1.Gαi1/3 mediation of Akt-mTOR activation is important for RSPO3-induced angiogenesis.
Gang XU ; Li-Na QI ; Mei-Qing ZHANG ; Xue-Yun LI ; Jin-Long CHAI ; Zhi-Qing ZHANG ; Xia CHEN ; Qian WANG ; Ke-Ran LI ; Cong CAO
Protein & Cell 2023;14(3):217-222
2.Inhibitory G proteins and their receptors: emerging therapeutic targets for obesity and diabetes.
Michelle E KIMPLE ; Joshua C NEUMAN ; Amelia K LINNEMANN ; Patrick J CASEY
Experimental & Molecular Medicine 2014;46(6):e102-
The worldwide prevalence of obesity is steadily increasing, nearly doubling between 1980 and 2008. Obesity is often associated with insulin resistance, a major risk factor for type 2 diabetes mellitus (T2DM): a costly chronic disease and serious public health problem. The underlying cause of T2DM is a failure of the beta cells of the pancreas to continue to produce enough insulin to counteract insulin resistance. Most current T2DM therapeutics do not prevent continued loss of insulin secretion capacity, and those that do have the potential to preserve beta cell mass and function are not effective in all patients. Therefore, developing new methods for preventing and treating obesity and T2DM is very timely and of great significance. There is now considerable literature demonstrating a link between inhibitory guanine nucleotide-binding protein (G protein) and G protein-coupled receptor (GPCR) signaling in insulin-responsive tissues and the pathogenesis of obesity and T2DM. These studies are suggesting new and emerging therapeutic targets for these conditions. In this review, we will discuss inhibitory G proteins and GPCRs that have primary actions in the beta cell and other peripheral sites as therapeutic targets for obesity and T2DM, improving satiety, insulin resistance and/or beta cell biology.
Animals
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Diabetes Mellitus, Type 2/drug therapy/*metabolism
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GTP-Binding Protein alpha Subunits/genetics/*metabolism
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Humans
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Insulin-Secreting Cells/metabolism
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Obesity/drug therapy/*metabolism
;
Receptor, Melatonin, MT2/genetics/*metabolism
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Receptors, Adrenergic, alpha-1/genetics/*metabolism
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Receptors, Prostaglandin/genetics/*metabolism
3.YME1L overexpression exerts pro-tumorigenic activity in glioma by promoting Gαi1 expression and Akt activation.
Fang LIU ; Gang CHEN ; Li-Na ZHOU ; Yin WANG ; Zhi-Qing ZHANG ; Xihu QIN ; Cong CAO
Protein & Cell 2023;14(3):223-229
Humans
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Brain Neoplasms/metabolism*
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Cell Line, Tumor
;
Cell Proliferation
;
Gene Expression Regulation, Neoplastic
;
Glioma/metabolism*
;
Proto-Oncogene Proteins c-akt/metabolism*
;
Signal Transduction
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ATPases Associated with Diverse Cellular Activities/metabolism*
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Mitochondrial Proteins/metabolism*
;
GTP-Binding Protein alpha Subunits/metabolism*
4.Efficient fusion expression of G13 domain derived from granulysin in Escherichia coli.
Xiaoqiang LIU ; Xiangdong ZHA ; Yazhong XIAO ; Jinhuan YANG ; Nengshu LI
Chinese Journal of Biotechnology 2009;25(2):235-241
The G13 domain derived from granulysin shows high antimicrobial activities against Gram-positive and Gram-negative bacteria but does not lyse Jurkat cells or liposomes. To explore a new approach for high expression of the G13 domain, we fused the sequence encoding G13 to thioredoxin (Trx) gene to construct the recombinant expression vector (pThioHisA-G13). A cyanogen bromide (CNBr) cleavage site was introduced between the Trx and G13 to facilitate final release of the recombinant G13. The recombinant expression vector, pThioHisA-G13, was transformed into E. coli BL21 (DE3). Upon induction by IPTG Trx-G13 fusion protein was expressed and took the form of inclusion bodies counting 58% (W/W) of total cellular proteins. The inclusion body was solved by urea (8 mol/L) and then cleaved by CNBr. We purified the recombinant peptide G13 by one-step cation exchange chromatography. Results of agarose diffuse assay analysis indicated that the recombinant G13 exhibited antibacterial activity. The procedure described in this study will provide a reliable and simple method for highly efficient production of some cationic antimicrobial peptides.
