The Ablation of T1R1 Reduces Lipid Accumulation Through Reducing the De Novo Lipid Synthesis and Improving Lipid Metabolism in Mice

DOI: https://doi.org/10.21203/rs.3.rs-961608/v1

Abstract

Background: That cells sense extracellular amino acids to regulate intracellular lipid metabolism has been a heated debate in terms of the study on amino acid nutrition. T1R1 is a membrane G protein-coupled receptor that senses amino acids in a variety of cells. In our study, T1R1-KO mice was used to explore the function of umami taste receptor in lipid metabolism.

Results: Compared with wild-type mice, T1R1-KO mice showed Significantly lighter adipose tissue weight, reduced serum triglycerides (TG) and total cholesterol (TC), as well as higher glucose tolerance on chow diet. Moreover, there were less lipid accumulation in adipose and liver tissue and shrink of the adipocyte size in T1R1-KO mice. And a decreased expression of lipogenesis genes (PPARγ, CEBPα, SREBP1) was found in both adipose and liver tissue. To further study the mechanism of T1R1 regulating liver lipid metabolism, proteomics analysis was introduced and the up-regulated proteins were enriched in lipid and steroid metabolism pathways of T1R1-KO mice. Further PRM verification analysis showed that the ablation of T1R1 reduced the de novo synthesis of lipids through BCKDHA and BCKDHB, and promoted lipid metabolism through CYP7B1 and IGFBP2.

Conclusions: Our results showed that the disruption of T1R1 in mice could reduce body lipid accumulation, and our data clarifies the role of umami receptors in lipid metabolism and could provide a basis for the research on nutrition and obesity.

Introduction

Amino acids are important nutrient involved in the body’s lipid metabolism and various diseases such as obesity, type 2 diabetes, atherosclerosis, cancer, non-alcoholic fatty liver disease (NFALD) and so on [13]. So far, the studies on amino acids are mainly conducted through adding external sources or interfering with their synthesis and decomposition, whereas little is known about the mechanism by which cells sense amino acids to perform their functions [4].

Umami taste receptor T1R1/T1R3, a G protein-coupled receptor (GPCR) formed by heterodimerization of T1R1 and T1R3 (both of them belong to the T1Rs class), has been reported to act as chemoreceptors for a variety of amino acids, especially L-glutamate [5, 6]. Recently, T1R1/T1R3 has been reported to have function expression in multiple tissues and organs (such as stomach, liver, muscle, adipose tissue, testis, heart, bone, and so on), in addition to the tongue taste bud cells [7, 8]. Therefore, T1R1/T1R3, the amino acids sensing receptor, may have effect on metabolic diseases such as obesity and aging [9].

Located in gut endocrine cells, T1R1/T1R3 functions as a sensor for Phe-, Leu-, and Glu-induced CCK secretion, which makes the body feel full and inhibit food intake [10, 11]. Furthermore, T1R1/T1R3 was reported to mediate the AAs activating ERK1/2 and mTORC1 signaling pathways in pancreas MIN6 cell line [12]. What’s more, the decrease in mTORC1 activity after T1R3 knockout also appeared in skeletal muscle and heart [13]. As the central regulator of mammalian cell growth, mTORC1 acts as an important signaling pathway downstream of amino acid nutrition and regulates lipid metabolism [14, 15]. These show that umami taste receptors may have extensively potential role in body lipid metabolism.

What should be emphasized is that T1R3 can also function as the sweet taste receptor by forming a heterodimer with T1R2, except being part of the umami taste receptor [16]. Simon et al. found that the deficiency of T1R2 or T1R3 could reduce fat mass and adipocyte area without affecting food intake and body weight following western dietary intervention [16]. The elimination of T1R2 does not affect the number of peripheral adipocyte cells but reduces the number of bone marrow adipocyte cells [17]. In terms of biological process, T1R3, the sugar-sensing receptor in 3T3-L1 cells, plays anti-adipogenic role through Gas instead of cAMP [18, 19]. What’s more, the disruption of T1R2 protect mice from HF/LC diet-induced hyperinsulinemia and decreased liver TG accumulation [20]. In the pancreas, glucose promotes intracellular ATP production and insulin secretion through T1R3 [21, 22]. These demonstrate the link between T1R3 as a sugar-sensing receptor and body lipid metabolism. Given the dual role of T1R3 in sensing sugars and amino acids, umami taste receptors may also play a role in the body's lipid metabolism. Therefore, T1R1 (GPR70) knockout mice become a good model for studying the function of umami taste receptors.

