hrp mutants were produced from strain JXOIII of Xanthomonas oryzae pv. oryzae (Xoo) and strain RS105 of X.o. pv. oryzicola (Xooc), respectively, by using diethyl sulfate (DES) as a mutagenic che ...hrp mutants were produced from strain JXOIII of Xanthomonas oryzae pv. oryzae (Xoo) and strain RS105 of X.o. pv. oryzicola (Xooc), respectively, by using diethyl sulfate (DES) as a mutagenic che mical. All the hrp mutants lost their pathogenicity on a susceptible host plant, rice (Shanyou63), and elicitation of the hypersensitive response (HR) on a nonhost plant, tobacco (NC89). Extracellular enzyme (amy lase, pectate lyase, proteinase, cellulase and lipase) activities of all the hrp mutants were similar to those of the corresponding wild type strains. The response of tobacco to cell sonicated integrations of the wild type strains and the hrp mutants demonstrated that there existed an HR eliciting substance which was heat stable and sensitive to protease. No HR appeared on tobacco after infiltration of the lipopolysaccharide (LPS) of both the wild strains and hrp mutants into tobacco leaves. The ability of the Xooc hrp mutants to induce HR on tobacco and cause streak disease on rice was restored by complementation with pUHRX245 from JXOIII genomic DNA library and by pUHRS138 from RS105 genomic DNA library, respectively. Subcloning of a 38.6 kb hrp fragment insert in pUHRX245 and a 39.3 kb insert in pUHRS138 revealed that a 3.3 kb Sac Ⅰ fragment from pUHRX245 and a 4.5 kb Bam HⅠ Kpn Ⅰ fragment from pUHRS138 were the minimal functional portions required for restoration of the ability of Xooc hrp mutants to induce HR on tobacco and cause disease on rice. The disease symptom caused by the conjugant (M1005 plus 3.3 kb) on rice was similar to that caused by the wild type of Xooc. It suggests that the two fragments contain the same hrp gene(s) and are responsible reciprocally for HR induction on tobacco and pathogenicity on rice.展开更多
AIM:To analyze gene expression profiles in an experimental pancreatitis and provide functional reversal of hypersensitivity with candidate gene endothelin-1 antagonists.METHODS:Dibutyltin dichloride(DBTC) is a chemica...AIM:To analyze gene expression profiles in an experimental pancreatitis and provide functional reversal of hypersensitivity with candidate gene endothelin-1 antagonists.METHODS:Dibutyltin dichloride(DBTC) is a chemical used as a polyvinyl carbonate stabilizer/catalyzer,biocide in agriculture,antifouling agent in paint and fabric.DBTC induces an acute pancreatitis flare through generation of reactive oxygen species.Lewis-inbred rats received a single i.v.injection with either DBTC or vehicle.Spinal cord and dorsal root ganglia(DRG) were taken at the peak of inflammation and processed for transcriptional profiling with a cDNA microarray biased for rat brain-specific genes.In a second study,groups of animals with DBTC-induced pancreatitis were treated with endothelin(ET) receptor antagonists [ET-A(BQ123) and ET-B BQ788)].Spontaneous pain related mechanical and thermal hypersensitivity were measured.Immunohistochemical analysis was performed using anti-ET-A and ET-B antibodies on sections from pancreatic tissues and DRG of the T10-12 spinal segments.RESULTS:Animals developed acute pancreatic inflammation persisting 7-10 d as confirmed by pathological studies(edema in parenchyma,loss of pancreatic architecture and islets,infiltration of inflammatory cells,neutrophil and mononuclear cells,degeneration,vacuolization and necrosis of acinar cells) and the painrelated behaviors(cutaneous secondary mechanical and thermal hypersensitivity).Gene expression profile was different in the spinal cord from animals with pancreatitis compared to the vehicle control group.Over 260 up-regulated and 60 down-regulated unique genes could be classified into 8 functional gene families:circulatory/acute phase/immunomodulatory;extracellular matrix;structural;channel/receptor/transporter;signaling transduction;transcription/translation-related;antioxidants/chaperones/heat shock;pancreatic and other enzymes.ET-1 was among the 52 candidate genes upregulated greater than 2-fold in animals with pancreatic inflammation and visceral pain-related behavior.Treatments with the ET-A(BQ123) and ET-B(BQ-788) antagonists revealed significant protection against inflammatory pain related mechanical and thermal hypersensitivity behaviors in animals with pancreatitis(P < 0.05).Open field spontaneous behavioral activity(at baseline,day 6 and 30 min after drug treatments(BQ123,BQ788) showed overall stable activity levels indicating that the drugs produced no undesirable effects on normal exploratory behaviors,except for a trend toward reduction of the active time and increase in resting time at the highest dose(300 μmol/L).Immunocytochemical localization revealed that expression of ET-A and ET-B receptors increased in DRG from animals with pancreatitis.Endothelin receptor localization was combined in dual staining with neuronal marker NeuN,and glia marker,glial fibrillary acidic protein.ET-A was expressed in the cell bodies and occasional nuclei of DRG neurons in na ve animals.However,phenotypic expression of ET-A receptor was greatly increased in neurons of all sizes in animals with pancreatitis.Similarly,ET-B receptor was localized in neurons and in the satellite glia,as well as in the Schwann cell glial myelin sheaths surrounding the axons passing through the DRG.CONCLUSION:Endothelin-receptor antagonists protect against inflammatory pain responses without interfering with normal exploratory behaviors.Candidate genes can serve as future biomarkers for diagnosis and/or targeted gene therapy.展开更多
