Translationally controlled tumor proteins (TCTP) and SNF1- related protein kinase (SnRK1) are conserved and widely present in eukaryotic cells. TCTP regulates cell division, plant growth and development, and mediates plant resistance against pathogen infection. SnRK1 participates in a range of physiological processes including sugar metabolism and resistance to abiotic and biotic stresses. Previous work in our laboratory demonstrated that wheat TCTP can respond to Puccinia triticina infection and induce host defense responses. In order to further investigate the mechanism of TaTCTP in wheat resistance to Puccinia triticina infection, we used TAP (tandem affinity purification) and mass spectrometry to screen the potential interactants of TaTCTP. A SNF1- related protein kinase (SnRK1) was identified as a potential interacting protein of TaTCTP. The results of yeast two-hybrid assay showed that TCTP could interact with SnRK1 in yeast, and the yeast carrying TCTP and SnRK1 could grow on SD/-Leu/-Trp/-His/-Ade (SD/-LWHA) medium. The fluorescence signal of the interaction between TCTP and SnRK1 was found to be distributed in the cytoplasm in the Bi-fluorescense complementation experiment. Co-IP experiments further showed that TCTP and SnRK1 could interact in plant cells. This study lays an important foundation for further studying the mechanism of TaTCTP in the interaction between wheat and Puccinia triticina, and it play a great influence on further improving the molecular mechanism of wheat resistant to Puccinia triticina.
Basidiomycota ; Humans ; Neoplasms ; Protein Biosynthesis ; Protein-Serine-Threonine Kinases ; Triticum

The aim of this study was to investigate the role of S100 calcium binding protein A16 (S100A16) in lipid metabolism in hepatocytes and its possible biological mechanism. HepG2 cells (human hepatoma cell line) were cultured with fatty acid to establish fatty acid culture model. The control model was cultured without fatty acid. Each model was divided into three groups and transfected with S100a16 over-expression, shRNA and vector plasmids, respectively. The concentration of triglyceride (TG) in the cells was measured by kit, and the lipid droplets was observed by oil red O staining. Immunoprecipitation and mass spectrometry were used to find the interesting proteins interacting with S100A16, and the interaction was verified by immunoprecipitation. The further mechanism was studied by Western blot and qRT-PCR. The results showed that the intracellular lipid droplet and TG concentrations in the fatty acid culture model were significantly higher than those in the control model. The accumulation of intracellular fat in the S100a16 over-expression group was significantly higher than that in the vector plasmid transfection group. There was an interaction between heat shock protein A5 (HSPA5) and S100A16. Over-expression of S100A16 up-regulated protein expression levels of HSPA5, inositol-requiring enzyme 1α (IRE1α) and pIREα1, which belong to endoplasmic reticulum stress HSPA5/IRE1α-XBP1 pathway. Meanwhile, over-expression of S100A16 up-regulated the mRNA expression levels of adipose synthesis-related gene Srebp1c, Acc and Fas. In the S100a16 shRNA plasmid transfection group, the above-mentioned protein and mRNA levels were lower than those of vector plasmid transfection group. These results suggest that S100A16 may promote lipid synthesis in HepG2 cells through endoplasmic reticulum stress HSPA5/IRE1α-XBP1 pathway.
Endoplasmic Reticulum Stress ; Endoribonucleases ; physiology ; Heat-Shock Proteins ; physiology ; Hep G2 Cells ; Humans ; Lipid Metabolism ; Protein-Serine-Threonine Kinases ; physiology ; S100 Proteins ; physiology ; Triglycerides ; biosynthesis ; X-Box Binding Protein 1 ; physiology