T1R1 knockout mice model was made to explore the function of amino acid receptor T1R1/T1R3 in body lipid metabolism. We found that the deletion of T1R1 in mice caused the reduction of circulating TG and TC content and the lipid accumulation in adipose tissue and liver tissue. Further studies showed that the decrease in BCKDHA and BCKDHB reduced the de novo lipogenesis and the increase of CYP7B1 and IGFBP2 improved lipid metabolism in T1R1-KO mice. This study provides a valuable insight into the function of amino acid sensing receptors in lipid metabolism.

Materials And Methods

Animals

The use of all animals in this study was approved by the EAMC (Committee of Experimental Animal Management) at Northwest Agriculture and Forestry University, China. The T1R1 heterozygous C57BL/6J mice were a kind gift from by Zhang Chunlei’s laboratory, which were purchased from Jackson laboratory [16, 23]. According to Zhao’s study of T1R1 gene ablation [16], primers were designed to identify the wildtype (T1R1+/+, WT), T1R1 knockout mice (T1R1−/−, T1R1-KO) and T1R1 heterozygous mice (T1R1+/−) (shown in Supporting Information Table 1). Only WT and T1R1-KO mice were used in this study. All mice were kept in the same environment.

Glucose tolerance test (GTT), insulin sensitivity test (ITT) and tissue collection

For the glucose tolerance test, mice were given intraperitoneal injection of glucose (2g/kg body weight) after overnight fasting. For the insulin sensitivity test, mice were fasted for 6 hours and subjected to intraperitoneal infection of insulin (0.75U/kg body weight). Blood glucose concentrations were measured at 0, 15, 30, 60, 90, 120 minutes by glucometer (yuwell 560, Jiangsu, China). After overnight fasting, all mice were weighed and then we collected blood, inguinal white adipose tissue (iWAT), gonadal white adipose tissue (gWAT), brown adipose tissue (BAT), and liver samples.

Histological analysis

Samples of iWAT, gWAT and BAT were fixed with fat fixative (Servicebio) for over 24 hours followed by general paraffin section and HE staining. The adipocyte dimmer was calculated using ImajeJ(US National Institutes of Health) software for about 10 fields of view for one slice. The liver tissue used for HE staining was paraffin sectioned after being fixed with 4% paraformaldehyde, while the frozen samples were stained with Oil Red O.

Real-time quantitative PCR (RT-qPCR)

Total RNA was extracted from tissues using TRIzol reagent (Invitrogen, Carlsbad, CA). The cDNA was synthesized with reverse transcription kits (TaKaRa, Otsu, Japan) according to the manufacturer’s instructions. For RT-qPCR, SYBR Green (Vazyme, Nanjing, China) was used with the Step One Plus system (ABI, MA, USA). The primers for RT-qPCR were listed in Supporting Information Table 1.

Western blot

Briefly, 20 µg of extracted protein of tissues was run on 10% SDS-PAGE gel and immunoblotted with the primary antibodies to T1R1 (SIGMA-MLORICH, Saint Louis, USA), PPARγ (Abcam, Cambridge, UK), C/EBPα (Abcam, Cambridge, UK), p-HSL (CST, Danvers, USA), ATGL (CST, Danvers, USA), LPL (Santa Cruz, Dallas, USA), FABP4 (Santa Cruz, Dallas, USA), and GAPDH (Boster, Wuhan, China).

Proteomics Sequencing Analysis

Proteomics sequencing services were provided by Jingjie Biotechnology Co., Ltd. (Hangzhou, China). Livers from three T1R1-KO mice and WT mice which were all 16week old were tested in this experiment. Extracted the total protein and arried out trypsin digestion, TMT/iTRAQ labeling, high-performance liquid chromatog raphy classification, mass spectrometry analysis, database search, Bioinformatics analysis, and so forth.

Parallel Reaction Monitoring (PRM) Analysis

Parallel Reaction Monitoring services were provided by Jingjie Biotechnology Co., Ltd. (Hangzhou, China). The liver tissue from 3 T1R1-KO and 3 WT mice were ground and lysed to extract protein. The extracted protein was subjected to trypsin digestion and LC-MS/MS detection and analysis.

Statistical Analysis

All results were analyzed using GraphPad Prism 8 software (GraphPad Software, San Diego, USA), and the data represent the mean ± standard error of the mean (SEM). Statistical significance was determined using Student’s t-test. Statistical differences are expressed as follows: *P < 0.05, **P < 0.01, and ***P < 0.001.

Results

Systemic genetic elimination of miceT1R1 gene reduced body lipid accumulation

As the key organs for lipid storage and metabolism, the adipose and liver tissue were tested to explore the effect of T1R1 knockout on the body's lipid metabolism. The mRNA expression of T1R1 was similar among different adipose tissues (iWAT, gWAT, BAT), but significant(P<0.05) higher in liver tissue (about 42 times that of iWAT) (Fig. 1A). To explore the effect of T1R1 knockout on the body metabolism under physiological conditions, T1R1-KO mice were produced (the knockout efficiency was shown in Fig. 1B). Through continuous recording, T1R1-KO mice showed just slight rise of body weight and no difference in daily feed intake than that of the WT mice (P >0.05) (Fig. 1C-D). However, when the mice were dissected, significant reduction in iWAT and gWAT were found not only in the anatomical photos but also in the weight statistics, while the BAT and liver showed no difference (P >0.05) (Fig. 1E-G).

