Coordination between the sporophytic tissue and the gametic pollen within anthers is tightly controlled to achieve the optimal pollen fitness. Glucose-6-phosphate/phosphate translocator(GPT) transports glucose-6-phosp...Coordination between the sporophytic tissue and the gametic pollen within anthers is tightly controlled to achieve the optimal pollen fitness. Glucose-6-phosphate/phosphate translocator(GPT) transports glucose-6-phosphate, a key precursor of starch and/or fatty acid biosynthesis, into plastids. Here, we report the functional characterization of Os GPT1 in the rice anther development and pollen fertility. Pollen grains from homozygous osgpt1 mutant plants fail to accumulate starch granules, resulting in pollen sterility. Genetic analyses reveal a sporophytic effect for this mutation. Os GPT1 is highly expressed in the tapetal layer of rice anther. Degeneration of the tapetum, an important process to provide cellular contents to support pollen development, is impeded in osgpt1 plants. In addition, defective intine and exine are observed in the pollen from osgpt1 plants. Expression levels of multiple genes that are important to tapetum degeneration or pollen wall formation are significantly decreased in osgpt1 anthers. Previously, we reported that At GPT1 plays a gametic function in the accumulation of lipid bodies in Arabidopsis pollen. This report highlights a sporophytic role of Os GPT1 in the tapetum degeneration and pollen development. The divergent functions of Os GPT1 and At GPT1 in pollen development might be a result of their independent evolution after monocots and dicots diverged.展开更多
In plants, non-green plastids in heterotrophic tissues are sites for starch and fatty acids biosynthesis,which are essential for plant development and reproduction. Distinct from chloroplasts, the metabolites for thes...In plants, non-green plastids in heterotrophic tissues are sites for starch and fatty acids biosynthesis,which are essential for plant development and reproduction. Distinct from chloroplasts, the metabolites for these processes in non-green plastids have to be imported through specific transporters. Glucose 6-Phosphate/Phosphate Translocator 1 is required for the uptake of cytosolic Glucose 6-Phosphate into non-green plastids. In Arabidopsis, GPT1 has been demonstrated to play essential roles in male, female gametophyte and embryo development. However, the roles of GPTs in other species are yet largely unknown. Here, we reported that rice OsGPT1 is indispensable for normal tapetal degeneration and pollen exine formation during anther and pollen development. OsGPT1 is localized in the plastid and distributed in the anther wall layers and late-stage pollen grains. Different from the gametic defects caused by mutation in At GPT1, disruption of OsGPT1 does not affect male and female gamete transmission as well as embryo development. On the contrary, osgpt1 mutant exhibits delayed tapetum degeneration,decreased Ubisch bodies formation and thinner pollen exine, leading to pollen abortion at the mature stage. Furthermore, the expression of several genes involved in tapetal programmed cell death(PCD)and sporopollenin formation is decreased in osgpt1. Our study suggests that OsGPT1 coordinates the development of anther sporophytic tissues and the male gametophyte by integrating carbohydrate and fatty acid metabolism in the plastid.展开更多
Background The present study aimed to investigate the detailed mode and specific sites for their binding as well as the functional relevance of this binding in the phenotypic proliferation of vascular smooth muscle ce...Background The present study aimed to investigate the detailed mode and specific sites for their binding as well as the functional relevance of this binding in the phenotypic proliferation of vascular smooth muscle cells(SMCs). Methods CREG knocked-down SMCs were employed to evaluate the biological activity of wtCREG and mCREG.Expressions of SMC differentiation markers SM myosin heavy chain(SM-MHC),SM-actin,heavy caldesmon and myocardin were determined by Western blotting using specific antibodies. Cellular growth of SMCs was assessed by bromide dewuridine (BrdU) incorporation and cell cycle analysis on fluorescence-activated cell sorting(FACS).A solid-phase binding assay was used to study the binding of CREG to extracellular domains of M6P/IGF2R.The cellular co-localization of the two recombinant CREGs with M6P/IGF2R was detected on SMC surface by immunoprecipitation and immunofluorescence analysis.Results The