MOLECULAR EVOLUTION OF GPCRS: GLP1/GLP1 receptors

Hwang, Jong-Ik,Yun, Seongsik,Moon, Mi Jin,Park, Cho Rong,Seong, Jae Young Journal of Endocrinology (Ltd. by Guarantee) 2014 Journal of molecular endocrinology Vol.52 No.3

<P>Glucagon-like peptide 1 (GLP1) is an intestinal incretin that regulates glucose homeostasis through stimulation of insulin secretion from pancreatic β-cells and inhibits appetite by acting on the brain. Thus, it is a promising therapeutic agent for the treatment of type 2 diabetes mellitus and obesity. Studies using synteny and reconstructed ancestral chromosomes suggest that families for GLP1 and its receptor (GLP1R) have emerged through two rounds (2R) of whole genome duplication and local gene duplications before and after 2R. Exon duplications have also contributed to the expansion of the peptide family members. Specific changes in the amino acid sequence following exon/gene/genome duplications have established distinct yet related peptide and receptor families. These specific changes also confer selective interactions between GLP1 and GLP1R. In this review, we present a possible macro (genome level)- and micro (gene/exon level)-evolution mechanisms of GLP1 and GLP1R, which allows them to acquire selective interactions between this ligand–receptor pair. This information may provide critical insight for the development of potent therapeutic agents targeting GLP1R.</P>

Angiotensin II stimulates the synthesis of vascular endothelial growth factor through the p38 mitogen activated protein kinase pathway in cultured mouse podocytes.

Kang, Young Sun,Park, Yun Gyu,Kim, Bo Kyung,Han, Sang Youb,Jee, Yi Hwa,Han, Kum Hyun,Lee, Mi Hwa,Song, Hye Kyoung,Cha, Dae Ryong,Kang, Shin Wook,Han, Dae Suk Journal of Endocrinology (Ltd. by Guarantee) 2006 Journal of molecular endocrinology Vol.36 No.2

<P>Angiotensin II (Ang-II) and vascular endothelial growth factor (VEGF) have an important role in the pathogenesis of diabetic nephropathy, but the signaling cascade of VEGF regulation in response to Ang-II in podocytes is largely unknown. In these experiments, we looked at the effect of Ang-II on the production of VEGF, and investigated whether VEGF production depends on the p38 mitogen activated protein kinase (MAPK) pathway in cultured mouse podocytes. Incubation of podocytes with Ang-II induced a rapid increase in VEGF mRNA expression and protein synthesis as well as its transcriptional activity in an Ang-II dose-dependent manner. To further define the role of angiotensin type 1 (AT1) and type 2 (AT2) receptors involved in Ang-II-mediated VEGF synthesis, the effects of selective AT1 and AT2 receptor antagonists were evaluated. Prior treatment with losartan significantly inhibited VEGF mRNA and protein synthesis induced by Ang-II, which suggests that the AT1 receptor is involved in Ang-II-mediated VEGF synthesis. Furthermore, stimulation of the cells with Ang-II increased both phosphorylation of p38 MAPK and MAP kinase kinase 3/6 (MKK3/6). Additionally, Ang-II enhanced the DNA binding activity to cAMP response element binding protein (CREB) and phosphorylation of CREB. In addition, to investigate the role of p38 MAPK in Ang-II-induced VEGF synthesis, podocytes were pretreated with or without the p38 MAPK inhibitor, SB203580 for 24 h to observe whether Ang-II-mediated VEGF synthesis was inhibited by blocking p38 MAPK. The addition of SB203580 led to a marked inhibition of the increased VEGF mRNA and protein production induced by Ang-II in a dose-dependent manner. Taken together, these results suggest that Ang-II stimulates the synthesis of VEGF in podocytes and the production of VEGF induced by Ang-II is mediated, in part, through the activation of the p38 MAPK pathway.</P>

KSR1 is coordinately regulated with Notch signaling and oxidative phosphorylation in thyroid cancer