The present study was designed to elucidate whether the mechanism by which osthole decreases collagenI/III contents and their ratio is regulating the TGF-β/Smad signaling pathway in TGF-β1-overexpressed mouse cardiac fibroblasts (CFs). These CFs were cultured and treated with different concentrations of osthole. Our results showed that the TGF-β1 expression in the CFs transfected with that the recombinant expression plasmids pcDNA3.1(+)-TGF-β1 was significantly enhanced. After the CFs were treated with 1.25-5 μg·mL of osthole for 24 h, the mRNA and protein expression levels of collagensIand III were reduced. The collagen I/III ratio was also reduced. The mRNA and protein expression levels of TGF-β1, TβRI, Smad2/3, P-Smad2/3, Smad4, and α-SMA were decreased, whereas the expression level of Smad7 was increased. These effects suggested that osthole could inhibit collagen I and III expression and reduce their ratio via the TGF-β/Smad signaling pathway in TGF-β1 overexpressed CFs. These effects of osthole may play beneficial roles in the prevention and treatment of myocardial fibrosis.
Actins ; genetics ; Animals ; Cells, Cultured ; Collagen ; biosynthesis ; genetics ; Coumarins ; pharmacology ; Fibroblasts ; drug effects ; metabolism ; Gene Expression Regulation ; drug effects ; Mice ; Myocardium ; cytology ; Protein-Serine-Threonine Kinases ; genetics ; RNA, Messenger ; genetics ; Real-Time Polymerase Chain Reaction ; Receptor, Transforming Growth Factor-beta Type I ; Receptors, Transforming Growth Factor beta ; genetics ; Signal Transduction ; drug effects ; Smad Proteins ; genetics ; Transforming Growth Factor beta1 ; genetics

Emerging evidence indicates that aldosterone and mineralocorticoid receptors (MRs) are associated with the pathogenesis of erectile dysfunction. However, the molecular mechanisms remain largely unknown. In this study, freshly isolated penile corpus cavernosum tissue from rats was treated with aldosterone, with or without MRs inhibitors. Nuclear factor (NF)-kappa B (NF-κB) activity was evaluated by real-time quantitative PCR, luciferase assay, and immunoblot. The results demonstrated that mRNA levels of the NF-κB target genes, including inhibitor of NF-κB alpha (IκB-α), NF-κB1, tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6), were higher after aldosterone treatment. Accordingly, phosphorylation of p65/RelA, IκB-α, and inhibitor of NF-κB kinase-β was markedly increased by aldosterone. Furthermore, knockdown of MRs prevented activation of the NF-κB canonical pathway by aldosterone. Consistent with this finding, ectopic overexpression of MRs enhanced the transcriptional activation of NF-κB by aldosterone. More importantly, the MRs antagonist, spironolactone blocked aldosterone-mediated activation of the canonical NF-κB pathway. In conclusion, aldosterone has an inflammatory effect in the corpus cavernosum penis, inducing NF-κB activation via an MRs-dependent pathway, which may be prevented by selective MRs antagonists. These data reveal the possible role of aldosterone in erectile dysfunction as well as its potential as a novel pharmacologic target for treatment.
Aldosterone/pharmacology* ; Animals ; Cytokines/biosynthesis* ; Gene Knockdown Techniques ; I-kappa B Kinase/antagonists & inhibitors* ; Interleukin-6/genetics* ; Male ; Mineralocorticoid Receptor Antagonists/pharmacology* ; NF-kappa B/genetics* ; Penis/metabolism* ; Protein Serine-Threonine Kinases/antagonists & inhibitors* ; RNA, Messenger/biosynthesis* ; Rats ; Rats, Inbred WKY ; Receptors, Mineralocorticoid/genetics* ; Signal Transduction/drug effects* ; Spironolactone/pharmacology* ; Transcriptional Activation ; Tumor Necrosis Factor-alpha/biosynthesis* ; NF-kappaB-Inducing Kinase