Anti-Infective Agents
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metabolism
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Antigens, Differentiation, T-Lymphocyte
;
genetics
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Cyanogen Bromide
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pharmacology
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Escherichia coli
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genetics
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metabolism
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GTP-Binding Protein alpha Subunits, G12-G13
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biosynthesis
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genetics
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Inclusion Bodies
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metabolism
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Protein Structure, Tertiary
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genetics
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Recombinant Fusion Proteins
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biosynthesis
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genetics
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Thioredoxins
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genetics
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Transfection
5.Stimulatory heterotrimeric G protein augments gamma ray-induced apoptosis by up-regulation of Bak expression via CREB and AP-1 in H1299 human lung cancer cells.
Yoon Jung CHOI ; So Young KIM ; Jung Min OH ; Yong Sung JUHNN
Experimental & Molecular Medicine 2009;41(8):592-600
Stimulatory heterotrimeric GTP-binding proteins (Gs protein) stimulate cAMP generation in response to various signals, and modulate various cellular phenomena such as proliferation and apoptosis. This study aimed to investigate the effect of Gs proteins on gamma ray-induced apoptosis of lung cancer cells and its molecular mechanism, as an attempt to develop a new strategy to improve the therapeutic efficacy of gamma radiation. Expression of constitutively active mutant of the alpha subunit of Gs (GalphasQL) augmented gamma ray-induced apoptosis via mitochondrial dependent pathway when assessed by clonogenic assay, FACS analysis of PI stained cells, and western blot analysis of the cytoplasmic translocation of cytochrome C and the cleavage of caspase-3 and ploy(ADP-ribose) polymerase (PARP) in H1299 human lung cancer cells. GalphasQL up-regulated the Bak expression at the levels of protein and mRNA. Treatment with inhibitors of PKA (H89), SP600125 (JNK inhibitor), and a CRE-decoy blocked GalphasQL-stimulated Bak reporter luciferase activity. Expression of GalphasQL increased basal and gamma ray-induced luciferase activity of cAMP response element binding protein (CREB) and AP-1, and the binding of CREB and AP-1 to Bak promoter. Furthermore, prostaglandin E2, a Galphas activating signal, was found to augment gamma ray-induced apoptosis, which was abolished by treatment with a prostanoid receptor antagonist. These results indicate that Galphas augments gamma ray-induced apoptosis by up-regulation of Bak expression via CREB and AP-1 in H1299 lung cancer cells, suggesting that the efficacy of radiotherapy of lung cancer may be improved by modulating Gs signaling pathway.
Apoptosis/*radiation effects
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Cell Line, Tumor
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Cyclic AMP Response Element-Binding Protein/metabolism
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GTP-Binding Protein alpha Subunits, Gs/*metabolism
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*Gamma Rays
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Heterotrimeric GTP-Binding Proteins/metabolism
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Humans
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Lung/*cytology/physiology/radiation effects
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Lung Neoplasms
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Transcription Factor AP-1/metabolism
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*Up-Regulation
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bcl-2 Homologous Antagonist-Killer Protein/*metabolism
6.Simultaneous deletion of floxed genes mediated by CaMKIIalpha-Cre in the brain and in male germ cells: application to conditional and conventional disruption of Goalpha.
