The rabbit is well known for producing diverse antibodies against various antigens including small molecules such as drugs and toxins,due to a robust immune response.Elucidating how hapten repeated immunization shapes...The rabbit is well known for producing diverse antibodies against various antigens including small molecules such as drugs and toxins,due to a robust immune response.Elucidating how hapten repeated immunization shapes the rabbit B cell receptor(BCR)repertoire is crucial to understanding rabbit immune response to small molecules and assisting rare antibody discovery/engineering.In this study,we enriched and sequenced chloramphenicol(CAP)-specific rabbit B cells following repeated immunization,and analyzed both CAP-specific repertoires combined with the structure and affinity features of V1S69/V1S37 germline-based BCRs.The length of rabbit complementarity-determining region 3 of heavy chain(CDRH3)increased after hapten immunization.Repeated immunization significantly reduced the diversity of CAP-specific rabbit BCR clonotypes,and changed the frequency of VDJ usage and the type of V(D)J recombination.The average number of mutations among VL is notably higher than that of VH genes in rabbits,however,they are both not changed along with repeated immunization.Moreover,repeated immunization resulted in an increase surface charge and a decrease in solvent accessible surface area,leading to improvement in the stability of the most abundant V1S69/V1S37 germline-based BCR,along with an affinity increase from an IC50 of 898.2 ng mL^(−1)at the 1st immunization to 4.16 ng mL^(−1)at the 6th immunization.The study provides a benchmark for rabbit repertoire-scale analyses and offers a method for antibody discovery of small molecules.展开更多
The characterization of the human T-cell receptor （TCR） repertoire has made remarkable progress, with most of the work focusing on the TCRβ chains. Here, we ana- lyzed the diversity and complexity of both the TCRa ...The characterization of the human T-cell receptor （TCR） repertoire has made remarkable progress, with most of the work focusing on the TCRβ chains. Here, we ana- lyzed the diversity and complexity of both the TCRa and TCRβ repertoires of three healthy donors. We found that the diversity of the TCRα repertoire is higher than that of the TCRβ repertoire, whereas the usages of the V and J genes tended to be preferential with similar TRAV and TRAJ patterns in all three donors. The V-J pairings, like the V and J gene usages, were slightly preferential. We also found that the TRDV1 gene rearranges with the majority of TRAJ genes, suggesting that TRDV1 is a shared TRAV/DV gene （TRAV42/DV1）. Moreover, we uncovered the presence of tandem TRBD （TRB D gene） usage in -2% of the productive human TCRβ CDR3 sequences.展开更多
T cells and T cell receptors(TCRs)play pivotal roles in adaptive immune responses against tumors.The development of next-generation sequencing technologies has enabled the analysis of the TCRb repertoire usage.Given t...T cells and T cell receptors(TCRs)play pivotal roles in adaptive immune responses against tumors.The development of next-generation sequencing technologies has enabled the analysis of the TCRb repertoire usage.Given the scarce investigations on the TCR repertoire in lung cancer tissues,in this study,we analyzed TCRb repertoires in lung cancer tissues and the matched distant non-tumor lung tissues(normal lung tissues)from 15 lung cancer patients.Based on our results,the general distribution of T cell clones was similar between cancer tissues and normal lung tissues;however,the proportion of highly expanded clones was significantly higher in normal lung tissues than in cancer tissues(0.021%±0.002%vs.0.016%±0.001%,P=0.0054,Wilcoxon signed rank test).In addition,a significantly higher TCR diversity was observed in cancer tissues than in normal lung tissues(431.37±305.96 vs.166.20±101.58,P=0.0075,Mann-Whitney U test).Moreover,younger patients had a significantly higher TCR diversity than older patients(640.7±295.3 vs.291.8±233.6,P=0.036,Mann-Whitney U test),and the higher TCR diversity in tumors was significantly associated with worse cancer outcomes.Thus,we provided a comprehensive comparison of the TCR repertoires between cancer tissues and matched normal lung tissues and demonstrated the presence of distinct T cell immune microenvironments in lung cancer patients.展开更多
基金approved by the Animal Ethics Committee of China Agricultural University and strictly conducted by Chinese laws and guidelines(AW32602202-2-2).