To further detect the effect of T1R1 ablation on the body's metabolism, the serum was collected for physical and chemical index analysis. Compared with WT mice, the level of TG and TC in T1R1-KO mice were significantly reduced. But we observed a significantly (P<0.05) increase in the high-density lipoprotein and no change(P>0.05) in glucose and Low-density lipoprotein (LDL) (Fig. 1H). What’s more, GTT and ITT showed that T1R1-KO mice had better glucose tolerance and insulin sensitivity (Fig. 1I and J) (P<0.05).

The ablation of T1R1 reduced adipogenesis process in adipose and liver tissue

To clarify the histological changes, adipose tissue and liver were paraffin sectioned and stained with HE or Oil Red O, and then the adipocytes were used for cell diameter analysis. Smaller cell diameter was detected in both iWAT and gWAT (Fig. 2A-C). Besides, the BAT and liver also showed reduced lipid accumulation (Fig. 2A and 2G). Furthermore, the key genes in the adipogenesis process (PPARγ, CEBPα, SREBP1) and lipolysis process (ATGL, LPL, HSL) were tested in iWAT and liver. For iWAT, we observed a significant (P<0.05) reduction of the mRNA levels of adipogenesis genes in T1R1-KO mice, along with either significantly (P<0.05) reduced protein levels (PPARγ, SREBP1) or no significantly (P<0.05) protein level (CEBPα) (Fig. 2D-F). While the lipolysis genes showed significantly reduced mRNA levels, but no significantly protein levels (ATGL, LPL) or significantly decreased protein level (p-HSL) (Fig. 2D-F). For liver, significantly reduced mRNA and protein levels of CEBPα and SREBP1 were detected in T1R1-KO mice, while there was no change in that of PPARγ (Fig. 2H-J). In addition, there were no difference between T1R1-KO and WT mice in the mRNA and protein levels of ATGL and LPL, except p-HSL showed a significant increase in mRNA level and no difference in protein level (Fig. 2H-J). Besides, the mTOR signaling was also reduced in T1R1-KO mice (Fig. 1B).

Up-regulated proteins identified by proteomics analysis were mainly enriched in lipid and steroid metabolism pathways

To explore the effect and mechanism of T1R1 on lipid metabolism, quantitative proteomics using liquid chromatography-tandem mass spectrometry (LC-MS) analysis was introduced in liver tissue (Fig. 3A). Principal component analysis (PCA) indicate that the data can be analyzed in the next step (Fig. 3B). About 4889 proteins were identified through mass spectrometry analysis, of which 4281 could be quantified (Fig. 3C). We filtered the proteins under the condition of 1.2 fold changes and significance P < 0.05, and 49 differentially (P<0.05) expressed proteins were screened, of which 24 were up-regulated and 25 were down-regulated (Fig. 3D and E).

To further explore the functional information of the differential proteins, we carried out location, annotation, and functional analysis. As shown in Fig. 4A, most of the differential proteins were located in nucleus (34.69%) and cytoplasm (26.53%), indicating that wild changes happened inside the cell following T1R1 ablation, even though T1R1 protein located in the membrane itself. Gene ontology (GO) enrichment analysis showed that, metabolic process was also affected except cellular process (Fig. 4B). Coordinated with the localization of T1R1 membrane receptors, the cellular component of differential proteins was also enriched in the cell membrane and extracellular region except cell and organelle (Fig. 4B). What’s more, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analysis showed that the 24 up-regulated proteins were mainly enriched in glutamatergic synapse pathway (Fig. 4C). And according to GO analysis, they were found to be mainly enriched in negative regulation of lipid metabolic process, regulation of steroid metabolic process, negative regulation of immunity and other processes embryonic development, muscle development (Fig. 4D).

