Brown adipocytes local response to thyroid hormone is required for adaptive thermogenesis in adult male mice
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eLife assessment
The manuscript by Yanis Zekri et al identifies the direct T3 target genes that are important in thyroid hormone signaling in brown adipose tissue (BAT). The findings reported in this manuscript are significant and fundamental to our understanding of thyroid hormone action in response to environmental changes. The strength of the evidence presented with the novel methodological approaches used makes the manuscript exceptional in the area of BAT biology and T3 regulation of adaptive thermogenesis.
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Abstract
Thyroid hormone (T3) and its nuclear receptors (TR) are important regulators of energy expenditure and adaptive thermogenesis, notably through their action in the brown adipose tissue (BAT). However, T3 acts in many other peripheral and central tissues which are also involved in energy expenditure. The general picture of how T3 regulates BAT thermogenesis is currently not fully established, notably due to the absence of extensive omics analyses and the lack of specific mice model. Here, we first used transcriptome and cistrome analyses to establish the list of T3/TR direct target genes in brown adipocytes. We then developed a novel model of transgenic mice, in which T3 signaling is specifically suppressed in brown adipocytes at adult stage. We addressed the capacity of these mice to mount a thermogenic response when challenged by either a cold exposure or a high-fat diet, and analyzed the associated changes in BAT transcriptome. We conclude that T3 plays a crucial role in the thermogenic response of the BAT, controlling the expression of genes involved in lipid and glucose metabolism and regulating BAT proliferation. The resulting picture provides an unprecedented view on the pathways by which T3 activates energy expenditure through an efficient adaptive thermogenesis in the BAT.
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Author Response
Reviewer #1 (Public Review):
Yanis Zekri et al have addressed an important question of the possible role of thyroid hormone (T3) and its nuclear receptor (TR) on local BAT thermogenesis and energy expenditure. In this well-written manuscript and well-carried work, the authors address the above question by A) by generating the BATKO mice by selectively eliminating TR signaling in BAT by knocking-in a TRα1L400R, a dominant negative version of the TRa1 receptor, and by floxing the ThRb gene. They characterized this mouse thoroughly to show that they totally lacked T3 responsiveness. Using qPCR they evaluated the selective abrogation of Thrb and Hr expression in BAT tissue relative to other tissue sites. B) Using time-course transcriptome analysis they then go on to enlist all the T3/TR direct target genes using …
Author Response
Reviewer #1 (Public Review):
Yanis Zekri et al have addressed an important question of the possible role of thyroid hormone (T3) and its nuclear receptor (TR) on local BAT thermogenesis and energy expenditure. In this well-written manuscript and well-carried work, the authors address the above question by A) by generating the BATKO mice by selectively eliminating TR signaling in BAT by knocking-in a TRα1L400R, a dominant negative version of the TRa1 receptor, and by floxing the ThRb gene. They characterized this mouse thoroughly to show that they totally lacked T3 responsiveness. Using qPCR they evaluated the selective abrogation of Thrb and Hr expression in BAT tissue relative to other tissue sites. B) Using time-course transcriptome analysis they then go on to enlist all the T3/TR direct target genes using well-defined criteria and further linking with their ChipSeq data they identified 639 putative target genes which are under the direct control of T3/TR signaling. Interestingly their gene analysis lead them to some target genes directly involved with UCP1 and PGC1α in addition to genes of many other metabolic processes related to BAT thermogenesis. The experiments on denervated BAT on wild-type PTU-fed was a rather neat experiment to eliminate the influence of noradrenergic terminal BAT target genes. Furthermore, the cold exposure experiments and the high-fat diets feeding with the series of complex analyses led them to the conclusion that BAT KO animals suffered from reduced efficiency of BAT adaptive thermogenesis. By comparing the BAT transcriptome of BATKO and CTRL mice after 24h at 4{degree sign}C, the authors further go on to show how BAT TR signaling controls other subsets of genes, especially a wide variety of metabolic regulations, especially lipolysis/fatty acid oxidation. Finally, EdU injection experiments showed a direct effect of T3 on BAT proliferation.