文摘The rabbit is well known for producing diverse antibodies against various antigens including small molecules such as drugs and toxins,due to a robust immune response.Elucidating how hapten repeated immunization shapes the rabbit B cell receptor(BCR)repertoire is crucial to understanding rabbit immune response to small molecules and assisting rare antibody discovery/engineering.In this study,we enriched and sequenced chloramphenicol(CAP)-specific rabbit B cells following repeated immunization,and analyzed both CAP-specific repertoires combined with the structure and affinity features of V1S69/V1S37 germline-based BCRs.The length of rabbit complementarity-determining region 3 of heavy chain(CDRH3)increased after hapten immunization.Repeated immunization significantly reduced the diversity of CAP-specific rabbit BCR clonotypes,and changed the frequency of VDJ usage and the type of V(D)J recombination.The average number of mutations among VL is notably higher than that of VH genes in rabbits,however,they are both not changed along with repeated immunization.Moreover,repeated immunization resulted in an increase surface charge and a decrease in solvent accessible surface area,leading to improvement in the stability of the most abundant V1S69/V1S37 germline-based BCR,along with an affinity increase from an IC50 of 898.2 ng mL^(−1)at the 1st immunization to 4.16 ng mL^(−1)at the 6th immunization.The study provides a benchmark for rabbit repertoire-scale analyses and offers a method for antibody discovery of small molecules.
基金We thank Dr. Christopher J. Vavrickafor and Boris Tefsen for their critical reading and revision of the manuscript and Dr. Miles P. Dav- enport for his inspiring discussions. This work is supported by the National Natural Science Foundation of China （NSFC, Grant No. 31030030）, the National Basic Research Program （973 Program） （No. 2013CB531500） and the National Natural Science Foundation of China （Grant No. 81373141 ）. G.F.G. is a leading principal investigator of the NSFC Innovative Research Group （Grant No. 81321063）.
文摘The characterization of the human T-cell receptor （TCR） repertoire has made remarkable progress, with most of the work focusing on the TCRβ chains. Here, we ana- lyzed the diversity and complexity of both the TCRa and TCRβ repertoires of three healthy donors. We found that the diversity of the TCRα repertoire is higher than that of the TCRβ repertoire, whereas the usages of the V and J genes tended to be preferential with similar TRAV and TRAJ patterns in all three donors. The V-J pairings, like the V and J gene usages, were slightly preferential. We also found that the TRDV1 gene rearranges with the majority of TRAJ genes, suggesting that TRDV1 is a shared TRAV/DV gene （TRAV42/DV1）. Moreover, we uncovered the presence of tandem TRBD （TRB D gene） usage in -2% of the productive human TCRβ CDR3 sequences.
基金supported by the CAMS Innovation Fund for Medical Sciences(CIFMS),China(Grant No.2016-I2M-1-001)the National Key Research and Development Program of China(Grant No.2017YFC0908401)the National Basic Research Program of China(973 Program,Grant No.2014CBA02004)
文摘T cells and T cell receptors(TCRs)play pivotal roles in adaptive immune responses against tumors.The development of next-generation sequencing technologies has enabled the analysis of the TCRb repertoire usage.Given the scarce investigations on the TCR repertoire in lung cancer tissues,in this study,we analyzed TCRb repertoires in lung cancer tissues and the matched distant non-tumor lung tissues(normal lung tissues)from 15 lung cancer patients.Based on our results,the general distribution of T cell clones was similar between cancer tissues and normal lung tissues;however,the proportion of highly expanded clones was significantly higher in normal lung tissues than in cancer tissues(0.021%±0.002%vs.0.016%±0.001%,P=0.0054,Wilcoxon signed rank test).In addition,a significantly higher TCR diversity was observed in cancer tissues than in normal lung tissues(431.37±305.96 vs.166.20±101.58,P=0.0075,Mann-Whitney U test).Moreover,younger patients had a significantly higher TCR diversity than older patients(640.7±295.3 vs.291.8±233.6,P=0.036,Mann-Whitney U test),and the higher TCR diversity in tumors was significantly associated with worse cancer outcomes.Thus,we provided a comprehensive comparison of the TCR repertoires between cancer tissues and matched normal lung tissues and demonstrated the presence of distinct T cell immune microenvironments in lung cancer patients.