CYP7B1, IGFBP2, BCKDHA and BCKDHB played important roles in the reduction of lipid deposition in T1R1 knockout mice

To further study the mechanism by which T1R1 regulates liver lipid metabolism, quantitative analysis based on PRM were performed to verify the proteins related to lipid metabolism in proteomics analysis in liver (Fig. 5A). As important regulators of cholesterol metabolism, oxysterol 7 α-hydroxylase (CYP7B1) was significantly up-regulated in T1R1-KO mice (Fig. 5B). And Insulin-like growth factor binding protein 2 (IGFBP2) was found to be significantly (P<0.05) up-regulated in T1R1-KO mice (Fig. 5C). In addition, key genes related to amino acid metabolism were also affected, of which Branched Chain Keto Acid Dehydrogenase E1 Subunit Alpha (BCKDHA) and Branched Chain Keto Acid Dehydrogenase E1 Subunit Beta (BCKDHB) were significantly (P<0.05) down-regulated (Fig. 5D and E). Additionally, there were no difference (P>0.05) of Acetoacetyl-CoA synthetase (AACS) which is a key factor that regulate lipid biosynthesis, and Malic Enzyme-1 (ME1) which produces NADPH for fatty acid and cholesterol biosynthetic (Fig. 5F and G).

Discussion

That cells sense extracellular nutrients to affect their own metabolism is a key topic for many human metabolic diseases [24]. GPCR signaling has been reported to regulate glucose and lipid metabolism and energy homeostasis in obesity and type 2 diabetes [25]. Class C GPCRs responsible for amino acid sensing, such as T1Rs, are considered as potential targets for the treatment of related diseases [26]. However, the role of T1Rs in various tissues and organs of the whole body is not completely understood [27]. Here we focused on the role of umami taste receptor T1R1/T1R3 on lipid metabolism, in order to explain the mechanism of cellular amino acid sensing and its effects.

Consistent with previous studies of T1R2-KO and T1R3-KO mice, we found T1R1-KO mice also showed insignificant body weight change, reduced fat mass and smaller adipocyte area [20]. Unlike T1R2-KO mice showing decreased liver TG level following a high-fat diet, our results showed T1R1-KO mice had lower serum TG and TC [20]. Besides, in contrast to T1R3 deleted mice exhibiting glucose intolerance and insulin resistance, it was observed that T1R1-KO mice exhibited better glucose tolerance and insulin sensitivity [28]. That’s because in addition to the function of umami taste receptors, T1R3 also forms sugar-sensitive receptors with T1R2 to perform functions, which mediates the sense of cells to glucose, and has been reported to affect the secretion of insulin by pancreatic islets to regulate sugar metabolism [ 20, 28, 29].

As the center organ of lipid metabolism, liver absorbs fatty acids from the blood and synthesizes them into triglycerides and cholesterol, the excess part of which is then stored in adipose tissue through blood circulation [15]. Therefore, when the body's lipid metabolism changes, the liver and adipose tissue are always checked first. Consistent with the adipose tissue performance of T1R2-KO and T1R3-KO mice, T1R1-KO mice also had deceased iWAT and gWAT fat mass and adipocyte area [20, 30]. Meanwhile, we found reduced lipid droplets in liver and BAT.

Generally, the decrease in lipid accumulation is attributed to the fact that the rate of lipogenesis is lower than that of lipolysis [31]. These processes are mainly regulated by a series of genes, for example the PPARγ, C/EBPα and SREBP1 in adipogenesis, and ATGL, LPL and HSL in lipolysis [3235]. Changes of these key genes in adipose and liver tissue suggested that the process of adipogenesis was significantly inhibited in T1R1-KO mice, which may partly explain the decrease in lipid deposition. Besides, consistent with the function of mTOR signaling as the downstream of T1R1/T1R3, we also found its reduction in T1R1-KO mice [13, 14].

Proteomics Sequencing analysis could explore extensive biological changes in cells or tissues, so as to easily probe the ways that genes affecting physiological changes [36]. These tools have never been used in previous studies on T1Rs family members’ function in cells or mice. Therefore, the use of proteomics sequencing was made in this study to further reveal the mechanism by which T1R1 affects body lipid metabolism. In accord with our prediction, the up-regulated proteins were enriched in processes of regulation of lipid metabolism, steroid metabolism and cellular ketone metabolism, which showed that the ablation of T1R1 may disturbed the lipid metabolism in liver. Besides, muscle cell development pathway has also been enriched with up-regulated proteins, which is consistent with the functional role of T1R3 as the downstream of muscle regulatory factors [37]. As the amino acid receptor which sense glutamate mostly, it’s predictable that the ablation of T1R1 would affect Glutamatergic synapse pathway [5]. The functional expression of T1R1/T1R3 in mouse neutrophils also showed its potential roles in immune signaling pathway [38].

Combined with the verification results of proteomics analysis by PRM, we found that CYP7B1 and IGFBP2 act as key regulators of T1R1 regulating liver lipid metabolism (Fig. 5A-C). The process in which the cholesterol is conversed into bile acids and excreted from the body is one of the important ways of cholesterol metabolism in mammals, of which oxysterol 7 α-hydroxylase (CYP7B1) acts as a key regulator [39]. As a secreted protein in the liver, IGFBP2 is believed to increase insulin sensitivity and reduce adipogenesis [40, 41]. Therefore, the increase in CYP7B1 and IGFBP2 explained the improvement of liver lipid metabolism.

