miR-133 links to energy balance through targeting Prdm16

Adipose (fat) tissues contain white, brown, and beige adipocytes. White adipocytes store extra energy in the form of lipids; ectopic and excessive accumulation of white adipose in the body leads to overweight and obesity, which often causes insulin resistance and Type 2 diabetes. Brown adipocytes, in contrast, contain hundreds of mitochondria per cell, dissipate energy into heat, and are negatively correlated to body weight and insulin resistance (Wu et al., 2013). Under cold temperature or β-adrenergic drug stimulation, a population of ‘beige’ or ‘brite’ adipocytes can be formed in the subcutaneous white adipose, containing intermediate levels of mitochondria and consume energy (Wu et al., 2013; Ye et al., 2013). Both brown and beige adipocytes are considered as good fats; and expansion or activation of these types of adipocytes thus represents a novel therapeutic strategy against obesity and diabetes.

Among many molecules that affect brown adipocyte biogenesis, Prdm16 is a key transcriptional regulator that drives the formation of both brown and beige adipocytes (Seale et al., 2008). Prdm16 also determines the bi-directional fate switch of muscle cells and brown adipocytes. Overexpression of Prdm16 transforms white adipocytes and myoblasts into brown adipocytes, while loss of Prdm16 conversely switches brown preadipocytes into muscle cells (Seale et al., 2008). Thus, Prdm16 represents a novel molecular nexus that can be stimulated to expand brown/beige adipocytes and increases energy expenditures. For example, the major Type 2 diabetic drug Rosiglitazone functions by stabilizing Prdm16 protein and promoting brown fat gene program in white adipocytes (Ohno et al., 2012). Mechanistically, Prdm16 functions as a PPARγ2 co-activator and defines a specific set of PPARγ2 transcriptional targets involved in brown adipogenesis. Given the importance of Prdm16 in brown adipocyte biogenesis, it is critical to understand how Prdm16 is regulated at the transcriptional and/or post-transcriptional levels. Recently, three groups have independently reported that miR-133a/b regulates Prdm16 expression and play roles in energy balance (Trajkovski et al., 2012; Liu et al., 2013; Yin et al., 2013).

Trajkovski et al. (2012) screened adipose microRNAs that respond to cold environment in a mouse model. The mice were maintained at 8°C for 24 h, whereas the control mice were housed under room temperature. Both miR-133a and miR-133b were downregulated ∼5 folds in brown adipose by cold exposure. The data indicated a possible physiological role of miR-133 in brown adipocyte function, especially in maintaining the body temperature. Indeed, with the use of immortalized brown adipocyte cell (PIBA), Trajkovski and colleagues showed that overexpression of miR-133 decreased brown adipogenesis, while inhibition of miR-133 increased brown adipocyte gene program including Prdm16, Ucp1, and Pparα. Therefore, miR-133 is an inhibitor of brown adipocyte differentiation. In an effort in looking for miR-133 target in brown adipocytes, a conserved binding site for miR-133 was found within the 3′ untranslated region (UTR) of Prdm16 mRNA. Luciferase assay showed that miR-133a and miR-133b recognized the 3′UTR of Prdm16. The increased brown adipogenesis by miR-133 inhibition could be fully rescued by knockdown of Prdm16. This study clearly documents the regulatory role of miR-133 in adipocytes through Prdm16; however, the functional significance of miR-133 in vivo is unexplored.

