressed in the leptin-receptor containing POMC neurons and contains STAT motifs in its promoter. Based on these preliminary findings, we propose that positive energy balance increases the expression of NhIh2, which in turn increases the transcription rate of the PC1 and PC2 genes. To test this hypothesis, we will ask in the first two specific aims, if NhIh2, PC1 and PC2 are increased in response to a positive energy balance and if the of the PC1 and PC2 genes is lost in N2KO animals. In specific aim 3, we will switch to a molecular investigation of the NhIh2 promoter to determine if AP1 and STAT3 transcription factors can bind to and transactivate the NhIh2 gene. We will then ask in specific aim 4 if the NhIh2 transcription factor, in cooperation with activated STAT3, binds to and transactivates the PC1 promoter. The proposed experiments use both molecular and animal-based approaches to ask a fundamental question about transcriptional and post- transcriptional mechanisms controlling body weight.