The host takes use of pattern recognition receptors (PRRs) to defend against pathogen invasion or cellular damage. Among microorganism-associated molecular patterns detected by host PRRs, nucleic acids derived from bacteria or viruses are tightly supervised, providing a fundamental mechanism of host defense. Pathogenic DNAs are supposed to be detected by DNA sensors that induce the activation of NFκB or TBK1-IRF3 pathway. DNA sensor cGAS is widely expressed in innate immune cells and is a key sensor of invading DNAs in several cell types. cGAS binds to DNA, followed by a conformational change that allows the synthesis of cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) from adenosine triphosphate and guanosine triphosphate. cGAMP is a strong activator of STING that can activate IRF3 and subsequent type I interferon production. Here we describe recent progresses in DNA sensors especially cGAS in the innate immune responses against pathogenic DNAs.
DNA, Bacterial ; immunology ; metabolism ; DNA, Viral ; immunology ; metabolism ; Gene Expression Regulation ; Host-Pathogen Interactions ; Humans ; Immunity, Innate ; Interferon Regulatory Factor-3 ; genetics ; immunology ; Interferon Type I ; biosynthesis ; immunology ; Membrane Proteins ; genetics ; immunology ; Models, Molecular ; NF-kappa B ; genetics ; immunology ; Nucleotides, Cyclic ; biosynthesis ; immunology ; Nucleotidyltransferases ; genetics ; immunology ; Protein Binding ; Protein-Serine-Threonine Kinases ; genetics ; immunology ; Signal Transduction

The deubiquitinating enzyme ubiquitin specific peptidase 15 (USP15) is regarded as a regulator of TGFβ signaling pathway. This process depends on Smad7, the inhibitory factor of the TGFβ signal, and type I TGFβ receptor (TβR-I), one of the receptors of TGFβ. The expression level of USP15 seems to play vital roles in the pathogenesis of many neoplasms, but so far there has been no report about USP15 in psoriasis. In this study, immunohistochemical staining of USP15, TβR-I and Smad7 was performed in 30 paraffin-embedded psoriasis specimens and 10 normal specimens to investigate the expression of USP15, TβR-I and Smad7 in psoriasis and to explore the relevance among them. And USP15 small interfering RNA (USP15 siRNA) was used to transfect Hacat cells to detect the mRNA expression of TβR-I and Smad7. Of 30 cases of psoriasis in active stage, 28, 24 and 26 cases were positive for USP15, TβR-I and Smad7 staining, respectively. The positive rates of USP15 and Smad7 were significantly higher in psoriasis specimens than in normal skin specimens (44.1%±26.0% vs. 6.1%±6.6%, 47.2%±27.1% vs. 6.6%±7.1%), and positive rate of TβR-I (20.3%±22.2%) in psoriasis was lower than that in normal skin specimens (46.7%±18.2%). There was a significant positive correlation between USP15 and Smad7 expression, and significant negative correlations between USP15 and TβR-expression, an I d between TβR- and Smad7 expression I in psoriasis. After transfection of USP15 siRNA in Hacat cells, the expression of TβR-mRNA was up I -regulated and that of Smad7 was down-regulated. It is concluded that USP15 may play a role in the pathogenesis of psoriasis through regulating the TβR-I/Smad7 pathway and there may be other cell signaling pathways interacting with USP15 to take part in the development of psoriasis.
Adult ; Cell Line ; Female ; Gene Expression ; Humans ; Immunohistochemistry ; Keratinocytes ; cytology ; metabolism ; Male ; Middle Aged ; Protein-Serine-Threonine Kinases ; biosynthesis ; genetics ; Psoriasis ; genetics ; metabolism ; RNA Interference ; Receptors, Transforming Growth Factor beta ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; genetics ; Skin ; metabolism ; Smad7 Protein ; biosynthesis ; genetics ; Ubiquitin-Specific Proteases ; biosynthesis ; genetics ; Young Adult