Lee, Jandee,Seol, Mi-Youn,Jeong, Seonhyang,Kwon, Hyeong Ju,Lee, Cho Rok,Ku, Cheol Ryong,Kang, Sang-Wook,Jeong, Jong Ju,Shin, Dong Yeob,Nam, Kee-Hyun,Lee, Eun Jig,Chung, Woong Youn,Jo, Young Suk Journal of Endocrinology (Ltd. by Guarantee) 2015 Journal of molecular endocrinology Vol.54 No.2

<P>Kinase suppressor of RAS1 (KSR1) is a scaffold protein implicated in RAS-mediated RAF activation. However, the molecular function of KSR in papillary thyroid cancer (PTC) is unknown. Thus, this study aimed to characterize the role of KSR1 in patients with PTC. qRT-PCR and immunohistochemistry (IHC) revealed inter-tumor heterogeneities in the expression of KSR1 in PTC tissues. Interestingly, BRAFV600E-positive PTC showed higher <I>KSR1</I> mRNA expression than BRAFV600E-negative PTC (<I>P</I><0.001). Gene Set Enrichment Analysis (GSEA) using public repositories showed that high KSR1 expression coordinately upregulated Notch signaling (nominal <I>P</I>=0.019, false discovery rate (FDR) <I>q</I>-value=0.165); this finding was supported by GeneNetwork analysis, indicating that <I>KSR1</I> expression is positively correlated with <I>NOTCH1</I> expression (<I>ρ</I>=0.677, <I>P</I>=6.15×10<SUP>−9</SUP>). siRNA against KSR1 (siKSR1) significantly decreased ERK phosphorylation induced by BRAFV600E, resulting in reduced expression of <I>NOTCH1</I> and <I>HES1</I>, targets of Notch signaling. GSEA revealed that high KSR1 expression was also associated with downregulation of genes related to oxidative phosphorylation (OxPhos). Consistent with this, electron microscopy showed that PTCs with high KSR1 expression exhibited structural defects of the mitochondrial cristae. Furthermore, siKSR1-transfected BCPAP and 8505C cells generated fewer colonies in colony-forming assays. In addition, GSEA showed that high expression of KSR2 and connector enhancer of KSR1 (CNKSR1) also coordinately upregulated Notch signaling (KSR2: nominal <I>P</I>=0.0097, FDR <I>q</I>-value=0.154 and CNKSR1: nominal <I>P</I><0.0001, FDR <I>q</I>-value=0.00554), and high CNKSR2 was associated with downregulation of the OxPhos gene set (nominal <I>P</I><0.0001, FDR <I>q</I>-value <0.0001). In conclusion, KSR1 is coordinately regulated with Notch signaling and OxPhos in PTC, because its scaffold function might be required to sustain the proliferative signaling and metabolic remodeling associated with this type of cancer.</P>

Visfatin, a novel adipokine, stimulates glucose uptake through the Ca²⁺-dependent AMPK–p38 MAPK pathway in C2C12 skeletal muscle cells

Lee, Jung Ok,Kim, Nami,Lee, Hye Jeong,Lee, Yong Woo,Kim, Joong Kwan,Kim, Hyung Ip,Lee, Soo Kyung,Kim, Su Jin,Park, Sun Hwa,Kim, Hyeon Soo Journal of Endocrinology (Ltd. by Guarantee) 2015 Journal of molecular endocrinology Vol.54 No.3

<P>Visfatin is a novel adipocytokine produced by visceral fat. In the present study, visfatin increased AMP-activated protein kinase (AMPK) phosphorylation in mouse C2C12 skeletal muscle cells. It also increased phosphorylation of the insulin receptor, whose knockdown blocked visfatin-induced AMPK phosphorylation and glucose uptake. Visfatin stimulated glucose uptake in differentiated skeletal muscle cells. However, inhibition of AMPK alpha 2 with an inhibitor or with knockdown of AMPK alpha 2 using siRNA blocked visfatin-induced glucose uptake, which indicates that visfatin stimulates glucose uptake through the AMPK alpha 2 pathway. Visfatin increased the intracellular Ca2+ concentration. STO-609, a calmodulin-dependent protein kinase kinase inhibitor, blocked visfatin-induced AMPK phosphorylation and glucose uptake. Visfatin-mediated activation of p38 MAPK was AMPK alpha 2-dependent. Furthermore, both inhibition and knockdown of p38 MAPK blocked visfatin-induced glucose uptake. Visfatin increased glucose transporter type 4 (GLUT4) mRNA and protein levels. In addition, visfatin stimulated the translocation of GLUT4 to the plasma membrane, and this effect was suppressed by AMPK alpha 2 inhibition. The present results indicate that visfatin plays an important role in glucose metabolism via the Ca2+-mediated AMPK-p38 MAPK pathway.</P>

