We investigated the microRNA172(miR172)-mediated regulatory network for the perception of changes in external and endogenous signals to identify a universally applicable floral regulation system in ornamental plants, ...We investigated the microRNA172(miR172)-mediated regulatory network for the perception of changes in external and endogenous signals to identify a universally applicable floral regulation system in ornamental plants, manipulation of which could be economically beneficial. Transgenic gloxinia plants, in which miR172 was either overexpressed or suppressed, were generated using Agrobacterium-mediated transformation. They were used to study the effect of altering the expression of this miRNA on time of flowering and to identify its mRNA target. Early or late flowering was observed in transgenic plants in which miR172 was overexpressed or suppressed, respectively. A full-length complementary DNA(cDNA) of gloxinia(Sinningia speciosa) APETALA2-like(SsAP2-like) was identified as a target of miR172. The altered expression levels of miR172 caused up-or down-regulation of SsAP2-like during flower development, which affected the time of flowering. Quantitative real-time reverse transcription PCR analysis of different gloxinia tissues revealed that the accumulation of SsAP2-like was negatively correlated with the expression of miR172 a, whereas the expression pattern of miR172 a was negatively correlated with that of miR156 a. Our results suggest that transgenic manipulation of miR172 could be used as a universal strategy for regulating time of flowering in ornamental plants.展开更多
BACKGROUND Hepatocellular carcinoma(HCC) is one of the most common malignant tumors with high mortality-to-incidence ratios. Nuclear factor erythroid 2-like 3(NFE2 L3), also known as NRF3, is a member of the cap ‘n&#...BACKGROUND Hepatocellular carcinoma(HCC) is one of the most common malignant tumors with high mortality-to-incidence ratios. Nuclear factor erythroid 2-like 3(NFE2 L3), also known as NRF3, is a member of the cap ‘n' collar basic-region leucine zipper family of transcription factors. NFE2 L3 is involved in the regulation of various biological processes, whereas its role in HCC has not been elucidated.AIM To explore the expression and biological function of NFE2 L3 in HCC.METHODS We analyzed the expression of NFE2 L3 in HCC tissues and its correlation with clinicopathological parameters based on The Cancer Genome Atlas(TCGA) data portal. Short hairpin RNA(shRNA) interference technology was utilized to knock down NFE2 L3 in vitro. Cell apoptosis, clone formation, proliferation, migration,and invasion assays were used to identify the biological effects of NFE2 L3 in BEL-7404 and SMMC-7721 cells. The expression of epithelial-mesenchymal transition(EMT) markers was examined by Western blot analysis.RESULTS TCGA analysis showed that NFE2 L3 expression was significantly positively correlated with tumor grade, T stage, and pathologic stage. The qPCR and Western blot results showed that both the mRNA and protein levels of NFE2 L3 were significantly decreased after shRNA-mediated knockdown in BEL-7404 and SMMC-7721 cells. The shRNA-mediated knockdown of NFE2 L3 could induce apoptosis and inhibit the clone formation and cell proliferation of SMMC-7721 and BEL-7404 cells. NFE2 L3 knockdown also significantly suppressed the migration, invasion, and EMT of the two cell lines.CONCLUSION Our study showed that shRNA-mediated knockdown of NFE2 L3 exhibited tumor-suppressing effects in HCC cells.展开更多
TDF1(transcription-drived fragment) was homologous to the predicted S. lycopersicum nonspecific lipid-transfer protein,nsLTP 2-like(91%), and it was significantly upregulated in response to C. fulvum(cladosporium fulv...TDF1(transcription-drived fragment) was homologous to the predicted S. lycopersicum nonspecific lipid-transfer protein,nsLTP 2-like(91%), and it was significantly upregulated in response to C. fulvum(cladosporium fulvum) infection in tomato plants.In this experiment, the full-length cDNA of nsLTP 2-like was cloned using RACE technology based on the sequence of TDF1(GenBank: JZ717725). A full-length, 625 bp(GenBank: KU366289), cDNA sequence, which with 98% similarity to nsLTP 2-like gene(GenBank: XM015233692) was obtained. This cDNA contains an ORF(open reading frame) with full-length of 345 bp, coding of 114 amino acids, including 12.3% Ala and Gly. Protein molecular weight was 11.51 ku, the isoelectric point(pI) was 8.99, and average overall hydrophilicity was 0.412, with one phosphorylation sites, belonging to volatile acidic nuclear protein. Secondary structure prediction showed that α-Helix accounts for 30.7%, extension chain for 12.28%, β-corner for 9.65%, and random coil for 47.37%. Through comparative analysis of the homology among species, it was found that the amino acid sequence of tomato nsLTP 2-like protein had a high similarity with other plants, and with a specific conserved sequence which might related features in nsLTP 2-like protein. It also be analyzed the gene expression pattern of tomato in different parts and under different stress conditions.The results showed that nsLTP 2-like gene was up-regulated in varying degrees, under the condition of cold stress, exogenous hormone spraying and cladosporium fulvum infection. Therefore, it was speculated that the gene played a role in response to abiotic and biotic stress in tomato.展开更多