Natural isoflavones and flavones are important dietary factors for prostate cancer prevention. We investigated the molecular mechanism of these compounds (genistein, biochanin-A and apigenin) in PC-3 (hormone-independent/p53 mutant type) and LNCaP (hormone-dependent/p53 wild type) prostate cancer cells. A cell growth rate and apoptotic activities were analyzed in different concentrations and exposure time to evaluate the antitumor activities of genistein, biochanin-A and apigenin. The real time PCR and Western blot analysis were performed to investigate whether the molecular mechanism of these compounds are involving the p21 and PLK-1 pathway. Apoptosis of prostate cancer cells was associated with p21 up-regulation and PLK-1 suppression. Exposure of genistein, biochanin-A and apigenin on LNCaP and PC-3 prostate cancer cells resulted in same pattern of cell cycle arrest and apoptosis. The inhibition effect for cell proliferation was slightly greater in LNCaP than PC-3 cells. In conclusion, flavonoids treatment induces up-regulation of p21 expression, and p21 inhibits transcription of PLK-1, which promotes apoptosis of cancer cells.
Antineoplastic Agents/*pharmacology ; Apigenin/pharmacology ; *Apoptosis ; Cell Cycle/drug effects ; Cell Cycle Proteins/biosynthesis/*genetics/metabolism ; Cell Line, Tumor ; Cell Proliferation ; Cyclin-Dependent Kinase Inhibitor p21/biosynthesis/*metabolism ; Flavonoids/*pharmacology ; Gene Expression Regulation, Neoplastic/drug effects ; Genistein/pharmacology ; Humans ; Male ; Prostatic Neoplasms/genetics/metabolism/*pathology ; Protein Kinase Inhibitors/pharmacology ; Protein-Serine-Threonine Kinases/biosynthesis/*genetics/metabolism ; Proto-Oncogene Proteins/biosynthesis/*genetics/metabolism ; Transcription, Genetic/drug effects

OBJECTIVETo investigate the effect of dracorhodin perchlorate (DP) on inhibiting high glucose-induced serum and glucocorticoid induced protein kinase 1 (SGK1) and fibronectin (FN) expression in human mesangial cells (HMC), and its mechanism of prevention and treatment on renal fibrosis in diabetic nephropathy (DN) .

METHODThe HMC were divided into normal glucose group (NG group, 5.5 mmol x L(-1) D-glucose), normal glucose +low DP group (NG + LDP group, 5.5 mmol x L(-1) D-glucose +7.5 micromol x L(-1) DP), normal glucose +high DP group (NG + HDP group, 5.5 mmol x L(-1) D-glucose + 15 micromol x L(-1) DP), high glucose group (HG group,25 mmol x L(-1) D-glucose), high glucose +low DP group (HG + LDP group, 25 mmol x L(-1) D-glucose + 7.5 micromol x L(-1) DP)and high glucose +high DP group (HG +HDP group, 25 mmol x L(-1) D-glucose + 15 micromol x L(-1) DP). Each group was examined at 24 hours. The levels of SGK1 and FN mRNA was detected by real-time fluorescence quantitative PCR,and the expression of SGK1 and FN protein was detected by Western blot or indirect immunofluorescence.

RESULTA basal level of SGK1 and FN in HMC were detected in NG group, and the level of SGK1 and FN mRNA and protein were not evidently different compared to that of NG group adding 7.5 micromol x L(-1) DP for 24 hours. On the other hand, the levels of SGK1 and FN mRNA and protein were obviously decreased by adding 15 micromol x L(-1) DP for 24 hours. Compared to NG group, the levels of SGK1 and FN mRNA and protein were increased in HG group after stimulating for 24 hours (P < 0.01). Compared to HG group, the level of SGK1 and FN mRNA and protein were evidently reduced in HG + LDP and HG + HDP groups by adding 7.5 micromol x L(-1) DP and 15 micromol x L(-1) DP for 24 hours (P < 0.01).

