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Characterization of CD8+ T cell differentiation following SIVΔnef vaccination by transcription factor expression profiling.

Billingsley JM, Rajakumar PA, Connole MA, Salisch NC, Adnan S, Kuzmichev YV, Hong HS, Reeves RK, Kang HJ, Li W, Li Q, Haase AT, Johnson RP - PLoS Pathog. (2015)

Bottom Line: The onset of protective immunity against pathogenic SIV challenge in SIVΔnef-vaccinated macaques is delayed for 15-20 weeks, a process that is related to qualitative changes in CD8+ T cell responses induced by SIVΔnef.Expression of transcription factors associated with effector differentiation, such as ID2 and RUNX3, were decreased over time, while expression of transcription factors associated with quiescence or memory differentiation, such as TCF7, BCOR and EOMES, increased.These data suggest transcription factor expression profiling is a novel method that can provide additional data complementary to the analysis of memory cell differentiation based on classical phenotypic markers.

View Article: PubMed Central - PubMed

Affiliation: Division of Immunology, New England Primate Research Center, Harvard Medical School, Southborough, Massachusetts, United States of America; Yerkes National Primate Research Center, Emory University, Atlanta, Georgia, United States of America.

ABSTRACT
The onset of protective immunity against pathogenic SIV challenge in SIVΔnef-vaccinated macaques is delayed for 15-20 weeks, a process that is related to qualitative changes in CD8+ T cell responses induced by SIVΔnef. As a novel approach to characterize cell differentiation following vaccination, we used multi-target qPCR to measure transcription factor expression in naïve and memory subsets of CD8++ T cells, and in SIV-specific CD8+ T cells obtained from SIVΔnef-vaccinated or wild type SIVmac239-infected macaques. Unsupervised clustering of expression profiles organized naïve and memory CD8+ T cells into groups concordant with cell surface phenotype. Transcription factor expression patterns in SIV-specific CD8+ T cells in SIVΔnef-vaccinated animals were distinct from those observed in purified CD8+ T cell subsets obtained from naïve animals, and were intermediate to expression profiles of purified central memory and effector memory T cells. Expression of transcription factors elicited by SIVΔnef vaccination also varied over time: cells obtained at later time points, temporally associated with greater protection, appeared more central-memory like than cells obtained at earlier time points, which appeared more effector memory-like. Expression of transcription factors associated with effector differentiation, such as ID2 and RUNX3, were decreased over time, while expression of transcription factors associated with quiescence or memory differentiation, such as TCF7, BCOR and EOMES, increased. CD8+ T cells specific for a more conserved epitope expressed higher levels of TBX21 and BATF, and appeared more effector-like than cells specific for an escaped epitope, consistent with continued activation by replicating vaccine virus. These data suggest transcription factor expression profiling is a novel method that can provide additional data complementary to the analysis of memory cell differentiation based on classical phenotypic markers. Additionally, these data support the hypothesis that ongoing stimulation by SIVΔnef promotes a distinct protective balance of CD8+ T cell differentiation and activation states.

No MeSH data available.


Related in: MedlinePlus

Principal component analysis of transcription factor expression profiles from SIV-specific MHC tetramer-sorted CD8+ T cells and sorted CD8+ T cell subsets.(A) Plot of principal components 1 vs. 2, and 2 vs. 3 for each of the expression profiles assessed in sorted naïve and memory CD8+ T cell subsets isolated from healthy control animals (n = 5), and SIV-specific MHC tetramer-sorted CD8+ T cells isolated from animals (n = 4) at week 5 or week 20 following SIVΔnef vaccination. Principal components 1, 2 and 3 explain 92% of cumulative total variance. (B) PCA loading factors for each transcription factor.
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ppat.1004740.g003: Principal component analysis of transcription factor expression profiles from SIV-specific MHC tetramer-sorted CD8+ T cells and sorted CD8+ T cell subsets.(A) Plot of principal components 1 vs. 2, and 2 vs. 3 for each of the expression profiles assessed in sorted naïve and memory CD8+ T cell subsets isolated from healthy control animals (n = 5), and SIV-specific MHC tetramer-sorted CD8+ T cells isolated from animals (n = 4) at week 5 or week 20 following SIVΔnef vaccination. Principal components 1, 2 and 3 explain 92% of cumulative total variance. (B) PCA loading factors for each transcription factor.