The endosperm plays essential roles in embryogenesis and seed germination and provides abundant resources for human food and industrial products. Identification of genes regulating the development of the endosperm and...The endosperm plays essential roles in embryogenesis and seed germination and provides abundant resources for human food and industrial products. Identification of genes regulating the development of the endosperm and elucidation of their functions is of great importance for maize genetics and breeding. This study showed that the genespecific imprinted gene, ETHYLENE-INSENSITIVE 2-like(EIN2-like), is maternally expressed in both endosperm and embryo. The maternally expressed pattern was maintained throughout later seed developmental stages. Bisulfite sequencing using DNA obtained from hybrid endosperm tissues showed that the upstream regions of the alleles of EIN2-like were highly methylated at symmetrical sites(CG and CHG). A differentially methylated region in the upstream part of the maternal allele of EIN2-like was identified and found to be hypomethylated. Expression analysis showed that EIN2-like was highly expressed in the maize endosperm as well as at different stages of cell differentiation(8–12 days after pollination) in the hybrid endosperm. These results suggest that the maternally expressed gene EIN2-like may play crucial roles in the regulation of seed development.展开更多
BACKGROUND Hepatocellular carcinoma(HCC)is one of the most aggressive tumors worldwide.Chronic inflammation contributes to tumor evolution,and the infiltration of tumor-associated macrophages(TAMs),also known as M2-li...BACKGROUND Hepatocellular carcinoma(HCC)is one of the most aggressive tumors worldwide.Chronic inflammation contributes to tumor evolution,and the infiltration of tumor-associated macrophages(TAMs),also known as M2-like macrophages,is associated with the most aggressive behavior.Therefore,these macrophages provide the primary growth and migratory factors to the tumor cells,including those of HCC.Current therapies are not well optimized for eliminating trans-formed cells or neutralizing the tumor immune microenvironment leukocytes,such as TAMs.Growth differentiation factor 11(GDF11)may represent a promi-sing dual therapeutic target due to its reported anti-tumorigenic and immuno-modulatory properties.AIM To characterize the effects of GDF11 in M2-like macrophages and the HCC cell interaction using a functional in vitro model.METHODS This research used THP-1 and Huh7 cell lines.We applied recombinant GDF11(50 ng/mL)every 24 hours on THP-1 differentiated macrophages with M2-like polarization using interleukin-4 and interleukin-13.Firstly,the GDF11 effects on signaling,viability,proliferation,metabolism,and redox state in macrophages were charac-terized.Subsequently,we extracted conditioned media(CM)from macrophages and performed indirect co-cultures with Huh7 cells.The functional parameters were proliferation and migration assays.Finally,we charac-terized secretion in the CM using the cytokine array membrane assay.RESULTS The present study demonstrated that GDF11 activates the canonical pathway Smad2/3 without cytotoxic or prolif-erative effects.We provide evidence that GDF11 also diminishes the pro-tumoral properties of M2-like macrophages.GDF11 promoted the reduction of the M2-like macrophage marker,cluster of differentiation 206,indicating a loss of pro-tumoral properties in these leukocytes.Furthermore,this molecule induced changes in metabolism and an increase in reactive oxygen species.Using CM derived from GDF11-treated M2-like macrophages,we observed a reduction in the proliferation and migratory capacity of liver cancer cells.Moreover,the cytokine profile was affected by GDF11 stimulus,demonstrating that this molecule alters the pro-tumoral properties of TAMs,which in turn impact the behavior of HCC-derived cells.CONCLUSION This in vitro study suggests that mitigating tumor-promoting or M2-like macrophages with GDF11 may be an effective strategy for controlling the aggressiveness of HCC.展开更多
基金Project supported by the National Natural Science Foundation of China(Nos.31171615 and 31401913)