Although Blufensins(Bln)have important functions in the response of plants to biotic stress the precise functioning of Bln in wheat remains largely unknown.Here we isolated a Bln gene(TaBln4)from Suwon 11 infected by ...Although Blufensins(Bln)have important functions in the response of plants to biotic stress the precise functioning of Bln in wheat remains largely unknown.Here we isolated a Bln gene(TaBln4)from Suwon 11 infected by Puccinia striiformis f.sp.tritici(Pst).Expression of TaBln4 increased in host plants at the early stage of infection with a virulent Pst race(CYR31)but was unchanged in response to infection by an avirulent race(CYR23).Transcription levels of TaBln4 were also regulated by hormone and abiotic stresses.Expression of TaBln4 in tobacco leaves suppressed Bax-induced programmed cell death.Knockdown of TaBln4 by virus-induced gene silencing inhibited colonization of race CYR31 by increasing the accumulation of H2O2 and formation of hypersensitive responses(HR).Transient overexpression of TaBln4 by a transient overexpression system(BSMV-VOX)increased the susceptibility of wheat to CYR31.Results from bimolecular fluorescence complementation and pull-down assays demonstrated that TaBLN4 interacted with calmodulin.Taken together,our results suggest that TaBln4 negatively regulates resistance in wheat to Pst in a reactive oxygen species(ROS)-and HR-dependent manner.展开更多
In this study, we analyze the binding of nuclear proteins isolated from the hydroxyurea (Hu)-induced and uninduced HEL cells to the DNasel hypersensitive sites Ⅲ (HS3 -14991 --14716 bp) and Ⅳ(HS4 -18586-18306 bp) in...In this study, we analyze the binding of nuclear proteins isolated from the hydroxyurea (Hu)-induced and uninduced HEL cells to the DNasel hypersensitive sites Ⅲ (HS3 -14991 --14716 bp) and Ⅳ(HS4 -18586-18306 bp) in the human p-globin gene locus control region (LCR). Using Western blot assay, we demonstrate that GATA-1 transcription factor in HEL cells is increased following the induction of Hu, while GATA-2 transcription factor is decreased. Based on both the competition EMSA and the Western blot assay, our data reveal that the nuclear protein isolated from the uninduced HEL cells, which can bind to the HS3 and HS4 core DNA sequences, is mainly GATA-2; however, the nuclear proteins isolated from the Hu-induced HEL cells, which can bind to the HS3 and HS4 core DNA sequences, are mainly GATA-1 and GATA-X (a kind of unknown GATA factor). These results suggest that the erythroid specific transcription factors (GATA-1 and GATA-2) in the Hu-induced and uninduced HEL cells can selectively bind to the HS3展开更多
HMG proteins are abundant chromosomal non-histone proteins. It has been suggested that the HMG proteins may play an important role in the structure and function of chromatin. In the present study, the binding of HMG p...HMG proteins are abundant chromosomal non-histone proteins. It has been suggested that the HMG proteins may play an important role in the structure and function of chromatin. In the present study, the binding of HMG proteins (HMG1/2 and HMG14/17) to the core DNA sequence of DNasel hypersensitive site 2 (HS2core DNA sequence, -10681-10970 bp) in the locus control region (LCR) of the human β-like globin gene cluster has been examined by using both the in vitro nucleosome reconstitution and the gel mobility shift assays. Here we show that HMG1/2 can bind to the naked HS2core DNA sequence, however, HMG 14/17 cannot. Using the in vitro nucleosome reconstitution we demonstrate that HMG14/17 can bind to the HS2core DNA sequence which is assembled into nucleosomes with the core histone octamer transferred from chicken erythrocytes. In contrast, HMG 1/2 cannot bind to the nucleosomes reconstituted in vitro with the HS2core DNA sequence. These results indicate that the binding patterns between HMG proteins and展开更多