CONCLUSIONDracorhodin perchlorate can inhibit high glucose-induced serum and glucocorticoid induced protein kinase 1 (SGK1) and fibronectin(FN) expression in human mesangial cells, and this may be part of the mechanism of preventing and treating renal fibrosis of DN.

Benzopyrans ; pharmacology ; Cell Line ; Diabetic Nephropathies ; drug therapy ; enzymology ; genetics ; metabolism ; Down-Regulation ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Fibronectins ; biosynthesis ; genetics ; Gene Expression ; drug effects ; Glucose ; metabolism ; Humans ; Immediate-Early Proteins ; genetics ; metabolism ; Mesangial Cells ; drug effects ; enzymology ; metabolism ; Perchlorates ; pharmacology ; Protein-Serine-Threonine Kinases ; genetics ; metabolism

OBJECTIVE: To observe the effeet of rapamycin, an inhibitor of mammalian target of rapamycin (mTOR), on mTOR and eukaryotic initiation factor-4E(eIF-4E)expression in coxsac-kievirus B3 (CVB3)-induced rat myocardial cells and to investigate the role of mTOR/eIF-4E signal pathway in viral myocarditis. METHODS: To construct a cell model of viral myocarditis with primary cultured myocardial cells. Myocardial cells infected by CVB3 were treated with 10 nmol/L rapamycin according to the cell toxicity test. The mTOR and eIF-4E expressions of cells were determined by RT-PCR and Western Blot. RESULTS: Rapamycin inhibited the degeneration of CVB3-induced myocardial cells. Expressions of mTOR and eIF-4E mRNA or protein in CVB3-induced myocardial cells were significantly upregulated compared with the control group (P < 0.05), and rapamycin (10 nmol/L) inhibited the upregulation (P < 0.05). CONCLUSION Rapamycin can downregulate the expressions of mTOR and eIF-4E in CVB3-induced myocardial cells, suggesting that mTOR/eIF-4E signal transduction may play an important role in viral myocarditis.
Animals ; Animals, Newborn ; Coxsackievirus Infections ; Enterovirus B, Human ; Eukaryotic Initiation Factor-4E ; biosynthesis ; Myocarditis ; metabolism ; virology ; Myocytes, Cardiac ; metabolism ; Protein Kinases ; biosynthesis ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Sirolimus ; pharmacology ; TOR Serine-Threonine Kinases

The expression of Aurora B in normal endometria and endometrial carcinomas and its relation with clinicopathologic parameters of endometrial carcinomas were investigated. Streptavidin-biotin peroxidase (SP) immunohistochemical technique was used to detect the expression of Aurora B in 10 cases of normal proliferative phase endometria, 10 cases of normal secretory phase endometria and 72 cases of endometrial carcinomas respectively. According to the 1988 International Federation of Gynecology and Obstetrics (FIGO) grade, there were 37 patients in grade 1, 23 in grade 2 and 12 in grade 3 respectively. According to the FIGO stage, there were 59 patients in stage I-II and 13 patients in stage III-IV. Aurora B was expressed in both normal proliferative phase endometria, secretory phase endometria and endometrial carcinomas, but its positive labeling index (PLI) in proliferative phase endometria was significantly higher than that in secretory phase endometria (P<0.01) and endometrial carcinomas (P<0.01). The PLI of Aurora B was lower in tumors with well differentiation (G(1)), low surgical staging (I-II), and 1/2 myometrial invasion (all P<0.01). Aurora B exerts its functions in the replication of normal endometrial glandular cells; Expression of Aurora B is significantly correlated with biologic behavior of endometrial carcinoma, indicating that Aurora B may be a promising prognostic factor in endometrial carcinoma.
Carcinoma/*metabolism ; Cell Proliferation ; Endometrial Neoplasms/*metabolism ; Endometrium/*metabolism ; Gene Expression Regulation ; Gene Expression Regulation, Neoplastic ; Immunohistochemistry ; Prognosis ; Protein-Serine-Threonine Kinases/*biosynthesis