Fusarium graminearum is an important plant pathogenic fungus that causes disease and yield reduction in many cereal crops, such as wheat and barley. Gyp8 stimulates GTP hydrolysis on Ypt1 in yeast. However, the functi...Fusarium graminearum is an important plant pathogenic fungus that causes disease and yield reduction in many cereal crops, such as wheat and barley. Gyp8 stimulates GTP hydrolysis on Ypt1 in yeast. However, the functions of Gyp8 in plant pathogenic fungi are still unknown. In this study, we investigated the roles of Fg Gyp8 in F. graminearum by genetic and pathological analyses. Through gene knockout and phenotypic analyses, we found that Fg Gyp8 is required for vegetative growth in F. graminearum. The conidiation, conidial size and number of septa per conidium of ΔFggyp8 mutant are significantly reduced when compared to the wild type PH-1. Furthermore, Fg Gyp8 is crucial for pathogenicity on wheat coleoptiles and wheat heads. Fg Gyp8 contains a conserved TBC domain. Domain deletion analysis showed that the TBC domain, C-and N-terminal regions of Fg Gyp8 are all important for its biological functions in F. graminearum. Moreover, we showed that Fg Gyp8 catalyzes the hydrolysis of the GTP on Fg Rab1 to GDP in vitro, indicating that Fg Gyp8 is a GTPase-activating protein(GAP) for Fg Rab1. In addition, we demonstrated that Fg Gyp8 is required for Fg Snc1-mediated fusion of secretory vesicles with the plasma membrane in F. graminearum. Finally, we showed that Fg Gyp8 has functional redundancy with another Fg Rab1 GAP, Fg Gyp1, in F. graminearum. Taken together, we conclude that Fg Gyp8 is required for vegetative growth, conidiogenesis, pathogenicity and acts as a GAP for Fg Rab1 in F. graminearum.展开更多
MgCdc42 （Cdc42 in Magnaporthe grisea）, with high homology to ScCdc42 （Cdc42 in Saccharomyces cerevisiae）, has been demonstrated to involve in the morphogenesis and infection process. To further understand the sign...MgCdc42 （Cdc42 in Magnaporthe grisea）, with high homology to ScCdc42 （Cdc42 in Saccharomyces cerevisiae）, has been demonstrated to involve in the morphogenesis and infection process. To further understand the signaling network, the putative MgCdc42-interacting proteins were analyzed. ScCdc42-interacting protein sequences were first used to BLAST against the M. grisea genome database to retrieve their corresponding analogs. Subsequently, conserved domains of these proteins were compared and expression patterns of their encoding genes in different MgCdc42 mutation states were analyzed by semiquantitative RT-PCR. All retrieved analogs of ScCdc42-interacting proteins from the M. grisea database have conserved domains as those in S. cerevisiae. Expression of their encoding genes increased in MgCdc42CA mutant and decreased in MgCdc42KO mutant. However, MgBeml, Chin1, and MgGicl in MgCdc42DN mutant had the same expression level as that in the wild type, although MgBem4, MgBoi2, MgCdc24, MgGic2, MgRgal, and Mst20 had decreased expression level, as expected. Overall, it is concluded that there may exist a similar Cdc42 signal pathway in M. grisea as in S. cerevisiae and MgCdc42 plays a key role in the pathway.展开更多
