Since traditional weight loss strategies have been unsuccessful at slowing the increasing rate of obesity, alternative strategies are in high demand. Brown adipose tissue (BAT) is a thermogenic tissue that efficiently expends energy. Thermogenesis in BAT occurs through the action of uncoupling protein 1 (UCP1). Identifying novel transcriptional regulators of UCP1 may lead to potential targets to increase BAT activity since its expression is under tight transcriptional control. Engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) is a modified CRISPR/Cas9 methodology that identifies novel transcriptional regulators. EnChIP uses dCas9 (lacks endonuclease activity) and guide RNAs (gRNAs) targeting a specific promoter region. The DNA-bound dCas9 can be immunoprecipitated, and the associated promoter region/transcriptional machinery can be sent for proteomic analysis to identify novel transcriptional regulators. Following β-adrenergic stimulation, which activates UCP1 gene transcription in brown adipocytes, we performed enChIP by using five gRNAs targeting the UCP1 promoter. EnChIP analysis of the UCP1 promoter revealed cysteine-rich protein 2 (CRIP2) as a putative transcriptional regulator of UCP1, which was validated by ChIP-PCR. We demonstrated that knocking down CRIP2 in brown adipocytes significantly decreased β-adrenergic-induced UCP1 expression and heat production. Mechanistically, we demonstrated that following β-adrenergic stimulation, CRIP2 was phosphorylated and translocated from the cytosol to the nucleus, where it interacted with peroxisomal proliferator-activated receptor gamma (PPARγ). Taken together, using the unbiased CRISPR-based enChIP approach, we identify CRIP2 as a novel transcriptional regulator that modulates UCP1 expression in response to β-adrenergic stimulation. These findings pave the foundation for developing new anti-obesity and antidiabetes therapies.


J. Darcy: None. C. Wang: None. M. Lundh: Employee; Self; Gubra. F. Shamsi: None. M. Lynes: None. T. d'Aprigny: None. B. Emanuelli: None. Y. Tseng: Other Relationship; Self; Chugai Pharmaceutical Co., Ltd.


National Institutes of Health (R01DK077097, R01DK102898, T32DK007260)

Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered. More information is available at