Lassa virus(LASV)belongs to the Mammarenavirus genus(family Arenaviridae)and causes severe hemorrhagic fever in humans.The glycoprotein complex(GPC)contains eleven N-linked glycans that play essential roles in GPC fun...Lassa virus(LASV)belongs to the Mammarenavirus genus(family Arenaviridae)and causes severe hemorrhagic fever in humans.The glycoprotein complex(GPC)contains eleven N-linked glycans that play essential roles in GPC functionalities such as cleavage,transport,receptor recognition,epitope shielding,and immune response.We used three mutagenesis strategies(asparagine to glutamine,asparagine to alanine,and serine/tyrosine to alanine mutants)to abolish individual glycan chain on GPC and found that all the three strategies led to cleavage inefficiency on the 2nd(N89),5th(N119),or 8th(N365)glycosylation motif.To evaluate N to Q mutagenesis for further research,it was found that deletion of the 2nd(N89Q)or 8th(N365Q)glycan completely inhibited the transduction efficiency of pseudotyped particles.We further investigated the role of individual glycan on GPC-mediated immune response by DNA immunization of mice.Deletion of the individual 1st(N79Q),3rd(N99Q),5th(N119Q),or 6th(N167Q)glycan significantly enhanced the proportion of effector CD4+cells,whereas deletion of the 1st(N79Q),2nd(N89Q),3rd(N99Q),4th(N109Q),5th(N119Q),6th(N167Q),or 9th(N373Q)glycan enhanced the proportion of CD8+effector T cells.Deletion of specific glycan improves the Th1-type immune response,and abolishment of glycan on GPC generally increases the antibody titer to the glycan-deficient GPC.However,the antibodies from either the mutant or WT GPC-immunized mice show little neutralization effect on wild-type LASV.The glycan residues on GPC provide an immune shield for the virus,and thus represent a target for the design and development of a vaccine.展开更多
Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments gro...Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments groups with 21 replicates,13 birds per replicate.The present research investigated effects of microbial muramidase or a precision glycan alone or in combination on growth performance,apparent total tract digestibility,total blood carotenoid content,intestinal villus length,meat quality and gut microbiota in broiler chickens.Treatments included:NC:negative control(basal diet group);PC:positive control(basal diet+0.02%probiotics);MR:basal diet+0.035%microbial muramidase;PG:basal diet+0.1%precision glycan;and MRPG:basal diet+0.025%MR+0.1%PG,respectively.Results MRPG group increased the body weight gain and feed intake(P<0.05)compared with NC group.Moreover,it significantly increased total serum carotenoid(P<0.05)and MRPG altered the microbial diversity in ileum contents.The MRPG treatment group increased the abundance of the phylum Firmicutes,and family Lachnospiraceae,Ruminococcaceae,Oscillospiraceae,Lactobacillaceae,Peptostreptococcaceae and decreased the abundance of the phylum Campilobacterota,Bacteroidota and family Bacteroidaceae.Compared with the NC group,the chickens fed MRPG showed significantly increased in duodenum villus length at end the trial.Conclusion In this study,overall results showed that the synergetic effects of MR and PG showed enhancing growth performance,total serum carotenoid level and altering gut microbiota composition of broilers.The current research indicates that co-supplementation of MR and PG in broiler diets enhances intestinal health,consequently leading to an increased broiler production.展开更多
Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone（THAP）and 2,5-dihydroxybenzoic acid（DHB）as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass...Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone（THAP）and 2,5-dihydroxybenzoic acid（DHB）as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer（MALDI-TOF MS）.High quality negative-ion spectra of commercial sialylated glycan were obtained with THAP as matrix.Detection limit of the glycan was less than 0.1 pmol.After methyl esterification of sialic acid（SA）residue,sialylated glycans were detected sensitively in the positive-ion mode using DHB as matrix.Neutral and sialylated glycans from the mixture of asialofetuin and fetuin were methylesterified and simultaneously recognized in one manipulation.Methyl esterification of SA residue offers a convenient and sensitive way to identify the structure of N-linked glycans for glycan profiling.展开更多
BACKGROUND Mass spectrometry-based proteomics and glycomics reveal post-translational modifications providing significant biological insights beyond the scope of genomic sequencing.AIM To characterize the N-linked gly...BACKGROUND Mass spectrometry-based proteomics and glycomics reveal post-translational modifications providing significant biological insights beyond the scope of genomic sequencing.AIM To characterize the N-linked glycoproteomic profile in esophageal squamous cell carcinoma(ESCC)via two complementary approaches.METHODS Using tandem multilectin affinity chromatography for enrichment of N-linked glycoproteins,we performed N-linked glycoproteomic profiling in ESCC tissues by two-dimensional gel electrophoresis(2-DE)-based and isobaric tags for relative and absolute quantification(iTRAQ)labeling-based mass spectrometry quantitation in parallel,followed by validation of candidate glycoprotein biomarkers by Western blot.RESULTS 2-DE-based and iTRAQ labeling-based quantitation identified 24 and 402 differentially expressed N-linked glycoproteins,respectively,with 15 in common,demonstrating the outperformance of iTRAQ labeling-based quantitation over 2-DE and complementarity of these two approaches.Proteomaps showed the distinct compositions of functional categories between proteins and glycoproteins with differential expression associated with ESCC.Western blot analysis validated the up-regulation of total procathepsin D and high-mannose procathepsin D,and the downregulation of total haptoglobin,high-mannose clusterin,and GlcNAc/sialic acid-containing fraction of 14-3-3ζ in ESCC tissues.The serum levels of glycosylated fractions of clusterin,prolinearginine-rich end leucine-rich repeat protein,and haptoglobin in patients with ESCC were remarkably higher than those in healthy controls.CONCLUSION Our study provides insights into the aberrant N-linked glycoproteome associated with ESCC,which will be a valuable resource for future investigations.展开更多
Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Rece...Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Recently,immunoregulatory molecule-based cancer immunotherapy has greatly improved treatment efficacy for solid cancers,thus enabling some patients to achieve persistent responses or cure.However,owing to the development of drug-resistance and the low response rate,immunotherapy against the available targets PD-1/PD-L1 or CTLA-4 has limited benefits.Although combination therapies have been proposed to enhance the response rate,severe adverse effects are observed.Thus,alternative immune checkpoints must be identified.The SIGLECs are a family of immunoregulatory receptors(known as glyco-immune checkpoints)discovered in recent years.This review systematically describes the molecular characteristics of the SIGLECs,and discusses recent progress in areas including synthetic ligands,monoclonal antibody inhibitors,and Chimeric antigen receptor T(CAR-T)cells,with a focus on available strategies for blocking the sialylated glycan-SIGLEC axis.Targeting glyco-immune checkpoints can expand the scope of immune checkpoints and provide multiple options for new drug development.展开更多
基金This work was supported by the National Key Research and Development Program of China(2018YFA0507204)the National Natural Sciences Foundation of China(31670165)+1 种基金Wuhan National Biosafety Laboratory,Chinese Academy of Sciences Advanced Customer Cultivation Project(2019ACCP-MS03)the Open Research Fund Program of the State Key Laboratory of Virology of China(2018IOV001).Author Contributions。
文摘Lassa virus(LASV)belongs to the Mammarenavirus genus(family Arenaviridae)and causes severe hemorrhagic fever in humans.The glycoprotein complex(GPC)contains eleven N-linked glycans that play essential roles in GPC functionalities such as cleavage,transport,receptor recognition,epitope shielding,and immune response.We used three mutagenesis strategies(asparagine to glutamine,asparagine to alanine,and serine/tyrosine to alanine mutants)to abolish individual glycan chain on GPC and found that all the three strategies led to cleavage inefficiency on the 2nd(N89),5th(N119),or 8th(N365)glycosylation motif.To evaluate N to Q mutagenesis for further research,it was found that deletion of the 2nd(N89Q)or 8th(N365Q)glycan completely inhibited the transduction efficiency of pseudotyped particles.We further investigated the role of individual glycan on GPC-mediated immune response by DNA immunization of mice.Deletion of the individual 1st(N79Q),3rd(N99Q),5th(N119Q),or 6th(N167Q)glycan significantly enhanced the proportion of effector CD4+cells,whereas deletion of the 1st(N79Q),2nd(N89Q),3rd(N99Q),4th(N109Q),5th(N119Q),6th(N167Q),or 9th(N373Q)glycan enhanced the proportion of CD8+effector T cells.Deletion of specific glycan improves the Th1-type immune response,and abolishment of glycan on GPC generally increases the antibody titer to the glycan-deficient GPC.However,the antibodies from either the mutant or WT GPC-immunized mice show little neutralization effect on wild-type LASV.The glycan residues on GPC provide an immune shield for the virus,and thus represent a target for the design and development of a vaccine.
基金supported by Basic Science Research Program through the National Research Foundation of Korea(NRF)funded by the Ministry of Education(NRF-RS-2023-00275307)。
文摘Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments groups with 21 replicates,13 birds per replicate.The present research investigated effects of microbial muramidase or a precision glycan alone or in combination on growth performance,apparent total tract digestibility,total blood carotenoid content,intestinal villus length,meat quality and gut microbiota in broiler chickens.Treatments included:NC:negative control(basal diet group);PC:positive control(basal diet+0.02%probiotics);MR:basal diet+0.035%microbial muramidase;PG:basal diet+0.1%precision glycan;and MRPG:basal diet+0.025%MR+0.1%PG,respectively.Results MRPG group increased the body weight gain and feed intake(P<0.05)compared with NC group.Moreover,it significantly increased total serum carotenoid(P<0.05)and MRPG altered the microbial diversity in ileum contents.The MRPG treatment group increased the abundance of the phylum Firmicutes,and family Lachnospiraceae,Ruminococcaceae,Oscillospiraceae,Lactobacillaceae,Peptostreptococcaceae and decreased the abundance of the phylum Campilobacterota,Bacteroidota and family Bacteroidaceae.Compared with the NC group,the chickens fed MRPG showed significantly increased in duodenum villus length at end the trial.Conclusion In this study,overall results showed that the synergetic effects of MR and PG showed enhancing growth performance,total serum carotenoid level and altering gut microbiota composition of broilers.The current research indicates that co-supplementation of MR and PG in broiler diets enhances intestinal health,consequently leading to an increased broiler production.