Since T1R1/T1R3 could also sense branched chain amino acids (BCAAs), the reduction of BCAAs metabolic enzyme activity (BCKDHA and BCKDHB) in T1R1-KO mice (Fig. 5D and E) was found, which decrease in the production of acetyl-CoA, thus leading to the reduction of raw materials of fat de novo synthesis [11, 42]. These changes showed the ablation of T1R1 in the liver of mice may reduce the de novo lipid synthesis process.

In summary, our results showed that the disruption of T1R1 in mice could reduce body lipid accumulation through reduce the lipid synthesis process and increase the lipid metabolism. In the liver, the loss of T1R1 causes the increase of CYP7B1 and IGFBP2 to enhance lipid metabolism, and causes the reduction of BCAA metabolic enzymes (BCKDHA and BCKDHB) to reduce the lipid synthesis process. Our study gives insights into the effect of amino acid sensing on the body's metabolism, and provides a supplement for T1Rs as a potential drug target for the treatment of type 2 diabetes.

Conclusion

In conclusion, we revealed that the disruption of T1R1 in mice could reduce body lipid accumulation through reduce the lipid synthesis process and increase the lipid metabolism. Besides, the decrease in BCKDHA and BCKDHB reduced the de novo lipogenesis, the increase of CYP7B1 and IGFBP2 improved lipid metabolism in T1R1-KO mice.

Abbreviations

TG: triglycerides; TC: total cholesterol; NFALD: non-alcoholic fatty liver disease; GPCR: G protein-coupled receptor; GTT: Glucose tolerance test; ITT: insulin sensitivity test; iWAT: inguinal white adipose tissue; gWAT: gonadal white adipose tissue; BAT: brown adipose tissue; RT-qPCR: Real-time quantitative PCR; LDL: Low-density lipoprotein; LC-MS: liquid chromatography −tandem mass spectrometry; PCA: Principal component analysis; KEGG: Kyoto Encyclopedia of Genes and Genomes; CYP7B1: cholesterol metabolism, oxysterol 7 α-hydroxylase; IGFBP2: Insulin-like growth factor binding protein 2;  BCKDHA: Branched Chain Keto Acid Dehydrogenase E1 Subunit Alpha; BCKDHB:  Branched Chain Keto Acid Dehydrogenase E1 Subunit Beta; AACSA: cetoacetyl-CoA synthetase; ME1: Malic Enzyme-1; GO: Gene ontology .

Declarations

Acknowledgements

We would like to thank Ms. Fan Lin for correcting the language of this article, and your patient modification has made the content of the article enjoyable.

Authors' contributions

Taiyong Yu is the leader of the project, conceived and designed the experiment. Lu Ma and Xuekai Tian performed experiments, analyzed data and wrote the manuscript. Fengxue Xi performed the genotyping. Yulin He, Dong Li, Jingchun Sun, Tiantian Yuan, Yaxin Wang contributed to the manuscript preparation. Fan Lin contributed to the language of this article. Chunlei Zhang supported the T1R1-KO mice. Gongshe Yang contributed to the revisions. All authors reviewed and approved the final manuscript.

Funding 

This study was supported by Key R&D Program of Shaanxi (2017ZDXM-NY-077,

2018ZDXM-NY-02-05) and National Key R&D Program of China (2017YFD0502003).

Availability of data and materials

The data produced or analyzed during the current study are available from the corresponding author by reasonable request.

Ethics approval and consent to participate

The animal protocol was approved by the Animal Care and Use Committee of the Feed Research Institute of Northwest A&F university.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no conflict of interest.

Authors' information (optional)

1Key Laboratory of Animal Gennetics, Breeding and Reproducation of ShaanxiProvince, Laboratory of Animal Fat Deposition & Muscle Development, College of Animal Science and Technology, Northwest A&F University, Yangling Shaanxi 712100, China

Associated content

The Supporting Information is available free of charge at

Table S1: Primers used for RT-qPCR analysis.