文摘We investigated the microRNA172(miR172)-mediated regulatory network for the perception of changes in external and endogenous signals to identify a universally applicable floral regulation system in ornamental plants, manipulation of which could be economically beneficial. Transgenic gloxinia plants, in which miR172 was either overexpressed or suppressed, were generated using Agrobacterium-mediated transformation. They were used to study the effect of altering the expression of this miRNA on time of flowering and to identify its mRNA target. Early or late flowering was observed in transgenic plants in which miR172 was overexpressed or suppressed, respectively. A full-length complementary DNA(cDNA) of gloxinia(Sinningia speciosa) APETALA2-like(SsAP2-like) was identified as a target of miR172. The altered expression levels of miR172 caused up-or down-regulation of SsAP2-like during flower development, which affected the time of flowering. Quantitative real-time reverse transcription PCR analysis of different gloxinia tissues revealed that the accumulation of SsAP2-like was negatively correlated with the expression of miR172 a, whereas the expression pattern of miR172 a was negatively correlated with that of miR156 a. Our results suggest that transgenic manipulation of miR172 could be used as a universal strategy for regulating time of flowering in ornamental plants.
基金the Changzhou High-Level Medical Talents Training Project,No.2016ZCLJ002
文摘BACKGROUND Hepatocellular carcinoma(HCC) is one of the most common malignant tumors with high mortality-to-incidence ratios. Nuclear factor erythroid 2-like 3(NFE2 L3), also known as NRF3, is a member of the cap ‘n' collar basic-region leucine zipper family of transcription factors. NFE2 L3 is involved in the regulation of various biological processes, whereas its role in HCC has not been elucidated.AIM To explore the expression and biological function of NFE2 L3 in HCC.METHODS We analyzed the expression of NFE2 L3 in HCC tissues and its correlation with clinicopathological parameters based on The Cancer Genome Atlas(TCGA) data portal. Short hairpin RNA(shRNA) interference technology was utilized to knock down NFE2 L3 in vitro. Cell apoptosis, clone formation, proliferation, migration,and invasion assays were used to identify the biological effects of NFE2 L3 in BEL-7404 and SMMC-7721 cells. The expression of epithelial-mesenchymal transition(EMT) markers was examined by Western blot analysis.RESULTS TCGA analysis showed that NFE2 L3 expression was significantly positively correlated with tumor grade, T stage, and pathologic stage. The qPCR and Western blot results showed that both the mRNA and protein levels of NFE2 L3 were significantly decreased after shRNA-mediated knockdown in BEL-7404 and SMMC-7721 cells. The shRNA-mediated knockdown of NFE2 L3 could induce apoptosis and inhibit the clone formation and cell proliferation of SMMC-7721 and BEL-7404 cells. NFE2 L3 knockdown also significantly suppressed the migration, invasion, and EMT of the two cell lines.CONCLUSION Our study showed that shRNA-mediated knockdown of NFE2 L3 exhibited tumor-suppressing effects in HCC cells.
基金Supported by the National Key R&D Program of China(2017YFD0101900)China Agriculture Research System(CARS-23-A-16)the Science Foundation of Heilongjiang Province(C2017024)
文摘TDF1(transcription-drived fragment) was homologous to the predicted S. lycopersicum nonspecific lipid-transfer protein,nsLTP 2-like(91%), and it was significantly upregulated in response to C. fulvum(cladosporium fulvum) infection in tomato plants.In this experiment, the full-length cDNA of nsLTP 2-like was cloned using RACE technology based on the sequence of TDF1(GenBank: JZ717725). A full-length, 625 bp(GenBank: KU366289), cDNA sequence, which with 98% similarity to nsLTP 2-like gene(GenBank: XM015233692) was obtained. This cDNA contains an ORF(open reading frame) with full-length of 345 bp, coding of 114 amino acids, including 12.3% Ala and Gly. Protein molecular weight was 11.51 ku, the isoelectric point(pI) was 8.99, and average overall hydrophilicity was 0.412, with one phosphorylation sites, belonging to volatile acidic nuclear protein. Secondary structure prediction showed that α-Helix accounts for 30.7%, extension chain for 12.28%, β-corner for 9.65%, and random coil for 47.37%. Through comparative analysis of the homology among species, it was found that the amino acid sequence of tomato nsLTP 2-like protein had a high similarity with other plants, and with a specific conserved sequence which might related features in nsLTP 2-like protein. It also be analyzed the gene expression pattern of tomato in different parts and under different stress conditions.The results showed that nsLTP 2-like gene was up-regulated in varying degrees, under the condition of cold stress, exogenous hormone spraying and cladosporium fulvum infection. Therefore, it was speculated that the gene played a role in response to abiotic and biotic stress in tomato.