molecular weight of wtCREG was around 30 kD while that of the mCREG was～25 kD.Treatment of wtCREG with PNGase F reduced its molecular weight from～30 kD to～25 kD,whereas PNGase F treatment had no effect on the molecular weight of mCREG.Both wtCREG and mCREG proteins enhanced SMC differentiation,inhibited BrdU incorporation,and arrested cell cycle progression when added to the culture medium.In CREG knocked-down SMCs,the amount of CREG detected by immunoblotting in M6P/IGF2R immunoprecipitates was significantly reduced when compared to normal cells.Both recombinant CREGs co-immunoprecipitated with M6P/IGF2R, although slightly reduced amount of the mutant CREG was detected in M6P/IGF2R immunoprecipitates.Immunostaining revealed that His-tagged CREGs co-localized with IGF2R on the cell surface in a glycosylation-independent manner.In vitro binding assay showed that CREGs bound to M6P/ IGF2R extracellular domains 7-10 and 11-13 in a glycosylation -dependent and -independent manner,respectively.Further blocking experiments using soluble M6P/IGF2R fragments and M6P/IGF2R neutralizing antibody indicated that the biological activities of recombinant CREGs in SMC growth and the up-regulation of SMC differentiation markers were all abolished by treatment with the M6P/IGF2R neutralizing antibody. However,although the growth inhibitory effect of wtCREG was nearly abolished by D7-10 or D11-13,the effect of mCREG was only reversed by Dll-13,indicating that the binding to domains 11-13 is required for CREG to modulate the proliferation of SMCs.Conclusions These data suggest that solubleCREG proteins can exert their biological function via binding to the extracellular domains 7-10 and 11-13 of cell surface M6P/IGF2R in both a glycosylation-dependent and -independent manner.展开更多
Objective:To investigate the correlation between the expression of glucose-6-phosphate dehydrogenase(G6PD)and the clinicopathological characteristics,prognosis and immune cell infiltration of hepatocellular carcinoma(...Objective:To investigate the correlation between the expression of glucose-6-phosphate dehydrogenase(G6PD)and the clinicopathological characteristics,prognosis and immune cell infiltration of hepatocellular carcinoma(HCC).Methods:The expression of G6PD in liver cancer tissues and normal tissues is extracted from TCGA and GEO databases,validated by immunohistochemistry,and the correlation between G6PD expression and clinical features is analyzed.The clinical significance of G6PD in liver cancer is assessed by Kaplan-Meier,Cox regression,and prognostic line graph models.Functional enrichment analysis is performed by protein-protein interaction(PPI)network,GO/KEGG,GSEA and for G6PD-associated differentially expressed genes(DEGs).TIMER and ssGSEA packages are used to assess the correlation between expression and the level of immune cell infiltration.Results:Analysis of TCGA and GEO datasets revealed that G6PD expression is significantly upregulated in hepatocellular carcinoma tissues(P<0.001).G6PD expression is associated with histological grade,pathological stage,T-stage,vascular infiltration,and AFP level(P<0.05);HCC patients in the low G6PD expression group had longer overall survival and better prognosis compared with the high G6PD expression group(P<0.05).The level of G6PD expression affects the levels of macrophages,dendritic cells,B cells,and follicular helper T cells in the tumor microenvironment.Conclusion:High expression of G6PD is a potential biomarker for poor prognosis of hepatocellular carcinoma,and G6PD may be a target for immunotherapy of HCC.展开更多
[Objective] The aim of this study was to perform genome-wide analysis of glucose-6-phosphate dehydrogenase（G6PDH） and reveal its evolution in Eucalyptus grandsis.[Method] The gene character,protein sequence and phyl...[Objective] The aim of this study was to perform genome-wide analysis of glucose-6-phosphate dehydrogenase（G6PDH） and reveal its evolution in Eucalyptus grandsis.[Method] The gene character,protein sequence and phylogenetic tree of G6PDH gene were analyzed by BLAST and other bioinformatics software within Eucalyptus grandsis whole genome database.[Result] Six G6PDH genes,including one cytomic type and five plastids,were detected in the E.grandsis genome.All the G6PDHs have conserved motifs of motif 1,motif 2,motif 3,motif 7,motif 9 and motif 11.Furthermore,promoter sequences of all E.grandsis G6PDH contain TATA box,enhancer,light-responsive,hormone-responsive and stress-responsive regulatory elements.[Conclusion] This study provided reference for the further revealing molecular function of E.grandsis G6PDH gene family展开更多