Objective： To investigate the relationship between the excitotoxicity and seruminducible kinase （SNK） and spine-associated Rap GTPase-activating protein （SPAR） pathway in primary hippocampal neuron injury induced...Objective： To investigate the relationship between the excitotoxicity and seruminducible kinase （SNK） and spine-associated Rap GTPase-activating protein （SPAR） pathway in primary hippocampal neuron injury induced by glutamate and furthermore, to explore the molecular mechanism of neuroprotection of Zibu Piyin Recipe （滋补脾阴方药, ZBPYR） and the relationship between ZBPYR and the morphological regulation of dendritic spines. Methods： The serum containing ZBPYR was prepared by seropharmacology. Reverse transcription and polymerase chain reaction （RT-PCR） was used to detect the expression of mRNA for SNK, SPAR, postsynaptic density protein 95 （PSD-95） and N-methyI-D-aspartate （NMDA） receptor subunits （NR1, NR2A and NR2B） in primary rat hippocampal neuron cultures after pretreatment with 10 μmol/L glutamate and ZBPYR serum. Results： ZBPYR serum pretreatment resulted in a significant down-regulation of glutamate-induced SNK mRNA expression （P〈0.05）. Significant up-regulation was seen on the mRNA expression of SPAR and PSD-95 （P〈0.05）. All these changes were dose-dependent. The mRNA expression of NR1, NR2A and NR2B was down-regulated to different degrees （P〈0.05）. Conclusion： The mechanism of effect of ZBPYR on glutamate-induced excitotoxicity may be related to the regulation of SNK-SPAR signal pathway. ZBPYR may play a role in protecting and maintaining the normal morphology and structure of dendritic spines, which may be achieved by inhibiting the excessive activation of NMDA receptors.展开更多
This study investigated the possible involvement of microRNAs in the regulation of genes that participate in peripheral neural regeneration. A microRNA microarray analysis was conducted and 23 microRNAs were identiife...This study investigated the possible involvement of microRNAs in the regulation of genes that participate in peripheral neural regeneration. A microRNA microarray analysis was conducted and 23 microRNAs were identiifed whose expression was signiifcantly changed in rat dorsal root ganglia after sciatic nerve transection. The expression of one of the downregulated microRNAs, microRNA-214, was validated using quantitative reverse transcriptase-PCR. MicroRNA-214 was predicted to target the 3′-untranslated region of Slit-Robo GTPase-activating protein 3. In situ hybridization veriifed that microRNA-214 was located in the cytoplasm of dorsal root ganglia primary neurons and was downregulated following sciatic nerve transection. Moreover, a com-bination of in situ hybridization and immunohistochemistry revealed that microRNA-214 and Slit-Robo GTPase-activating protein 3 were co-localized in dorsal root ganglion primary neu-rons. Western blot analysis suggested that Slit-Robo GTPase-activating protein 3 was upregulated in dorsal root ganglion neurons after sciatic nerve transection. These data demonstrate that mi-croRNA-214 is located and differentially expressed in dorsal root ganglion primary neurons and may participate in regulating the gene expression of Slit-Robo GTPase-activating protein 3 after sciatic nerve transection.展开更多
Polar auxin transport,which depends on polarized subcellular distribution of AUXIN RESISTANT 1/LIKE AUX1（AUX1/LAX） influx carriers and PIN-FORMED（PIN） efflux carriers,mediates various processes of plant growth and...Polar auxin transport,which depends on polarized subcellular distribution of AUXIN RESISTANT 1/LIKE AUX1（AUX1/LAX） influx carriers and PIN-FORMED（PIN） efflux carriers,mediates various processes of plant growth and development.Endosomal recycling of PIN1 is mediated by an adenosine diphosphate（ADP）ribosylation factor（ARF）-GTPase exchange factor protein,GNOM.However,the mediation of auxin influx carrier recycling is poorly understood.Here,we report that overexpression of OsAGAP,an ARF-GTPase-activating protein in rice,stimulates vesicle transport from the plasma membrane to the Golgi apparatus in protoplasts and transgenic plants and induces the accumulation of early endosomes and AUX1.AUX1 endosomes could partially colocalize with FM4-64 labeled early endosome after actin