Mentions: To integrate the expression profiles of the SIV-specific cells with the sorted CD8+ subsets, we applied principal component analysis (PCA) to the combined data sets. Plotting principal components 1 vs, 2, and principal components 2 vs. 3, (PC1, PC2, PC3; Fig. 3A, S1 Video) segregated the data into distinct clusters. The data points representing the sorted CD8+ T cells occupy the periphery of the PC1 vs. PC2 plot, and segregate into separate clusters based upon cell differentiation stage. The naïve cells segregate from the memory cells along the PC1 axis, whereas the memory cells segregate along the PC2 axis, with the transitional memory cells positioned intermediately between the central and effector cells. The PC1 and PC2 loading factors (Fig. 3B) indicate that in this analysis, differential expression of LEF1, TCF7, PRDM1 and TBX21 strongly influence segregation of naïve from memory cells, whereas differential expression of ID2, RUNX3, AHR and LEF1 strongly influence segregation of memory cell subsets. The SIV-specific CD8+ T cells cluster with the sorted memory cells on the PC1 axis, and are positioned intermediately between central memory and effector memory cells on the PC2 axis. This intermediate position on the PC2 axis in part reflects the combined expression profiles of different memory subsets present in the SIV-specific cell samples. However, the SIV-specific samples significantly segregate from any sorted memory subset, particularly on the PC3 axis (p<0.001), indicating that the transcription factor expression profiles of the SIV-specific cells are distinct from the sorted subsets and are not solely comprised of proportions of memory subsets. The PC3 loading factors (Fig. 3B) indicate that in this analysis, the differential expression of NFIL3, IRF4, LEF1 and EOMES influence segregation of SIV-specific cells from the sorted naïve and memory subsets. The SIV-specific cells form two clusters, generally organized by week post-infection. The week 5 and week 20 post-vaccination samples occupy significantly different positions in PCA space (p<0.01). The week 5 post-vaccination cells, temporally associated with less protection to challenge, have greater PC2 values, indicating a more effector-like profile, whereas the week 20 post-vaccination cells have lesser PC2 values indicating a more central-memory like profile. The samples also significantly segregate based on epitope specificity (p<0.01). The Gag CM9-specific cells have overall higher PC2 values indicating a more effector-like phenotype, whereas the Tat SL8-specific cells have lower PC2 values indicating a more central memory-like phenotype. The changes in expression profiles from week 5 to week 20 are consistent with SIV-specific CD8+ T cells becoming overall more central memory-like and less effector-like over time following vaccination. Similarly, the differences seen between Gag CM9- and Tat SL8-specific cells are consistent with the kinetics of epitope escape and likely reflect the loss of antigen stimulation of the Tat-specific cells versus the ongoing stimulation of the Gag-specific cells.


Characterization of CD8+ T cell differentiation following SIVΔnef vaccination by transcription factor expression profiling.

Billingsley JM, Rajakumar PA, Connole MA, Salisch NC, Adnan S, Kuzmichev YV, Hong HS, Reeves RK, Kang HJ, Li W, Li Q, Haase AT, Johnson RP - PLoS Pathog. (2015)

Principal component analysis of transcription factor expression profiles from SIV-specific MHC tetramer-sorted CD8+ T cells and sorted CD8+ T cell subsets.(A) Plot of principal components 1 vs. 2, and 2 vs. 3 for each of the expression profiles assessed in sorted naïve and memory CD8+ T cell subsets isolated from healthy control animals (n = 5), and SIV-specific MHC tetramer-sorted CD8+ T cells isolated from animals (n = 4) at week 5 or week 20 following SIVΔnef vaccination. Principal components 1, 2 and 3 explain 92% of cumulative total variance. (B) PCA loading factors for each transcription factor.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4358830&req=5