Chan Il CHOI ; Sang Phil YOON ; Jung Mi CHOI ; Sung Soo KIM ; Young Don LEE ; Lutz BIRNBAUMER ; Haeyoung SUH-KIM
Experimental & Molecular Medicine 2014;46(5):e93-
The Cre/LoxP system is a well-established approach to spatially and temporally control genetic inactivation. The calcium/calmodulin-dependent protein kinase II alpha subunit (CaMKIIalpha) promoter limits expression to specific regions of the forebrain and thus has been utilized for the brain-specific inactivation of the genes. Here, we show that CaMKIIalpha-Cre can be utilized for simultaneous inactivation of genes in the adult brain and in male germ cells. Double transgenic Rosa26(+/stop-lacZ)::CaMKIIalpha-Cre(+/Cre) mice generated by crossing CaMKIIalpha-Cre(+/Cre) mice with floxed ROSA26 lacZ reporter (Rosa26(+/stop-lacZ)) mice exhibited lacZ expression in the brain and testis. When these mice were mated to wild-type females, about 27% of the offspring were whole body blue by X-gal staining without inheriting the Cre transgene. These results indicate that recombination can occur in the germ cells of male Rosa26(+/stop-lacZ)::CaMKIIalpha-Cre(+/Cre) mice. Similarly, when double transgenic Gnao(+/f)::CaMKIIalpha-Cre(+/Cre) mice carrying a floxed Go-alpha gene (Gnao(f/f)) were backcrossed to wild-type females, approximately 22% of the offspring carried the disrupted allele (Gnao(Delta)) without inheriting the Cre transgene. The Gnao(Delta/Delta) mice closely resembled conventional Go-alpha knockout mice (Gnao(-/-)) with respect to impairment of their behavior. Thus, we conclude that CaMKIIalpha-Cre mice afford recombination for both tissue- and time-controlled inactivation of floxed target genes in the brain and for their permanent disruption. This work also emphasizes that extra caution should be exercised in utilizing CaMKIIalpha-Cre mice as breeding pairs.
Animals
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Brain/*metabolism
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Calcium-Calmodulin-Dependent Protein Kinase Type 2/genetics
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Female
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GTP-Binding Protein alpha Subunits, Gi-Go/*genetics
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*Gene Deletion
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Gene Knockout Techniques/*methods
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Male
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Mice
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RNA, Untranslated/genetics
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Recombination, Genetic
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Spermatozoa/*metabolism
7.An inhibitory compound against the interaction between Galpha(s) and the third intracellular loop region of serotonin receptor subtype 6 (5-HT(6)) disrupts the signaling pathway of 5-HT(6).
Yun Hee CHOI ; Hatan KANG ; Won Kyu LEE ; Taehyun KIM ; Hyewhon RHIM ; Yeon Gyu YU
Experimental & Molecular Medicine 2007;39(3):335-342
Serotonin receptor subtype 6 (5-HT(6)) is a neurotransmitter receptor, which is involved in various brain functions such as memory and mood. It mediates signaling via the interaction with a stimulatory G-protein. Especially, the third intracellular loop (iL3) of 5-HT(6) and the alpha subunit of stimulatory G protein (Galpha(s)) are responsible for the signaling process of 5-HT(6). Chemical compounds that could inhibit the interaction between the iL3 region of 5-HT(6) and Galpha(s) were screened from a chemical library consisted of 5,600 synthetic compounds. One of the identified compounds bound to Galpha(s) and effectively blocked the interaction between Galpha(s) and the iL3 region of 5-HT(6). The identified compound was further shown to reduce the serotonin-induced accumulation of cAMP in 293T cells transformed with 5-HT(6) cDNA. It also lowered the Ca2+ efflux induced by serotonin in cells expressing 5-HT(6) and chimeric Galpha(s5/q). These results indicate that the interaction between the iL3 of 5-HT(6) and Galpha(s) can be exploited for screening of regulatory compounds against the signaling pathway of 5-HT(6).