Molecular cloning and functional characterization of a type-I neurotensin receptor (NTR) and a novel NTR from the bullfrog brain.

Li, J H,Sicard, F,Salam, M A,Baek, M,LePrince, J,Vaudry, H,Kim, K,Kwon, H B,Seong, J Y Journal of Endocrinology (Ltd. by Guarantee) 2005 Journal of molecular endocrinology Vol.34 No.3

<P>Neurotensin (NT) is a tridecapeptide that functions as a neurotransmitter and neuromodulator in the nervous system. To date, three different types of NT receptor (NTR), NTR1, NTR2 and NTR3, have been identified only in mammalian species. In the present study we isolated the cDNAs for an NTR1 and a novel NTR in the bullfrog brain, designated bfNTR1 and bfNTR4 respectively. bfNTR1 and bfNTR4 encode 422- and 399-amino acid residue proteins respectively. bfNTR1 has a 64% amino acid identity with mammalian NTR1, and 34-37% identity with mammalian NTR2. bfNTR4 exhibits 43% and 45-47% identity with mammalian NTR1 and NTR2 respectively. Both receptors are mainly expressed in the brain and pituitary. bfNTR1 triggers both CRE-luc, a protein kinase A (PKA)-specific reporter, and c-fos-luc, a PKC-specific reporter, activities, indicating that bfNTR1 can activate PKA- and PKC-linked signaling pathways. However, bfNTR4 appears to be preferentially coupled to the PKA-linked pathway as it induces a higher CRE-luc activity than c-fos-luc activity. bfNTRs exhibit different pharmacological properties as compared with mammalian NTRs. Mammalian NTR1 but not NTR2 responds to NT, whereas both bfNTR1 and bfNTR4 show a high sensitivity to NT. SR 48692 and SR 142948A, antagonists for mammalian NTR1 but agonists for mammalian NTR2, function as antagonists for both bfNTR1 and bfNTR4. In conclusion, this report provides the first molecular, pharmacological and functional characterization of two NTRs in a non-mammalian vertebrate. These data should help to elucidate the phylogenetic history of the G protein-coupled NTRs in the vertebrate lineage as well as the structural features that determine their pharmacological properties.</P>

Biological activities of recombinant Manchurian trout FSH and LH: their receptor specificity, steroidogenic and vitellogenic potencies.

Ko, Hyeyeon,Park, WooDong,Kim, Dae-Jung,Kobayashi, Makito,Sohn, Young Chang Journal of Endocrinology (Ltd. by Guarantee) 2007 Journal of molecular endocrinology Vol.38 No.1