A full-length cDNA encoding fructose-6-phosphate, 2-kinase/fructose-2,6-bisphosphatase from maize (Zea mays L.) was cloned by the methods of reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplifica...A full-length cDNA encoding fructose-6-phosphate, 2-kinase/fructose-2,6-bisphosphatase from maize (Zea mays L.) was cloned by the methods of reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE), and designated as mF2KP. The encoded protein is composed of two regions. Its COOH-terminal region is catalytic region and homologous to the enzymes from other eukaryotes; and its NH 2-terminal region is common and special region only in plant. A truncated fragment of mF2KP covering integrated catalytic region was expressed in Escherichia coli. The fusion protein had the activities of fructose-6-phosphate, 2-kinase as well as fructose-2,6-bisphosphatase. Northern blot showed that the transcript level of mF2KP in seedlings initiated from strong-vigor seeds is lower than that from weak-vigor seeds.展开更多
基金supported by the grants from National Natural Science Foundation of China (31922005)Zhejiang Provincial Natural Science Foundation of China (LR18C020001)+2 种基金Zhejiang University K.P.Chao’s High Technology Development Foundation (2018RC016)the Young Elite Scientist Sponsorship Program by CAST (2018QNRC001)111 Project (B14027) to J.X。
文摘Coordination between the sporophytic tissue and the gametic pollen within anthers is tightly controlled to achieve the optimal pollen fitness. Glucose-6-phosphate/phosphate translocator(GPT) transports glucose-6-phosphate, a key precursor of starch and/or fatty acid biosynthesis, into plastids. Here, we report the functional characterization of Os GPT1 in the rice anther development and pollen fertility. Pollen grains from homozygous osgpt1 mutant plants fail to accumulate starch granules, resulting in pollen sterility. Genetic analyses reveal a sporophytic effect for this mutation. Os GPT1 is highly expressed in the tapetal layer of rice anther. Degeneration of the tapetum, an important process to provide cellular contents to support pollen development, is impeded in osgpt1 plants. In addition, defective intine and exine are observed in the pollen from osgpt1 plants. Expression levels of multiple genes that are important to tapetum degeneration or pollen wall formation are significantly decreased in osgpt1 anthers. Previously, we reported that At GPT1 plays a gametic function in the accumulation of lipid bodies in Arabidopsis pollen. This report highlights a sporophytic role of Os GPT1 in the tapetum degeneration and pollen development. The divergent functions of Os GPT1 and At GPT1 in pollen development might be a result of their independent evolution after monocots and dicots diverged.
基金supported the National Natural Science Foundation of China (U19A2031)the National Key Research and Development Program of China (2016YFD0100903)。
文摘In plants, non-green plastids in heterotrophic tissues are sites for starch and fatty acids biosynthesis,which are essential for plant development and reproduction. Distinct from chloroplasts, the metabolites for these processes in non-green plastids have to be imported through specific transporters. Glucose 6-Phosphate/Phosphate Translocator 1 is required for the uptake of cytosolic Glucose 6-Phosphate into non-green plastids. In Arabidopsis, GPT1 has been demonstrated to play essential roles in male, female gametophyte and embryo development. However, the roles of GPTs in other species are yet largely unknown. Here, we reported that rice OsGPT1 is indispensable for normal tapetal degeneration and pollen exine formation during anther and pollen development. OsGPT1 is localized in the plastid and distributed in the anther wall layers and late-stage pollen grains. Different from the gametic defects caused by mutation in At GPT1, disruption of OsGPT1 does not affect male and female gamete transmission as well as embryo development. On the contrary, osgpt1 mutant exhibits delayed tapetum degeneration,decreased Ubisch bodies formation and thinner pollen exine, leading to pollen abortion at the mature stage. Furthermore, the expression of several genes involved in tapetal programmed cell death(PCD)and sporopollenin formation is decreased in osgpt1. Our study suggests that OsGPT1 coordinates the development of anther sporophytic tissues and the male gametophyte by integrating carbohydrate and fatty acid metabolism in the plastid.