References

  1. Gagging, M., F. Carli, C. Rosso, E. Buzzigoli, M. Marietti, V. Della Latta, D. Ciociaro, M. L. Abate, R. Gambino, M. Cassader, E. Bugianesi, and A. Gastaldelli. 2018. Altered amino acid concentrations in NAFLD: Impact of obesity and insulin resistance. Hepatology (Baltimore, Md.) 67(1):145-158. https:// doi.org/10.1002/hep.29465.
  2. Alves, A., A. Bassot, A. L. Bulteau, L. Pirola, and B. Morio. 2019. Glycine Metabolism and Its Alterations in Obesity and Metabolic Diseases. Nutrients 11(6). https:// doi.org/10.3390/nu11061356.
  3. Rose, A. J. 2019. Amino Acid Nutrition and Metabolism in Health and Disease. Nutrients 11(11). https://doi.org/ 10.3390/nu11112623.
  4. Cai J, Wang D, Zhao FQ, Liang S, Liu J. 2020. AMPK-mTOR pathway is involved in glucose-modulated amino acid sensing and utilization in the mammary glands of lactating goats. J Anim Sci Biotechnol. 11:32. https://doi.org/10.1186/s40104-020-0434-6.
  5. Smith, K. R., and A. C. Spector. 2014. The importance of the presence of a 5'-ribonucleotide and the contribution of the T1R1 + T1R3 heterodimer and an additional low-affinity receptor in the taste detection of L-glutamate as assessed psychophysically. The Journal of neuroscience : the official journal of the Society for Neuroscience 34(39):13234-13245. https://doi.org/10.1523/jneurosci.0417-14.2014.
  6. Blonde, G. D., and A. C. Spector. 2017. An Examination of the Role of L-Glutamate and Inosine 5'-Monophosphate in Hedonic Taste-Guided Behavior by Mice Lacking the T1R1 + T1R3 Receptor. Chemical senses 42(5):393-404. https://doi.org/10.1093/chemse/bjx015.
  7. Calvo, S. S., and J. M. Egan. 2015. The endocrinology of taste receptors. Nature reviews. Endocrinology 11(4):213-227. https://doi.org/10.1038/nrendo.2015.7.
  8. Riera, C. E., A. J. T. i. E. Dillin, and Metabolism. 2016. Emerging Role of Sensory Perception in Aging and Metabolism.294-303.https://doi.org/10.1016/j.tem.2016.03.007.
  9. Wauson, E. M., M. L. Guerra, J. Dyachok, K. McGlynn, J. Giles, E. M. Ross, and M. H. Cobb. 2015. Differential Regulation of ERK1/2 and mTORC1 Through T1R1/T1R3 in MIN6 Cells. Molecular endocrinology (Baltimore, Md.) 29(8):1114-1122. https://doi.org/10.1210/me.2014-1181.
  10. Daly, K., M. Al-Rammahi, A. Moran, M. Marcello, Y. Ninomiya, and S. P. Shirazi-Beechey. 2013. Sensing of amino acids by the gut-expressed taste receptor T1R1-T1R3 stimulates CCK secretion. American journal of physiology. Gastrointestinal and liver physiology 304(3):G271-282. https://doi.org/10.1152/ajpgi.00074.2012.
  11. Tian, M., J. Heng, H. Song, Y. Zhang, F. Chen, W. Guan, and S. Zhang. 2019. Branched chain amino acids stimulate gut satiety hormone cholecystokinin secretion through activation of the umami taste receptor T1R1/T1R3 using an in vitro porcine jejunum model. Food & function 10(6):3356-3367. https://doi.org/10.1039/c9fo00228f.
  12. Wauson, E. M., M. L. Guerra, J. Dyachok, K. McGlynn, J. Giles, E. M. Ross, and M. H. Cobb. 2015. Differential Regulation of ERK1/2 and mTORC1 Through T1R1/T1R3 in MIN6 Cells. Molecular endocrinology (Baltimore, Md.) 29(8):1114-1122. https://doi.org/10.1210/me.2014-1181.
  13. Wauson, E. M., E. Zaganjor, A. Y. Lee, M. L. Guerra, A. B. Ghosh, A. L. Bookout, C. P. Chambers, A. Jivan, K. McGlynn, M. R. Hutchison, R. J. Deberardinis, and M. H. Cobb. 2012. The G protein-coupled taste receptor T1R1/T1R3 regulates mTORC1 and autophagy. Molecular cell 47(6):851-862. https://doi.org/10.1016/j.molcel.2012.08.001.
  14. Zheng, L., W. Zhang, Y. Zhou, F. Li, H. Wei, and J. Peng. 2016. Recent Advances in Understanding Amino Acid Sensing Mechanisms that Regulate mTORC1. International journal of molecular sciences 17(10).https://doi.org/10.3390/ijms17101636.
  15. Han, J., and Y. Wang. 2018. mTORC1 signaling in hepatic lipid metabolism. Protein & cell 9(2):145-151. https://doi.org/10.1007/s13238-017-0409-3.