Animals
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Calcium/metabolism
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Cell Line
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Cephalosporins/*pharmacology
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Cricetinae
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Cricetulus
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Cyclic AMP/biosynthesis
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GTP-Binding Protein alpha Subunits, Gs/antagonists & inhibitors/*metabolism
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Humans
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Receptors, Serotonin/*drug effects/metabolism/*physiology
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Serotonin/pharmacology
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Serotonin Antagonists/pharmacology
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Signal Transduction
8.Anti-arrhythmic effect of acupuncture pretreatment in the rat of myocardial ischemia the post-receptor signaling pathway of beta-adrenergic receptor.
Jun-hong GAO ; Wei-xing FU ; Zhi-gao JIN ; Xiao-chun YU
Chinese Acupuncture & Moxibustion 2006;26(6):431-435
OBJECTIVETo observe anti-arrhythmic effect of acupuncture pretreatment in the rat of myocardial ischemia and reperfusion (MIR) and to explore the role of cAMP and Gsa protein in beta-adrenergic receptor signaling.
METHODSMIR was produced by ligation and reperfusion of the left anterior descending coronary artery in the rat. Arrhythmic score, content of cAMP and Gsalpha protein in ischemic myocardium were compared among the normal control (NC), ischemia and reperfusion (IR), electroacupuncture (EA) and EA plus propranolol (EAP) groups.
RESULTSThe arrhythmic score in the IR group at 10 min after reperfusion was higher than the NC group (P < 0.01); in the EA group the score was decreased (P < 0.01 vs the IR group); the score in the EAP group was similar to the IR group, much higher than the EA group (P < 0.01). The similar results for the contents of cAMP and Gsalpha protein were found in the ischemic myocardium. It is suggested that EA pretreatment significantly attenuates the arrhythmic incidence rate and the enhancement of the contents of myocardial cAMP and Gsalpha protein induced by MIR, and the attenuating effect is significantly inhibited by the intraperitoneal pretreatment of propranolol, a specific beta-adrenoceptor antagonist.
CONCLUSIONPretreatment of EA can produce anti-arrhythmic effect in the rat of MIR, which is mediated by the post-receptor signaling pathway of beta-adrenergic receptor.
Acupuncture Therapy ; Animals ; Arrhythmias, Cardiac ; prevention & control ; Calcium ; metabolism ; Cyclic AMP ; analysis ; GTP-Binding Protein alpha Subunits, Gs ; analysis ; Male ; Myocardial Ischemia ; metabolism ; therapy ; Rats ; Rats, Wistar ; Receptors, Adrenergic, beta ; physiology ; Signal Transduction ; physiology
9.Estrogen reduced myocardial damage by regulating Gαs-cAMP pathway in isoprenaline injured rats.
Li-Li SANG ; Chun-Le ZHOU ; Lu FU ; Jing-Min CHONG ; Lin ZHANG ; Xian-Cun CAO ; Hong SUN
Acta Physiologica Sinica 2014;66(5):583-588
The aim of the present study is to explore the mechanism of estrogen on regulating cardiac function disorder by adjusting the stimulating adenylate cyclase G α protein (Gαs)-cycle adenosine monophosphate (cAMP) signal pathway. Adult female rats were randomly divided into five groups: sham group, ovariectomized group (OVX), OVX and 17β-estradiol given group (OVX+E₂), OVX and isoprenaline injected group (OVX+ISO), OVX and 17β-estradiol, isoprenaline injected group (OVX+E₂+ISO). Rats were ovariectomized, and two weeks later, OVX+E₂group was injected with E₂, OVX+ISO group was injected with ISO, OVX+E₂+ISO group was injected with E₂and ISO. Another four weeks later, the hemodynamic parameters were monitored by carotid artery intubation: left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), maximal differentials of left ventricular developed pressure (+dp/dt(max)), and minimal differentials of left ventricular developed pressure (-dp/dt(max)). Brain natriuretic peptide (BNP) and cAMP concentration in plasma were determined; Gα(s) protein expression in myocardium was determined. The results showed that the hemodynamic parameters, the concentration of BNP and cAMP in plasma had no significant changes after ovariectomy compared with sham group. But after isoprenaline injection in ovariectomized rats, LVSP and +dp/dt(max) declined (P < 0.01), LVEDP and -dp/dt(max) elevated (P < 0.01); plasma BNP concentration increased (P < 0.01); plasma cAMP concentration decreased (P < 0.01), compared with OVX group. Further estrogen supplements improved the heart function treated by isoprenaline: LVSP and +dp/dt(max) elevated (P < 0.01), LVEDP and -dp/dtmax declined (P < 0.05, P < 0.01); the plasma BNP concentration decreased (P < 0.01); the plasma cAMP concentration increased (P < 0.01). Estrogen had no significant influence on Gαs protein expression. The results suggest that estrogen can alleviate myocardial injury and regulate cardiac function disorder by increasing cAMP level, finally improved the excessive suppression of myocardium.