基金the Major Research Projects of Chongqing (CSTC2016shms-ztzx80013, CSTC2016shms-ztzx80016) for financial support
文摘The endosperm plays essential roles in embryogenesis and seed germination and provides abundant resources for human food and industrial products. Identification of genes regulating the development of the endosperm and elucidation of their functions is of great importance for maize genetics and breeding. This study showed that the genespecific imprinted gene, ETHYLENE-INSENSITIVE 2-like(EIN2-like), is maternally expressed in both endosperm and embryo. The maternally expressed pattern was maintained throughout later seed developmental stages. Bisulfite sequencing using DNA obtained from hybrid endosperm tissues showed that the upstream regions of the alleles of EIN2-like were highly methylated at symmetrical sites(CG and CHG). A differentially methylated region in the upstream part of the maternal allele of EIN2-like was identified and found to be hypomethylated. Expression analysis showed that EIN2-like was highly expressed in the maize endosperm as well as at different stages of cell differentiation(8–12 days after pollination) in the hybrid endosperm. These results suggest that the maternally expressed gene EIN2-like may play crucial roles in the regulation of seed development.
文摘BACKGROUND Hepatocellular carcinoma(HCC)is one of the most aggressive tumors worldwide.Chronic inflammation contributes to tumor evolution,and the infiltration of tumor-associated macrophages(TAMs),also known as M2-like macrophages,is associated with the most aggressive behavior.Therefore,these macrophages provide the primary growth and migratory factors to the tumor cells,including those of HCC.Current therapies are not well optimized for eliminating trans-formed cells or neutralizing the tumor immune microenvironment leukocytes,such as TAMs.Growth differentiation factor 11(GDF11)may represent a promi-sing dual therapeutic target due to its reported anti-tumorigenic and immuno-modulatory properties.AIM To characterize the effects of GDF11 in M2-like macrophages and the HCC cell interaction using a functional in vitro model.METHODS This research used THP-1 and Huh7 cell lines.We applied recombinant GDF11(50 ng/mL)every 24 hours on THP-1 differentiated macrophages with M2-like polarization using interleukin-4 and interleukin-13.Firstly,the GDF11 effects on signaling,viability,proliferation,metabolism,and redox state in macrophages were charac-terized.Subsequently,we extracted conditioned media(CM)from macrophages and performed indirect co-cultures with Huh7 cells.The functional parameters were proliferation and migration assays.Finally,we charac-terized secretion in the CM using the cytokine array membrane assay.RESULTS The present study demonstrated that GDF11 activates the canonical pathway Smad2/3 without cytotoxic or prolif-erative effects.We provide evidence that GDF11 also diminishes the pro-tumoral properties of M2-like macrophages.GDF11 promoted the reduction of the M2-like macrophage marker,cluster of differentiation 206,indicating a loss of pro-tumoral properties in these leukocytes.Furthermore,this molecule induced changes in metabolism and an increase in reactive oxygen species.Using CM derived from GDF11-treated M2-like macrophages,we observed a reduction in the proliferation and migratory capacity of liver cancer cells.Moreover,the cytokine profile was affected by GDF11 stimulus,demonstrating that this molecule alters the pro-tumoral properties of TAMs,which in turn impact the behavior of HCC-derived cells.CONCLUSION This in vitro study suggests that mitigating tumor-promoting or M2-like macrophages with GDF11 may be an effective strategy for controlling the aggressiveness of HCC.