文摘hrp mutants were produced from strain JXOIII of Xanthomonas oryzae pv. oryzae (Xoo) and strain RS105 of X.o. pv. oryzicola (Xooc), respectively, by using diethyl sulfate (DES) as a mutagenic che mical. All the hrp mutants lost their pathogenicity on a susceptible host plant, rice (Shanyou63), and elicitation of the hypersensitive response (HR) on a nonhost plant, tobacco (NC89). Extracellular enzyme (amy lase, pectate lyase, proteinase, cellulase and lipase) activities of all the hrp mutants were similar to those of the corresponding wild type strains. The response of tobacco to cell sonicated integrations of the wild type strains and the hrp mutants demonstrated that there existed an HR eliciting substance which was heat stable and sensitive to protease. No HR appeared on tobacco after infiltration of the lipopolysaccharide (LPS) of both the wild strains and hrp mutants into tobacco leaves. The ability of the Xooc hrp mutants to induce HR on tobacco and cause streak disease on rice was restored by complementation with pUHRX245 from JXOIII genomic DNA library and by pUHRS138 from RS105 genomic DNA library, respectively. Subcloning of a 38.6 kb hrp fragment insert in pUHRX245 and a 39.3 kb insert in pUHRS138 revealed that a 3.3 kb Sac Ⅰ fragment from pUHRX245 and a 4.5 kb Bam HⅠ Kpn Ⅰ fragment from pUHRS138 were the minimal functional portions required for restoration of the ability of Xooc hrp mutants to induce HR on tobacco and cause disease on rice. The disease symptom caused by the conjugant (M1005 plus 3.3 kb) on rice was similar to that caused by the wild type of Xooc. It suggests that the two fragments contain the same hrp gene(s) and are responsible reciprocally for HR induction on tobacco and pathogenicity on rice.
基金Supported by National Institutes of Health Grants,No. NS039041,to Westlund KN and DE19177,to Oz HS
文摘AIM:To analyze gene expression profiles in an experimental pancreatitis and provide functional reversal of hypersensitivity with candidate gene endothelin-1 antagonists.METHODS:Dibutyltin dichloride(DBTC) is a chemical used as a polyvinyl carbonate stabilizer/catalyzer,biocide in agriculture,antifouling agent in paint and fabric.DBTC induces an acute pancreatitis flare through generation of reactive oxygen species.Lewis-inbred rats received a single i.v.injection with either DBTC or vehicle.Spinal cord and dorsal root ganglia(DRG) were taken at the peak of inflammation and processed for transcriptional profiling with a cDNA microarray biased for rat brain-specific genes.In a second study,groups of animals with DBTC-induced pancreatitis were treated with endothelin(ET) receptor antagonists [ET-A(BQ123) and ET-B BQ788)].Spontaneous pain related mechanical and thermal hypersensitivity were measured.Immunohistochemical analysis was performed using anti-ET-A and ET-B antibodies on sections from pancreatic tissues and DRG of the T10-12 spinal segments.RESULTS:Animals developed acute pancreatic inflammation persisting 7-10 d as confirmed by pathological studies(edema in parenchyma,loss of pancreatic architecture and islets,infiltration of inflammatory cells,neutrophil and mononuclear cells,degeneration,vacuolization and necrosis of acinar cells) and the painrelated behaviors(cutaneous secondary mechanical and thermal hypersensitivity).Gene expression profile was different in the spinal cord from animals with pancreatitis compared to the vehicle control group.Over 260 up-regulated and 60 down-regulated unique genes could be classified into 8 functional gene families:circulatory/acute phase/immunomodulatory;extracellular matrix;structural;channel/receptor/transporter;signaling transduction;transcription/translation-related;antioxidants/chaperones/heat shock;pancreatic and other enzymes.ET-1 was among the 52 candidate genes upregulated greater than 2-fold in animals with pancreatic inflammation and visceral pain-related behavior.Treatments with the ET-A(BQ123) and ET-B(BQ-788) antagonists revealed significant protection against inflammatory pain related mechanical and thermal hypersensitivity behaviors in animals with pancreatitis(P < 0.05).Open field spontaneous behavioral activity(at baseline,day 6 and 30 min after drug treatments(BQ123,BQ788) showed overall stable activity levels indicating that the drugs produced no undesirable effects on normal exploratory behaviors,except for a trend toward reduction of the active time and increase in resting time at the highest dose(300 μmol/L).Immunocytochemical localization revealed that expression of ET-A and ET-B receptors increased in DRG from animals with pancreatitis.Endothelin receptor localization was combined in dual staining with neuronal marker NeuN,and glia marker,glial fibrillary acidic protein.ET-A was expressed in the cell bodies and occasional nuclei of DRG neurons in na ve animals.However,phenotypic expression of ET-A receptor was greatly increased in neurons of all sizes in animals with pancreatitis.Similarly,ET-B receptor was localized in neurons and in the satellite glia,as well as in the Schwann cell glial myelin sheaths surrounding the axons passing through the DRG.CONCLUSION:Endothelin-receptor antagonists protect against inflammatory pain responses without interfering with normal exploratory behaviors.Candidate genes can serve as future biomarkers for diagnosis and/or targeted gene therapy.