  16. Zhao, G. Q., Y. Zhang, M. A. Hoon, J. Chandrashekar, I. Erlenbach, N. J. Ryba, and C. S. Zuker. 2003. The receptors for mammalian sweet and umami taste. Cell 115(3):255-266. https://doi.org/10.1016/s0092-8674(03)00844-4.
  17. Simon, B. R., B. S. Learman, S. D. Parlee, E. L. Scheller, H. Mori, W. P. Cawthorn, X. Ning, V. Krishnan, Y. L. Ma, B. Tyrberg, and O. A. MacDougald. 2014. Sweet taste receptor deficient mice have decreased adiposity and increased bone mass. PloS one 9(1):e86454. https://doi.org/10.1371/journal.pone.0086454.
  18. Masubuchi, Y., Y. Nakagawa, J. Ma, T. Sasaki, T. Kitamura, Y. Yamamoto, H. Kurose, I. Kojima, and H. Shibata. 2013. A novel regulatory function of sweet taste-sensing receptor in adipogenic differentiation of 3T3-L1 cells. PloS one 8(1):e54500. https:// doi.org/10.1371/journal.pone.0054500.
  19. Masubuchi, Y., Y. Nakagawa, J. Medina, M. Nagasawa, I. Kojima, M. M. Rasenick, T. Inagaki, and H. Shibata. 2017. T1R3 homomeric sweet taste receptor regulates adipogenesis through Gαs-mediated microtubules disassembly and Rho activation in 3T3-L1 cells. PloS one 12(5):e0176841. https://doi.org/10.1371/journal.pone.0176841.
  20. Smith, K. R., T. Hussain, E. Karimian Azari, J. L. Steiner, J. E. Ayala, R. E. Pratley, and G. A. Kyriazis. 2016. Disruption of the sugar-sensing receptor T1R2 attenuates metabolic derangements associated with diet-induced obesity. American journal of physiology. Endocrinology and metabolism 310(8):E688-e698. https://doi.org/10.1152/ajpendo.00484.2015.
  21. Medina, A., Y. Nakagawa, J. Ma, L. Li, K. Hamano, T. Akimoto, Y. Ninomiya, and I. Kojima. 2014. Expression of the glucose-sensing receptor T1R3 in pancreatic islet: changes in the expression levels in various nutritional and metabolic states. Endocrine journal 61(8):797-805. https://doi.org/10.1507/endocrj.ej14-0221.
  22. Nakagawa, Y., Y. Ohtsu, M. Nagasawa, H. Shibata, and I. Kojima. 2014. Glucose promotes its own metabolism by acting on the cell-surface glucose-sensing receptor T1R3. Endocrine journal 61(2):119-131. https://doi.org/10.1507/endocrj.ej13-0431.
  23. Liu, J., Y. Wang, D. Li, Y. Wang, M. Li, C. Chen, X. Fang, H. Chen, and C. Zhang. 2017. Milk protein synthesis is regulated by T1R1/T1R3, a G protein-coupled taste receptor, through the mTOR pathway in the mouse mammary gland. Molecular nutrition & food research 61(9). https://doi.org/10.1002/mnfr.201601017.
  24. Husted, A. S., M. Trauelsen, O. Rudenko, S. A. Hjorth, and T. W. Schwartz. 2017. GPCR-Mediated Signaling of Metabolites. Cell metabolism 25(4):777-796. https://doi.org/10.1016/j.cmet.2017.03.008.
  25. Barella, L. F., S. Jain, T. Kimura, and S. P. Pydi. 2021. Metabolic roles of G protein-coupled receptor signaling in obesity and type 2 diabetes. The FEBS journal 288(8):2622-2644. https://doi.org/10.1111/febs.15800.
  26. Conigrave AD, Hampson DR. 2010. Broad-spectrum amino acid-sensing class C G-protein coupled receptors: molecular mechanisms, physiological significance and options for drug development. Pharmacol Ther 127(3):252-60. https://doi.org/10.1016/j.pharmthera.
  27. Dotson CD, Vigues S, Steinle NI, Munger SD. 2010. T1R and T2R receptors: the modulation of incretin hormones and potential targets for the treatment of type 2 diabetes mellitus. Curr Opin Investig Drugs 11(4):447-54. https://doi.org/10.1016/j.clinthera.2010.04.006.
  28. Murovets, V. O., A. A. Bachmanov, S. V. Travnikov, A. A. Churikova, and V. A. Zolotarev. 2014. The Involvement of the T1R3 Receptor Protein in the Control of Glucose Metabolism in Mice at Different Levels of Glycemia. Journal of evolutionary biochemistry and physiology 50(4):334-344. https://doi.org/10.1134/s002209301404006.
  29. Henquin, J. C. 2012. Do pancreatic β cells "taste" nutrients to secrete insulin? Science signaling 5(239):pe36. https://doi.org/10.1126/scisignal.2003325.