<P>Gonadotropins (GTHs), FSH and LH, play central roles in vertebrate reproduction. Here, we report the production of biologically-active recombinant FSH (r-mtFSH) and LH (r-mtLH) of an endangered salmon species, Manchurian trout (Brachymystax lenok), by baculovirus in silkworm (Bombyx mori) larvae. The biological activities of the recombinant hormones were analyzed using COS-7 cell line transiently expressing either amago salmon FSH or LH receptor. The steroidogenic potency of the r-mtFSH and r-mtLH was examined by a culture system using rainbow trout follicles in vitro. In vivo, bioactivity was assessed by measuring ovarian weight, oocyte diameter, and plasma steroid hormone levels in female rainbow trout. Moreover, inducing potency of milt production were examined in vivo using goldfish. Our results demonstrated that the r-mtFSH and r-mtLH were successfully produced in the baculovirus-silkworm system and recognized by their cognate receptors specifically in vitro. The production of estradiol-17beta (E2) and testosterone (T) was stimulated by the r-mtFSH and r-mtLH respectively, from the full-grown follicles of rainbow trout, whereas both E2 and T were increased by relatively higher doses of the recombinant hormones from the follicles of the maturing stage. In in vivo assay, injection of the r-mtFSH but not r-mtLH increased ovarian weight, oocyte diameter, and plasma E2 levels in immature rainbow trout. Injection of both r-mtFSH and r-mtLH induced milt production in male goldfish. In conclusion, the present study strongly suggests that the r-mtFSH and r-mtLH have distinct biological properties, such as a specific responsiveness for the cognate receptor, steroidogenic, and vitellogenic activities for ovarian follicles in salmonids. These recombinant FSH and LH may be applied for future studies on the gonadal development and maturation in fishes as well as the endangered salmon species.</P>

Analysis of gene expression profiles in insulin-sensitive tissues from pre-diabetic and diabetic Zucker diabetic fatty rats.

Suh, Young Ho,Kim, Younyoung,Bang, Jeong Hyun,Choi, Kyoung Suk,Lee, June Woo,Kim, Won-Ho,Oh, Tae Jeong,An, Sungwhan,Jung, Myeong Ho Journal of Endocrinology (Ltd. by Guarantee) 2005 Journal of molecular endocrinology Vol.34 No.2

<P>Insulin resistance occurs early in the disease process, preceding the development of type 2 diabetes. Therefore, the identification of molecules that contribute to insulin resistance and leading up to type 2 diabetes is important to elucidate the molecular pathogenesis of the disease. To this end, we characterized gene expression profiles from insulin-sensitive tissues, including adipose tissue, skeletal muscle, and liver tissue of Zucker diabetic fatty (ZDF) rats, a well characterized type 2 diabetes animal model. Gene expression profiles from ZDF rats at 6 weeks (pre-diabetes), 12 weeks (diabetes), and 20 weeks (late-stage diabetes) were compared with age- and sex-matched Zucker lean control (ZLC) rats using 5000 cDNA chips. Differentially regulated genes demonstrating > 1.3-fold change at age were identified and categorized through hierarchical clustering analysis. Our results showed that while expression of lipolytic genes was elevated in adipose tissue of diabetic ZDF rats at 12 weeks of age, expression of lipogenic genes was decreased in liver but increased in skeletal muscle of 12 week old diabetic ZDF rats. These results suggest that impairment of hepatic lipogenesis accompanied with the reduced lipogenesis of adipose tissue may contribute to development of diabetes in ZDF rats by increasing lipogenesis in skeletal muscle. Moreover, expression of antioxidant defense genes was decreased in the liver of 12-week old diabetic ZDF rats as well as in the adipose tissue of ZDF rats both at 6 and 12 weeks of age. Cytochrome P450 (CYP) genes were also significantly reduced in 12 week old diabetic liver of ZDF rats. Genes involved in glucose utilization were downregulated in skeletal muscle of diabetic ZDF rats, and the hepatic gluconeogenic gene was upregulated in diabetic ZDF rats. Genes commonly expressed in all three tissue types were also observed. These profilings might provide better fundamental understanding of insulin resistance and development of type 2 diabetes.</P>

Ghrelin gene products rescue cultured adult rat hippocampal neural stem cells from high glucose insult

Kim, Sehee,Kim, Chanyang,Park, Seungjoon Journal of Endocrinology (Ltd. by Guarantee) 2016 Journal of molecular endocrinology Vol.57 No.3