I think it was well thought and well-designed study for understanding the complex action of cell-autonomous T3 regulation of adaptive thermogenesis. The conclusions of this paper are well supported by the data provided.
Thank you very much for this very pertinent and kind summary of our work.
Reviewer #2 (Public Review):
The authors designed this study to identify the direct T3 target genes that underlie the T3 actions in the brown adipose tissue (BAT). The unique model used (dominant negative TRa knock-in and a TRb knock-out) allows for the isolation of BAT-specific actions from other well-known systemic effects on thermogenesis, including the central nervous system. The strengths of the study reside in the novel methodological approach. Previous studies of T3 actions in the BAT used animal models that did not allow for full isolation of BAT-specific effects of T3. A limitation however is the combination of TRa knock-in (which causes permanent suppression of TRa-dependent genes) with the TRb knockout, which only prevents T3 induction of TRb-dependent genes. Nonetheless, the results were impressive with the identification of about 1,500 genes differentially expressed in the BAT, among which UCP1 and PGC1a were the two main ones. Although it has been known that both UCP1 and PGC1a are downstream targets of T3, the work establishes through an ingenious approach the critical direct role played by T3 in BAT thermogenesis. In addition, the genetic approach utilized here is of great value and could be easily expanded to other tissues and systems.
Thank you for this very pertinent summary of our work. We just want to clarify one point: we do not believe that Ucp1 is, quantitatively, one of the main genes regulated by T3 in the BAT. First, it does not belong to the set of the most induced genes after T3/T4 injection in mice. Most importantly, Ucp1 expression was not altered in BATKO mice exposed at 4°C according to unbiased RNAseq analysis. Only targeted qRT-PCR analysis could evidence a modest change. We do not call into question the crucial role of Ucp1 in BAT thermogenesis. However, we think that our approach put into perspective the relevance of Ucp1 in the T3-dependent control of BAT thermogenesis, suggesting that other mechanisms might be more directly linked to T3 activity.
Reviewer #3 (Public Review):
This paper details the importance of thyroid hormone signaling in BAT in response to environmental and nutritional stress. The authors utilize a novel genetic model to specifically target BAT and impair thyroid hormone signaling. The physiologic insight is of great interest. The role of the sympathetic nervous system in the BAT response is not fully addressed but it appears that cell-autonomous signaling mediates TH signaling in response to hyperthyroidism. The link cistromically between the TR and PGC1 is also novel and of interest.
Thank you very much for your kind comments that are highly appreciated.
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eLife assessment
The manuscript by Yanis Zekri et al identifies the direct T3 target genes that are important in thyroid hormone signaling in brown adipose tissue (BAT). The findings reported in this manuscript are significant and fundamental to our understanding of thyroid hormone action in response to environmental changes. The strength of the evidence presented with the novel methodological approaches used makes the manuscript exceptional in the area of BAT biology and T3 regulation of adaptive thermogenesis.