ppat.1004740.g003: Principal component analysis of transcription factor expression profiles from SIV-specific MHC tetramer-sorted CD8+ T cells and sorted CD8+ T cell subsets.(A) Plot of principal components 1 vs. 2, and 2 vs. 3 for each of the expression profiles assessed in sorted naïve and memory CD8+ T cell subsets isolated from healthy control animals (n = 5), and SIV-specific MHC tetramer-sorted CD8+ T cells isolated from animals (n = 4) at week 5 or week 20 following SIVΔnef vaccination. Principal components 1, 2 and 3 explain 92% of cumulative total variance. (B) PCA loading factors for each transcription factor.
Mentions: To integrate the expression profiles of the SIV-specific cells with the sorted CD8+ subsets, we applied principal component analysis (PCA) to the combined data sets. Plotting principal components 1 vs, 2, and principal components 2 vs. 3, (PC1, PC2, PC3; Fig. 3A, S1 Video) segregated the data into distinct clusters. The data points representing the sorted CD8+ T cells occupy the periphery of the PC1 vs. PC2 plot, and segregate into separate clusters based upon cell differentiation stage. The naïve cells segregate from the memory cells along the PC1 axis, whereas the memory cells segregate along the PC2 axis, with the transitional memory cells positioned intermediately between the central and effector cells. The PC1 and PC2 loading factors (Fig. 3B) indicate that in this analysis, differential expression of LEF1, TCF7, PRDM1 and TBX21 strongly influence segregation of naïve from memory cells, whereas differential expression of ID2, RUNX3, AHR and LEF1 strongly influence segregation of memory cell subsets. The SIV-specific CD8+ T cells cluster with the sorted memory cells on the PC1 axis, and are positioned intermediately between central memory and effector memory cells on the PC2 axis. This intermediate position on the PC2 axis in part reflects the combined expression profiles of different memory subsets present in the SIV-specific cell samples. However, the SIV-specific samples significantly segregate from any sorted memory subset, particularly on the PC3 axis (p<0.001), indicating that the transcription factor expression profiles of the SIV-specific cells are distinct from the sorted subsets and are not solely comprised of proportions of memory subsets. The PC3 loading factors (Fig. 3B) indicate that in this analysis, the differential expression of NFIL3, IRF4, LEF1 and EOMES influence segregation of SIV-specific cells from the sorted naïve and memory subsets. The SIV-specific cells form two clusters, generally organized by week post-infection. The week 5 and week 20 post-vaccination samples occupy significantly different positions in PCA space (p<0.01). The week 5 post-vaccination cells, temporally associated with less protection to challenge, have greater PC2 values, indicating a more effector-like profile, whereas the week 20 post-vaccination cells have lesser PC2 values indicating a more central-memory like profile. The samples also significantly segregate based on epitope specificity (p<0.01). The Gag CM9-specific cells have overall higher PC2 values indicating a more effector-like phenotype, whereas the Tat SL8-specific cells have lower PC2 values indicating a more central memory-like phenotype. The changes in expression profiles from week 5 to week 20 are consistent with SIV-specific CD8+ T cells becoming overall more central memory-like and less effector-like over time following vaccination. Similarly, the differences seen between Gag CM9- and Tat SL8-specific cells are consistent with the kinetics of epitope escape and likely reflect the loss of antigen stimulation of the Tat-specific cells versus the ongoing stimulation of the Gag-specific cells.

Bottom Line: The onset of protective immunity against pathogenic SIV challenge in SIVΔnef-vaccinated macaques is delayed for 15-20 weeks, a process that is related to qualitative changes in CD8+ T cell responses induced by SIVΔnef.Expression of transcription factors associated with effector differentiation, such as ID2 and RUNX3, were decreased over time, while expression of transcription factors associated with quiescence or memory differentiation, such as TCF7, BCOR and EOMES, increased.These data suggest transcription factor expression profiling is a novel method that can provide additional data complementary to the analysis of memory cell differentiation based on classical phenotypic markers.

View Article: PubMed Central - PubMed

Affiliation: Division of Immunology, New England Primate Research Center, Harvard Medical School, Southborough, Massachusetts, United States of America; Yerkes National Primate Research Center, Emory University, Atlanta, Georgia, United States of America.

ABSTRACT
The onset of protective immunity against pathogenic SIV challenge in SIVΔnef-vaccinated macaques is delayed for 15-20 weeks, a process that is related to qualitative changes in CD8+ T cell responses induced by SIVΔnef. As a novel approach to characterize cell differentiation following vaccination, we used multi-target qPCR to measure transcription factor expression in naïve and memory subsets of CD8++ T cells, and in SIV-specific CD8+ T cells obtained from SIVΔnef-vaccinated or wild type SIVmac239-infected macaques. Unsupervised clustering of expression profiles organized naïve and memory CD8+ T cells into groups concordant with cell surface phenotype. Transcription factor expression patterns in SIV-specific CD8+ T cells in SIVΔnef-vaccinated animals were distinct from those observed in purified CD8+ T cell subsets obtained from naïve animals, and were intermediate to expression profiles of purified central memory and effector memory T cells. Expression of transcription factors elicited by SIVΔnef vaccination also varied over time: cells obtained at later time points, temporally associated with greater protection, appeared more central-memory like than cells obtained at earlier time points, which appeared more effector memory-like. Expression of transcription factors associated with effector differentiation, such as ID2 and RUNX3, were decreased over time, while expression of transcription factors associated with quiescence or memory differentiation, such as TCF7, BCOR and EOMES, increased. CD8+ T cells specific for a more conserved epitope expressed higher levels of TBX21 and BATF, and appeared more effector-like than cells specific for an escaped epitope, consistent with continued activation by replicating vaccine virus. These data suggest transcription factor expression profiling is a novel method that can provide additional data complementary to the analysis of memory cell differentiation based on classical phenotypic markers. Additionally, these data support the hypothesis that ongoing stimulation by SIVΔnef promotes a distinct protective balance of CD8+ T cell differentiation and activation states.

No MeSH data available.


Related in: MedlinePlus