Animals
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Cyclic AMP
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blood
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Estradiol
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pharmacology
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Estrogens
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pharmacology
;
Female
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GTP-Binding Protein alpha Subunits, Gs
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metabolism
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Hemodynamics
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Isoproterenol
;
adverse effects
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Myocardium
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pathology
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Natriuretic Peptide, Brain
;
blood
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Ovariectomy
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Rats
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Signal Transduction
10.Abnormal expression of c-myc, p53, p16 protein and GNAS1 gene mutation in fibrous dysplasia.
Juan TANG ; Hong-ye ZHAO ; Li ZHENG ; Hui-zhen ZHANG ; Zhi-ming JIANG
Chinese Journal of Pathology 2009;38(5):292-297
OBJECTIVETo study the significance of c-myc, p53 and p16 protein expression in fibrous dysplasia, to detect the GNAS1 gene mutation in fibrous dysplasia, and to explore the property of fibrous dysplasia.
METHODSThe expression of c-myc, p53 and p16 protein was evaluated by immunohistochemistry SP method in 35 cases of fibrous dysplasia including 1 FD with malignancy, 1 Mazabraud syndrome and 20 control cases (10 cases of bony callus, 10 cases of osteosarcoma). Genomic DNA extraction, PCR amplification and gene sequencing were used to detect GNAS1 gene mutation in 35 cases of fibrous dysplasia.
RESULTSC-myc protein immunoreactivity was detected in 91 percentage of FD (P = 0.001). Compared with the negative control group, the difference was significant. P16 positive was detected in 34 FD cases (P = 0.001). The difference was significant as compared with the positive control group. Positive p53 protein expression was detected in the only 1 case of fibrous dysplasia with malignant transformation. PCR amplification was successful in 12 of 35 FD cases. Two of the 12 FD cases were detected to have GNAS1 gene mutation, in which 1 case was FD of Mazabraud syndrome, 1 case was a monostotic lesion.
CONCLUSIONSC-myc could be another protooncogene in addition to c-fos in the fibrous dysplasia disease. P53 protein overexpression could be useful in the diagnosis of FD malignancy and in the prediction of the prognosis of FD. The abnormal expression of the gene p16 might play an important role in the formation of FD. The GNAS1 mutation exist in FD. All of the results indicate that FD could be a neoplasia disease, caused by multiple factors leading to a dysfunction of bone development.
Adolescent ; Adult ; Child ; Chromogranins ; Cyclin-Dependent Kinase Inhibitor p16 ; metabolism ; Female ; Fibrous Dysplasia of Bone ; genetics ; metabolism ; pathology ; GTP-Binding Protein alpha Subunits, Gs ; genetics ; Humans ; Male ; Middle Aged ; Mutation ; Osteosarcoma ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins c-myc ; metabolism ; Tumor Suppressor Protein p53 ; metabolism ; Young Adult