  30. Eaton, M. S., N. Weinstein, J. B. Newby, M. M. Plattes, H. E. Foster, J. W. Arthur, T. D. Ward, S. R. Shively, R. Shor, J. Nathan, H. M. Davis, L. I. Plotkin, E. M. Wauson, B. J. Dewar, A. Broege, and J. W. Lowery. 2018. Loss of the nutrient sensor TAS1R3 leads to reduced bone resorption. Journal of physiology and biochemistry 74(1):3-8. https://doi.org/10.1007/s13105-017-0596-7.
  31. Moseti, D., A. Regassa, and W. K. Kim. 2016. Molecular Regulation of Adipogenesis and Potential Anti-Adipogenic Bioactive Molecules. International journal of molecular sciences 17(1). https://doi.org/10.3390/ijms17010124.
  32. Engelking, L. J., M. J. Cantoria, Y. Xu, and G. Liang. 2018. Developmental and extrahepatic physiological functions of SREBP pathway genes in mice. Seminars in cell & developmental biology 81:98-109. https://doi.org/10.1016/j.semcdb.2017.07.011.
  33. Tsoli, M., M. M. Swarbrick, and G. R. Robertson. 2016. Lipolytic and thermogenic depletion of adipose tissue in cancer cachexia. Seminars in cell & developmental biology 54:68-81. https://doi.org/10.1016/j.semcdb.2015.10.039.
  34. Chiu, Y. J., H. H. Tu, M. L. Kung, H. J. Wu, and Y. W. Chen. 2021. Fluoxetine ameliorates high-fat diet-induced metabolic abnormalities partially via reduced adipose triglyceride lipase-mediated adipocyte lipolysis. Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie 141:111848. https://doi.org/10.1016/j.biopha.2021.111848.
  35. Kristensen, K. K., K. Z. Leth-Espensen, A. Kumari, A. L. Grønnemose, A. M. Lund-Winther, S. G. Young, and M. Ploug. 2021. GPIHBP1 and ANGPTL4 Utilize Protein Disorder to Orchestrate Order in Plasma Triglyceride Metabolism and Regulate Compartmentalization of LPL Activity. Frontiers in cell and developmental biology 9:702508. https://doi.org/10.3389/fcell.2021.702508.
  36. Zheng A, Zhang A, Chen Z, Pirzado SA, Chang W, Cai H, Bryden WL, Liu G. 2021. Molecular mechanisms of growth depression in broiler chickens (Gallus Gallus domesticus) mediated by immune stress: a hepatic proteome study. J Anim Sci Biotechnol. 12(1):90. https://doi.org/10.1186/s40104-021-00591-1.
  37. Kokabu, S., J. W. Lowery, T. Toyono, Y. Seta, S. Hitomi, T. Sato, Y. Enoki, M. Okubo, Y. Fukushima, and T. Yoda. 2015. Muscle regulatory factors regulate T1R3 taste receptor expression. Biochemical and biophysical research communications 468(4):568-573. https://doi.org/10.1016/j.bbrc.2015.10.142.
  38. Lee, N., Y. S. Jung, H. Y. Lee, N. Kang, Y. J. Park, J. S. Hwang, Y. Y. Bahk, J. Koo, and Y. S. Bae. 2014. Mouse neutrophils express functional umami taste receptor T1R1/T1R3. BMB reports 47(11):649-654. https://doi.org/10.5483/bmbrep.2014.47.11.185.
  39. Pandak, W. M., and G. Kakiyama. 2019. The acidic pathway of bile acid synthesis: Not just an alternative pathway. Liver research 3(2):88-98. https://doi.org/10.1016/j.livres.2019.05.001.
  40. Fahlbusch, P., B. Knebel, T. Hörbelt, D. M. Barbosa, A. Nikolic, S. Jacob, H. Al-Hasani, F. Van de Velde, Y. Van Nieuwenhove, D. Müller-Wieland, B. Lapauw, D. M. Ouwens, and J. Kotzka. 2020. Physiological Disturbance in Fatty Liver Energy Metabolism Converges on IGFBP2 Abundance and Regulation in Mice and Men. International journal of molecular sciences 21(11). https://doi.org/10.3390/ijms21114144.
  41. Yang, J., W. Zhou, Y. Wu, L. Xu, Y. Wang, Z. Xu, and Y. Yang. 2020. Circulating IGFBP-2 levels are inversely associated with the incidence of nonalcoholic fatty liver disease: A cohort study. The Journal of international medical research 48(8):300060520935219. https://doi.org/10.1177/0300060520935219.
  42. Green, C. R., M. Wallace, A. S. Divakaruni, S. A. Phillips, A. N. Murphy, T. P. Ciaraldi, and C. M. Metallo. 2016. Branched-chain amino acid catabolism fuels adipocyte differentiation and lipogenesis. Nature chemical biology 12(1):15-21. https://doi.org/10.1038/nchembio.1961.