<P>Adult hippocampal neurogenesis is decreased in type 2 diabetes, and this impairment appears to be important in cognitive dysfunction. Previous studies suggest that ghrelin gene products (acylated ghrelin (AG), unacylated ghrelin (UAG) and obestatin (OB)) promote neurogenesis. Therefore, we hypothesize that ghrelin gene products may reduce the harmful effects of high glucose (HG) on hippocampal neural stem cells (NSCs). The aim of this study was to investigate the role of these peptides on the survival of cultured hippocampal NSCs exposed to HG insult. Treatment of hippocampal NSCs with AG, UAG or OB inhibited HG-induced cell death and apoptosis. Exposure of cells to the growth hormone secretagogue receptor 1a antagonist abolished the protective effects of AG against HG toxicity, whereas those of UAG or OB were preserved. All three peptides attenuated HG-induced decrease in BrdU-labeled and phosphohistone-H3-labeled cells. We also investigated the effects of ghrelin gene products on the regulation of apoptosis at the mitochondrial level. AG, UAG or OB rescued hippocampal NSCs from HG insult by inhibiting intracellular and mitochondrial reactive oxygen species generation and stabilizing mitochondrial transmembrane potential. In addition, cells treated with ghrelin gene products showed an increased Bcl-2 and decreased Bax levels, thereby increasing the Bcl-2/Bax ratio, inhibiting cytochrome c release and preventing caspase-3 activation. Finally, AG-, UAG- or OB-mediated protection was dependent on the activities of adenosine monophosphate-activated protein kinase/uncoupling protein 2 pathway. Our data indicate that ghrelin gene products may act as survival factors that preserve mitochondrial function and inhibit oxidative stress-induced apoptosis.</P>

LECT2 promotes inflammation and insulin resistance in adipocytes via P38 pathways

Jung, Tae Woo,Chung, Yoon Hee,Kim, Hyoung-Chun,Abd El-Aty, A M,Jeong, Ji Hoon Journal of Endocrinology (Ltd. by Guarantee) 2018 Journal of molecular endocrinology Vol.61 No.1

<P>Leukocyte cell-derived chemotaxin 2 (LECT2) is a recently identified novel hepatokine that causes insulin resistance in skeletal muscle by activating c-Jun N-terminal kinase (JNK), thereby driving atherosclerotic inflammation. However, the role of LECT2 in inflammation and insulin resistance in adipocytes has not been investigated. In this study, we report that LECT2 treatment of differentiated 3T3-L1 cells stimulates P38 phosphorylation in a dose-dependent manner. LECT2 also enhanced inflammation markers such as I kappa B phosphorylation, nuclear factor kappa beta (NF-kappa B) phosphorylation and IL-6 expression. Moreover, LECT2 treatment impaired insulin signaling in differentiated 3T3-L1 cells, as evidenced by the decreased levels of insulin receptor substrate (IRS-1) and Akt phosphorylation and reduced insulin-stimulated glucose uptake. Furthermore, LECT2 augmented lipid accumulation during 3T3-L1 cell differentiation by activating SREBP1c-mediated signaling. All these effects were significantly abrogated by siRNA-mediated silencing of P38, CD209 expression or a JNK inhibitor. Our findings suggest that LECT2 stimulates inflammation and insulin resistance in adipocytes via activation of a CD209/P38-dependent pathway. Thus, these results suggest effective therapeutic targets for treating inflammation-mediated insulin resistance.</P>

Recent insight into the correlation of SREBP-mediated lipid metabolism and innate immune response

Park, Hyeon Young,Kang, Hye Suk,Im, Seung-Soon Journal of Endocrinology (Ltd. by Guarantee) 2018 Journal of molecular endocrinology Vol.61 No.3

<P>Fatty acids are essential nutrients that contribute to several intracellular functions. Fatty acid synthesis and oxidation are known to be regulated by sterol regulatory element-binding proteins (SREBPs), which play a pivotal role in the regulation of cellular triglyceride synthesis and cholesterol biogenesis. Recent studies point to a multifunctional role of SREBPs in the pathogenesis of metabolic diseases, such as obesity, type II diabetes and cancer as well as in immune responses. Notably, fatty acid metabolic intermediates are involved in energy homeostasis and pathophysiological conditions. In particular, intracellular fatty acid metabolism affects an inflammatory response, thereby influencing metabolic diseases. The objective of this review is to summarize the recent advances in our understanding of the dual role of SREBPs in both lipid metabolism and inflammation-mediated metabolic diseases.</P>