Meanwhile, Yin et al. (2013) reported that miR-133 determines the cell-fate choice of muscle stem cells through targeting Prdm16. Muscle stem cells are universally labeled by Pax7 expression. Yin and colleagues found that a small portion (0.1%) of Pax7-expressing cells do not form the muscle but form brown adipocytes expressing Prdm16 and Ucp1. What factors determine the cell-fate switch from muscle progenitors to brown adipocytes? RNA-Seq results showed that both miR-133a and miR-133b were differentially expressed between muscle progenitors and brown preadipocytes and could potentially target the Prdm16 mRNA. During skeletal muscle regeneration, inhibition of miR-133 by antisense oligonucleotide (ASO) increased Prdm16 expression in muscle progenitor cells and led to the formation of brown adipocytes in muscles, as evidenced by lipid accumulation and UCP1 expression (Figure 1A). Lineage tracing experiment showed that the newly formed brown adipocytes were converted from Pax7-expressing muscle stem cells. The brown adipose within skeletal muscles could efficiently consume glucose as manifested by [¹⁸F]-FDG PET imaging, suggesting that the newly formed brown fats were metabolically active. Strikingly, mice with ectopic brown adipose in the muscle were more resistant to high-fat diet-induced body weight gain in a 26-week observation and had improved glucose tolerance. This study elegantly demonstrates that miR-133 has a physiological role in the muscle; pharmacological inhibition of miR-133 results in a cell-fate change from muscle cells to brown adipocytes.

Figure 1.

miR-133 links to energy balance through targeting Prdm16. (A) In regenerative skeletal muscles, the muscle stem cells are activated and miR-133 is dramatically upregulated, which represses Prdm16 expression. The muscle progenitors proliferate and differentiate into myotubes. Administration of ASO against miR-133 decreases miR-133 expression, resulting in Prdm16 upregulation in muscle progenitors. The muscle progenitors switch the cell-fate from myocytes to brown adipocytes, which consume glucose and improve insulin sensitivity. (B) Along the commitment and differentiation of brown adipocyte progenitors, a decreased level of miR-133 leads to the upregulation of Prdm16, stimulating the biogenesis of brown adipocytes. When miR-133 is overexpressed, Prdm16 decreases and accordingly the generated adipocytes are less thermogenic. (C) Loss of miR-133a in white adipose promotes Prdm16 upregulation, which drives the formation of beige adipocytes in subcutaneous white adipose. The beige adipocytes increase thermogenesis and dissipate energy into heat, which leads to better insulin sensitivity and decreased fasting glucose levels.

Consistently, Liu et al. (2013) demonstrated the in vivo role of miR-133a in the browning of white adipocytes in a mouse model. A decline of miR-133a expression was accompanied by an increase of Prdm16 along the commitment and differentiation of brown preadipocytes (Figure 1B). Such a correlation fits very well with the well-established ‘microRNA-mRNA’ mutual exclusion model. Using a mouse model that has 75% reduction in miR-133a expression, Liu and colleagues found that miR-133a knockdown mice developed multilocular UCP1-positive beige adipocytes that were interspersed in the subcutaneous white fat (Figure 1C). At the molecular level, the mice exhibited elevated expression of brown, thermogenic, and lipolysis gene programs in subcutaneous fats. After 5 days of exposure at 4°C, miR-133a knockdown mice more robustly activated the thermogenic and lipolysis gene programs in subcutaneous adipose. These data suggest that miR-133a knockdown mice have elevated thermogenesis and energy expenditure. Consequently, these mice have much lower fasting glucose levels and increased insulin sensitivity compared with wild-type mice. This study provides the genetic evidence to support the miR-133-Prdm16 axis as a central regulator of adipocyte energy expenditure.

The three studies together elucidate a crucial role of miR-133 in the regulation of Prdm16, adipose browning, glucose metabolism, and energy balance, suggesting that miR-133 can be a potential drug target. As miR-133 is necessary for proper development and function of both cardiac and skeletal muscles, the side effects of miR-133 inhibition on these muscles must be taken into account. Luckily, while miR-133a knockdown mice improve insulin sensitivity and glucose disposal, no obvious cardiac and skeletal muscle defects are observed in these mice. Future studies are necessary to determine whether inhibition of miR-133 similarly targets Prdm16 in human adipocytes. If so, small molecules could be designed to specifically inhibit miR-133 generation or maturation for the treatment of obesity and Type 2 diabetes.

[The study was supported by funding from the National Institute of Health (NIH R01AR060652) and United States Department of Agriculture (USDA 2009-35206-05218) to S.K.]