基金Supported by the National Natural Science Foundation of China(30800193)Grant from Centre for International Mobility(CIMO),Finland
文摘Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone（THAP）and 2,5-dihydroxybenzoic acid（DHB）as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer（MALDI-TOF MS）.High quality negative-ion spectra of commercial sialylated glycan were obtained with THAP as matrix.Detection limit of the glycan was less than 0.1 pmol.After methyl esterification of sialic acid（SA）residue,sialylated glycans were detected sensitively in the positive-ion mode using DHB as matrix.Neutral and sialylated glycans from the mixture of asialofetuin and fetuin were methylesterified and simultaneously recognized in one manipulation.Methyl esterification of SA residue offers a convenient and sensitive way to identify the structure of N-linked glycans for glycan profiling.
基金Supported by National Natural Science Foundation of China,No.81072039 and No.81872037.
文摘BACKGROUND Mass spectrometry-based proteomics and glycomics reveal post-translational modifications providing significant biological insights beyond the scope of genomic sequencing.AIM To characterize the N-linked glycoproteomic profile in esophageal squamous cell carcinoma(ESCC)via two complementary approaches.METHODS Using tandem multilectin affinity chromatography for enrichment of N-linked glycoproteins,we performed N-linked glycoproteomic profiling in ESCC tissues by two-dimensional gel electrophoresis(2-DE)-based and isobaric tags for relative and absolute quantification(iTRAQ)labeling-based mass spectrometry quantitation in parallel,followed by validation of candidate glycoprotein biomarkers by Western blot.RESULTS 2-DE-based and iTRAQ labeling-based quantitation identified 24 and 402 differentially expressed N-linked glycoproteins,respectively,with 15 in common,demonstrating the outperformance of iTRAQ labeling-based quantitation over 2-DE and complementarity of these two approaches.Proteomaps showed the distinct compositions of functional categories between proteins and glycoproteins with differential expression associated with ESCC.Western blot analysis validated the up-regulation of total procathepsin D and high-mannose procathepsin D,and the downregulation of total haptoglobin,high-mannose clusterin,and GlcNAc/sialic acid-containing fraction of 14-3-3ζ in ESCC tissues.The serum levels of glycosylated fractions of clusterin,prolinearginine-rich end leucine-rich repeat protein,and haptoglobin in patients with ESCC were remarkably higher than those in healthy controls.CONCLUSION Our study provides insights into the aberrant N-linked glycoproteome associated with ESCC,which will be a valuable resource for future investigations.
基金supported by the Shanghai Science and Technology Committee (Grant Nos. 20DZ2201900 to Y.Y. and 23ZR1432500 to W.P.)National Natural Science Foundation of China (Grant Nos. 82072602 to Y.Y.+4 种基金91853121, 21977066, and 22177069 to W.P.)Innovation Foundation of Translational Medicine of Shanghai Jiao Tong University School of Medicine(Grant No. TM202001 to Y.Y.)Collaborative Innovation Center for Clinical and Translational Science by Chinese Ministry of Education&Shanghai (Grant No. CCTS-2022202 to Y.Y.)Shanghai Pilot Program for Basic Research-Shanghai Jiao Tong University (Grant No. 21TQ1400210 to W.P.)Medical-Engineering Interdisciplinary Research Foundation of Shanghai Jiao Tong University (Grant No. YG2022ZD001 to W.P.)
文摘Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Recently,immunoregulatory molecule-based cancer immunotherapy has greatly improved treatment efficacy for solid cancers,thus enabling some patients to achieve persistent responses or cure.However,owing to the development of drug-resistance and the low response rate,immunotherapy against the available targets PD-1/PD-L1 or CTLA-4 has limited benefits.Although combination therapies have been proposed to enhance the response rate,severe adverse effects are observed.Thus,alternative immune checkpoints must be identified.The SIGLECs are a family of immunoregulatory receptors(known as glyco-immune checkpoints)discovered in recent years.This review systematically describes the molecular characteristics of the SIGLECs,and discusses recent progress in areas including synthetic ligands,monoclonal antibody inhibitors,and Chimeric antigen receptor T(CAR-T)cells,with a focus on available strategies for blocking the sialylated glycan-SIGLEC axis.Targeting glyco-immune checkpoints can expand the scope of immune checkpoints and provide multiple options for new drug development.