基金supported by the National Key Research and Development Program of China(2021YFD1401000)the International Science and Technology Cooperation Project of Shaanxi Provincial Key R&D Plan-Key Project(2020KWZ-009)+1 种基金the Shaanxi Innovation Team Project(2018TD-004)the 111 Project of the Ministry of Education of China(B07049).
文摘Although Blufensins(Bln)have important functions in the response of plants to biotic stress the precise functioning of Bln in wheat remains largely unknown.Here we isolated a Bln gene(TaBln4)from Suwon 11 infected by Puccinia striiformis f.sp.tritici(Pst).Expression of TaBln4 increased in host plants at the early stage of infection with a virulent Pst race(CYR31)but was unchanged in response to infection by an avirulent race(CYR23).Transcription levels of TaBln4 were also regulated by hormone and abiotic stresses.Expression of TaBln4 in tobacco leaves suppressed Bax-induced programmed cell death.Knockdown of TaBln4 by virus-induced gene silencing inhibited colonization of race CYR31 by increasing the accumulation of H2O2 and formation of hypersensitive responses(HR).Transient overexpression of TaBln4 by a transient overexpression system(BSMV-VOX)increased the susceptibility of wheat to CYR31.Results from bimolecular fluorescence complementation and pull-down assays demonstrated that TaBLN4 interacted with calmodulin.Taken together,our results suggest that TaBln4 negatively regulates resistance in wheat to Pst in a reactive oxygen species(ROS)-and HR-dependent manner.
文摘In this study, we analyze the binding of nuclear proteins isolated from the hydroxyurea (Hu)-induced and uninduced HEL cells to the DNasel hypersensitive sites Ⅲ (HS3 -14991 --14716 bp) and Ⅳ(HS4 -18586-18306 bp) in the human p-globin gene locus control region (LCR). Using Western blot assay, we demonstrate that GATA-1 transcription factor in HEL cells is increased following the induction of Hu, while GATA-2 transcription factor is decreased. Based on both the competition EMSA and the Western blot assay, our data reveal that the nuclear protein isolated from the uninduced HEL cells, which can bind to the HS3 and HS4 core DNA sequences, is mainly GATA-2; however, the nuclear proteins isolated from the Hu-induced HEL cells, which can bind to the HS3 and HS4 core DNA sequences, are mainly GATA-1 and GATA-X (a kind of unknown GATA factor). These results suggest that the erythroid specific transcription factors (GATA-1 and GATA-2) in the Hu-induced and uninduced HEL cells can selectively bind to the HS3
文摘HMG proteins are abundant chromosomal non-histone proteins. It has been suggested that the HMG proteins may play an important role in the structure and function of chromatin. In the present study, the binding of HMG proteins (HMG1/2 and HMG14/17) to the core DNA sequence of DNasel hypersensitive site 2 (HS2core DNA sequence, -10681-10970 bp) in the locus control region (LCR) of the human β-like globin gene cluster has been examined by using both the in vitro nucleosome reconstitution and the gel mobility shift assays. Here we show that HMG1/2 can bind to the naked HS2core DNA sequence, however, HMG 14/17 cannot. Using the in vitro nucleosome reconstitution we demonstrate that HMG14/17 can bind to the HS2core DNA sequence which is assembled into nucleosomes with the core histone octamer transferred from chicken erythrocytes. In contrast, HMG 1/2 cannot bind to the nucleosomes reconstituted in vitro with the HS2core DNA sequence. These results indicate that the binding patterns between HMG proteins and