disruption.Furthermore,OsAGAP is involved in actin cytoskeletal organization,and its overexpression tends to reduce the thickness and bundling of actin filaments.Fluorescence recovery after photobleaching analysis revealed exocytosis of the AUX1 recycling endosome was not affected in the OsAGAP overexpression cells,and was only slightly promoted when the actin filaments were completely disrupted by Lat B.Thus,we propose that AUX1 accumulation in the OsAGAP overexpression and actin disrupted cells may be due to the fact that endocytosis of the auxin influx carrier AUX1 early endosome was greatly promoted by actin cytoskeleton disruption.展开更多
基金National Natural Science Foundation of China (31970141)the Natural Science Foundation of Fujian Province, China (2020J06047)+1 种基金the Foundation of Minjiang University, China (MJY19019)the Foundation of Fujian Agriculture and Forestry University, China (KFb22050XA)。
文摘Fusarium graminearum is an important plant pathogenic fungus that causes disease and yield reduction in many cereal crops, such as wheat and barley. Gyp8 stimulates GTP hydrolysis on Ypt1 in yeast. However, the functions of Gyp8 in plant pathogenic fungi are still unknown. In this study, we investigated the roles of Fg Gyp8 in F. graminearum by genetic and pathological analyses. Through gene knockout and phenotypic analyses, we found that Fg Gyp8 is required for vegetative growth in F. graminearum. The conidiation, conidial size and number of septa per conidium of ΔFggyp8 mutant are significantly reduced when compared to the wild type PH-1. Furthermore, Fg Gyp8 is crucial for pathogenicity on wheat coleoptiles and wheat heads. Fg Gyp8 contains a conserved TBC domain. Domain deletion analysis showed that the TBC domain, C-and N-terminal regions of Fg Gyp8 are all important for its biological functions in F. graminearum. Moreover, we showed that Fg Gyp8 catalyzes the hydrolysis of the GTP on Fg Rab1 to GDP in vitro, indicating that Fg Gyp8 is a GTPase-activating protein(GAP) for Fg Rab1. In addition, we demonstrated that Fg Gyp8 is required for Fg Snc1-mediated fusion of secretory vesicles with the plasma membrane in F. graminearum. Finally, we showed that Fg Gyp8 has functional redundancy with another Fg Rab1 GAP, Fg Gyp1, in F. graminearum. Taken together, we conclude that Fg Gyp8 is required for vegetative growth, conidiogenesis, pathogenicity and acts as a GAP for Fg Rab1 in F. graminearum.
基金the National Natural Science Foundation of China to Wang Zonghua （30070030, 30470066）.
文摘MgCdc42 （Cdc42 in Magnaporthe grisea）, with high homology to ScCdc42 （Cdc42 in Saccharomyces cerevisiae）, has been demonstrated to involve in the morphogenesis and infection process. To further understand the signaling network, the putative MgCdc42-interacting proteins were analyzed. ScCdc42-interacting protein sequences were first used to BLAST against the M. grisea genome database to retrieve their corresponding analogs. Subsequently, conserved domains of these proteins were compared and expression patterns of their encoding genes in different MgCdc42 mutation states were analyzed by semiquantitative RT-PCR. All retrieved analogs of ScCdc42-interacting proteins from the M. grisea database have conserved domains as those in S. cerevisiae. Expression of their encoding genes increased in MgCdc42CA mutant and decreased in MgCdc42KO mutant. However, MgBeml, Chin1, and MgGicl in MgCdc42DN mutant had the same expression level as that in the wild type, although MgBem4, MgBoi2, MgCdc24, MgGic2, MgRgal, and Mst20 had decreased expression level, as expected. Overall, it is concluded that there may exist a similar Cdc42 signal pathway in M. grisea as in S. cerevisiae and MgCdc42 plays a key role in the pathway.