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Reviewer #1 (Public Review):
Yanis Zekri et al have addressed an important question of the possible role of thyroid hormone (T3) and its nuclear receptor (TR) on local BAT thermogenesis and energy expenditure. In this well-written manuscript and well-carried work, the authors address the above question by A) by generating the BATKO mice by selectively eliminating TR signaling in BAT by knocking-in a TRα1L400R, a dominant negative version of the TRa1 receptor, and by floxing the ThRb gene. They characterized this mouse thoroughly to show that they totally lacked T3 responsiveness. Using qPCR they evaluated the selective abrogation of Thrb and Hr expression in BAT tissue relative to other tissue sites. B) Using time-course transcriptome analysis they then go on to enlist all the T3/TR direct target genes using well-defined criteria and …
Reviewer #1 (Public Review):
Yanis Zekri et al have addressed an important question of the possible role of thyroid hormone (T3) and its nuclear receptor (TR) on local BAT thermogenesis and energy expenditure. In this well-written manuscript and well-carried work, the authors address the above question by A) by generating the BATKO mice by selectively eliminating TR signaling in BAT by knocking-in a TRα1L400R, a dominant negative version of the TRa1 receptor, and by floxing the ThRb gene. They characterized this mouse thoroughly to show that they totally lacked T3 responsiveness. Using qPCR they evaluated the selective abrogation of Thrb and Hr expression in BAT tissue relative to other tissue sites. B) Using time-course transcriptome analysis they then go on to enlist all the T3/TR direct target genes using well-defined criteria and further linking with their ChipSeq data they identified 639 putative target genes which are under the direct control of T3/TR signaling. Interestingly their gene analysis lead them to some target genes directly involved with UCP1 and PGC1α in addition to genes of many other metabolic processes related to BAT thermogenesis. The experiments on denervated BAT on wild-type PTU-fed was a rather neat experiment to eliminate the influence of noradrenergic terminal BAT target genes. Furthermore, the cold exposure experiments and the high-fat diets feeding with the series of complex analyses led them to the conclusion that BAT KO animals suffered from reduced efficiency of BAT adaptive thermogenesis. By comparing the BAT transcriptome of BATKO and CTRL mice after 24h at 4{degree sign}C, the authors further go on to show how BAT TR signaling controls other subsets of genes, especially a wide variety of metabolic regulations, especially lipolysis/fatty acid oxidation. Finally, EdU injection experiments showed a direct effect of T3 on BAT proliferation.
I think it was well thought and well-designed study for understanding the complex action of cell-autonomous T3 regulation of adaptive thermogenesis. The conclusions of this paper are well supported by the data provided.
-
Reviewer #2 (Public Review):
The authors designed this study to identify the direct T3 target genes that underlie the T3 actions in the brown adipose tissue (BAT). The unique model used (dominant negative TRa knock-in and a TRb knock-out) allows for the isolation of BAT-specific actions from other well-known systemic effects on thermogenesis, including the central nervous system. The strengths of the study reside in the novel methodological approach. Previous studies of T3 actions in the BAT used animal models that did not allow for full isolation of BAT-specific effects of T3. A limitation however is the combination of TRa knock-in (which causes permanent suppression of TRa-dependent genes) with the TRb knockout, which only prevents T3 induction of TRb-dependent genes. Nonetheless, the results were impressive with the identification of …
Reviewer #2 (Public Review):
The authors designed this study to identify the direct T3 target genes that underlie the T3 actions in the brown adipose tissue (BAT). The unique model used (dominant negative TRa knock-in and a TRb knock-out) allows for the isolation of BAT-specific actions from other well-known systemic effects on thermogenesis, including the central nervous system. The strengths of the study reside in the novel methodological approach. Previous studies of T3 actions in the BAT used animal models that did not allow for full isolation of BAT-specific effects of T3. A limitation however is the combination of TRa knock-in (which causes permanent suppression of TRa-dependent genes) with the TRb knockout, which only prevents T3 induction of TRb-dependent genes. Nonetheless, the results were impressive with the identification of about 1,500 genes differentially expressed in the BAT, among which UCP1 and PGC1a were the two main ones. Although it has been known that both UCP1 and PGC1a are downstream targets of T3, the work establishes through an ingenious approach the critical direct role played by T3 in BAT thermogenesis. In addition, the genetic approach utilized here is of great value and could be easily expanded to other tissues and systems.
-
Reviewer #3 (Public Review):
This paper details the importance of thyroid hormone signaling in BAT in response to environmental and nutritional stress. The authors utilize a novel genetic model to specifically target BAT and impair thyroid hormone signaling. The physiologic insight is of great interest. The role of the sympathetic nervous system in the BAT response is not fully addressed but it appears that cell-autonomous signaling mediates TH signaling in response to hyperthyroidism. The link cistromically between the TR and PGC1 is also novel and of interest.
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