文摘Background The present study aimed to investigate the detailed mode and specific sites for their binding as well as the functional relevance of this binding in the phenotypic proliferation of vascular smooth muscle cells(SMCs). Methods CREG knocked-down SMCs were employed to evaluate the biological activity of wtCREG and mCREG.Expressions of SMC differentiation markers SM myosin heavy chain(SM-MHC),SM-actin,heavy caldesmon and myocardin were determined by Western blotting using specific antibodies. Cellular growth of SMCs was assessed by bromide dewuridine (BrdU) incorporation and cell cycle analysis on fluorescence-activated cell sorting(FACS).A solid-phase binding assay was used to study the binding of CREG to extracellular domains of M6P/IGF2R.The cellular co-localization of the two recombinant CREGs with M6P/IGF2R was detected on SMC surface by immunoprecipitation and immunofluorescence analysis.Results The molecular weight of wtCREG was around 30 kD while that of the mCREG was～25 kD.Treatment of wtCREG with PNGase F reduced its molecular weight from～30 kD to～25 kD,whereas PNGase F treatment had no effect on the molecular weight of mCREG.Both wtCREG and mCREG proteins enhanced SMC differentiation,inhibited BrdU incorporation,and arrested cell cycle progression when added to the culture medium.In CREG knocked-down SMCs,the amount of CREG detected by immunoblotting in M6P/IGF2R immunoprecipitates was significantly reduced when compared to normal cells.Both recombinant CREGs co-immunoprecipitated with M6P/IGF2R, although slightly reduced amount of the mutant CREG was detected in M6P/IGF2R immunoprecipitates.Immunostaining revealed that His-tagged CREGs co-localized with IGF2R on the cell surface in a glycosylation-independent manner.In vitro binding assay showed that CREGs bound to M6P/ IGF2R extracellular domains 7-10 and 11-13 in a glycosylation -dependent and -independent manner,respectively.Further blocking experiments using soluble M6P/IGF2R fragments and M6P/IGF2R neutralizing antibody indicated that the biological activities of recombinant CREGs in SMC growth and the up-regulation of SMC differentiation markers were all abolished by treatment with the M6P/IGF2R neutralizing antibody. However,although the growth inhibitory effect of wtCREG was nearly abolished by D7-10 or D11-13,the effect of mCREG was only reversed by Dll-13,indicating that the binding to domains 11-13 is required for CREG to modulate the proliferation of SMCs.Conclusions These data suggest that solubleCREG proteins can exert their biological function via binding to the extracellular domains 7-10 and 11-13 of cell surface M6P/IGF2R in both a glycosylation-dependent and -independent manner.
文摘Objective:To investigate the correlation between the expression of glucose-6-phosphate dehydrogenase(G6PD)and the clinicopathological characteristics,prognosis and immune cell infiltration of hepatocellular carcinoma(HCC).Methods:The expression of G6PD in liver cancer tissues and normal tissues is extracted from TCGA and GEO databases,validated by immunohistochemistry,and the correlation between G6PD expression and clinical features is analyzed.The clinical significance of G6PD in liver cancer is assessed by Kaplan-Meier,Cox regression,and prognostic line graph models.Functional enrichment analysis is performed by protein-protein interaction(PPI)network,GO/KEGG,GSEA and for G6PD-associated differentially expressed genes(DEGs).TIMER and ssGSEA packages are used to assess the correlation between expression and the level of immune cell infiltration.Results:Analysis of TCGA and GEO datasets revealed that G6PD expression is significantly upregulated in hepatocellular carcinoma tissues(P<0.001).G6PD expression is associated with histological grade,pathological stage,T-stage,vascular infiltration,and AFP level(P<0.05);HCC patients in the low G6PD expression group had longer overall survival and better prognosis compared with the high G6PD expression group(P<0.05).The level of G6PD expression affects the levels of macrophages,dendritic cells,B cells,and follicular helper T cells in the tumor microenvironment.Conclusion:High expression of G6PD is a potential biomarker for poor prognosis of hepatocellular carcinoma,and G6PD may be a target for immunotherapy of HCC.
基金Supported by Seeding Raising Project from Guangdong Provincial Department(LYM10040)Open Research Project of Key Laboratory for Genetics and Breeding in Forest Trees and Ornamental Plants,MOE,Beijing Forestry University(FOP2010-4)~~
文摘[Objective] The aim of this study was to perform genome-wide analysis of glucose-6-phosphate dehydrogenase（G6PDH） and reveal its evolution in Eucalyptus grandsis.[Method] The gene character,protein sequence and phylogenetic tree of G6PDH gene were analyzed by BLAST and other bioinformatics software within Eucalyptus grandsis whole genome database.[Result] Six G6PDH genes,including one cytomic type and five plastids,were detected in the E.grandsis genome.All the G6PDHs have conserved motifs of motif 1,motif 2,motif 3,motif 7,motif 9 and motif 11.Furthermore,promoter sequences of all E.grandsis G6PDH contain TATA box,enhancer,light-responsive,hormone-responsive and stress-responsive regulatory elements.[Conclusion] This study provided reference for the further revealing molecular function of E.grandsis G6PDH gene family
文摘A full-length cDNA encoding fructose-6-phosphate, 2-kinase/fructose-2,6-bisphosphatase from maize (Zea mays L.) was cloned by the methods of reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE), and designated as mF2KP. The encoded protein is composed of two regions. Its COOH-terminal region is catalytic region and homologous to the enzymes from other eukaryotes; and its NH 2-terminal region is common and special region only in plant. A truncated fragment of mF2KP covering integrated catalytic region was expressed in Escherichia coli. The fusion protein had the activities of fructose-6-phosphate, 2-kinase as well as fructose-2,6-bisphosphatase. Northern blot showed that the transcript level of mF2KP in seedlings initiated from strong-vigor seeds is lower than that from weak-vigor seeds.