基金the National Natural Science Foundation of China(No.30472255)
文摘Objective： To investigate the relationship between the excitotoxicity and seruminducible kinase （SNK） and spine-associated Rap GTPase-activating protein （SPAR） pathway in primary hippocampal neuron injury induced by glutamate and furthermore, to explore the molecular mechanism of neuroprotection of Zibu Piyin Recipe （滋补脾阴方药, ZBPYR） and the relationship between ZBPYR and the morphological regulation of dendritic spines. Methods： The serum containing ZBPYR was prepared by seropharmacology. Reverse transcription and polymerase chain reaction （RT-PCR） was used to detect the expression of mRNA for SNK, SPAR, postsynaptic density protein 95 （PSD-95） and N-methyI-D-aspartate （NMDA） receptor subunits （NR1, NR2A and NR2B） in primary rat hippocampal neuron cultures after pretreatment with 10 μmol/L glutamate and ZBPYR serum. Results： ZBPYR serum pretreatment resulted in a significant down-regulation of glutamate-induced SNK mRNA expression （P〈0.05）. Significant up-regulation was seen on the mRNA expression of SPAR and PSD-95 （P〈0.05）. All these changes were dose-dependent. The mRNA expression of NR1, NR2A and NR2B was down-regulated to different degrees （P〈0.05）. Conclusion： The mechanism of effect of ZBPYR on glutamate-induced excitotoxicity may be related to the regulation of SNK-SPAR signal pathway. ZBPYR may play a role in protecting and maintaining the normal morphology and structure of dendritic spines, which may be achieved by inhibiting the excessive activation of NMDA receptors.
基金supported by the National Natural Science Foundation of China,No.81160158 and 30860290
文摘This study investigated the possible involvement of microRNAs in the regulation of genes that participate in peripheral neural regeneration. A microRNA microarray analysis was conducted and 23 microRNAs were identiifed whose expression was signiifcantly changed in rat dorsal root ganglia after sciatic nerve transection. The expression of one of the downregulated microRNAs, microRNA-214, was validated using quantitative reverse transcriptase-PCR. MicroRNA-214 was predicted to target the 3′-untranslated region of Slit-Robo GTPase-activating protein 3. In situ hybridization veriifed that microRNA-214 was located in the cytoplasm of dorsal root ganglia primary neurons and was downregulated following sciatic nerve transection. Moreover, a com-bination of in situ hybridization and immunohistochemistry revealed that microRNA-214 and Slit-Robo GTPase-activating protein 3 were co-localized in dorsal root ganglion primary neu-rons. Western blot analysis suggested that Slit-Robo GTPase-activating protein 3 was upregulated in dorsal root ganglion neurons after sciatic nerve transection. These data demonstrate that mi-croRNA-214 is located and differentially expressed in dorsal root ganglion primary neurons and may participate in regulating the gene expression of Slit-Robo GTPase-activating protein 3 after sciatic nerve transection.
基金supported by the Innovative Program of the Chinese Academy of Sciences (KSCX2-YW-N-041)the National Natural Science Foundation of China(30670197)
文摘Polar auxin transport,which depends on polarized subcellular distribution of AUXIN RESISTANT 1/LIKE AUX1（AUX1/LAX） influx carriers and PIN-FORMED（PIN） efflux carriers,mediates various processes of plant growth and development.Endosomal recycling of PIN1 is mediated by an adenosine diphosphate（ADP）ribosylation factor（ARF）-GTPase exchange factor protein,GNOM.However,the mediation of auxin influx carrier recycling is poorly understood.Here,we report that overexpression of OsAGAP,an ARF-GTPase-activating protein in rice,stimulates vesicle transport from the plasma membrane to the Golgi apparatus in protoplasts and transgenic plants and induces the accumulation of early endosomes and AUX1.AUX1 endosomes could partially colocalize with FM4-64 labeled early endosome after actin disruption.Furthermore,OsAGAP is involved in actin cytoskeletal organization,and its overexpression tends to reduce the thickness and bundling of actin filaments.Fluorescence recovery after photobleaching analysis revealed exocytosis of the AUX1 recycling endosome was not affected in the OsAGAP overexpression cells,and was only slightly promoted when the actin filaments were completely disrupted by Lat B.Thus,we propose that AUX1 accumulation in the OsAGAP overexpression and actin disrupted cells may be due to the fact that endocytosis of the auxin influx carrier AUX1 early endosome was greatly promoted by actin cytoskeleton disruption.
