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Analysis of immunoglobulin transcripts and hypermutation following SHIV(AD8) infection and protein-plus-adjuvant immunization.

Francica JR, Sheng Z, Zhang Z, Nishimura Y, Shingai M, Ramesh A, Keele BF, Schmidt SD, Flynn BJ, Darko S, Lynch RM, Yamamoto T, Matus-Nicodemos R, Wolinsky D, NISC Comparative Sequencing ProgramNason M, Valiante NM, Malyala P, De Gregorio E, Barnett SW, Singh M, O'Hagan DT, Koup RA, Mascola JR, Martin MA, Kepler TB, Douek DC, Shapiro L, Seder RA - Nat Commun (2015)

Bottom Line: Here we use two nonhuman primate models to compare the roles of antigen persistence, diversity and innate immunity.The use of adjuvants results in increased ELISA titres but does not affect the mean SHM levels or CDR H3 lengths.Our study shows how the ontogeny of Env-specific B cells can be tracked, and provides insights into the requirements for developing neutralizing antibodies that should facilitate translation to human vaccine studies.

View Article: PubMed Central - PubMed

Affiliation: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
Developing predictive animal models to assess how candidate vaccines and infection influence the ontogenies of Envelope (Env)-specific antibodies is critical for the development of an HIV vaccine. Here we use two nonhuman primate models to compare the roles of antigen persistence, diversity and innate immunity. We perform longitudinal analyses of HIV Env-specific B-cell receptor responses to SHIV(AD8) infection and Env protein vaccination with eight different adjuvants. A subset of the SHIV(AD8)-infected animals with higher viral loads and greater Env diversity show increased neutralization associated with increasing somatic hypermutation (SHM) levels over time. The use of adjuvants results in increased ELISA titres but does not affect the mean SHM levels or CDR H3 lengths. Our study shows how the ontogeny of Env-specific B cells can be tracked, and provides insights into the requirements for developing neutralizing antibodies that should facilitate translation to human vaccine studies.

No MeSH data available.


Related in: MedlinePlus

Next-generation sequencing of antigen-specific B cells after SHIVAD8 infection.Data are shown from four good neutralizers, red; and four poor neutralizers, blue. (a,b) SHM for each animal over time; each symbol represents the average percent divergence from germline for Env-specific (gp120+) (a) or nonspecific (gp120−) (b) sequences from a given animal; error bars indicate s.e.m. (c) Histogram representation of SHM distribution from all sequences from 90–110 weeks post infection; binning averaged in 2% increments. (d) CDR H3 length distribution of Env-specific sequences from individual animals for all time points combined. Colour/symbol scheme as in a; binning averaged in 3-aa increments; arrow indicates population of sequences with long CDR H3 regions. (e) Proportion of unique sequences with long CDR H3 regions from DCF1 at the indicated time points. χ2-test with Yates correction was used to calculate P values between proportions from Env+ or − samples at each time point; n.s., not significant; *P<0.0001. (f) Long CDR H3 regions from DCF1 may be derived in at least two unique ways. Example 1 depicts the CDR H3 region of sequence 4594, arising from V(DD)J recombination. Example 2 depicts the CDR H3 region of sequence 107, arising from N-addition. Bold text indicates mature antibody sequence; red stars indicate predicted tyrosine sulfation; CDR H3 charge is shown. (g) Divergence over time of long CDR H3 antibodies from the parent lineage that includes sequence 4594. A phylogenic tree was constructed by maximum likelihood and rooted to the IGHV4D*01 allele and is colour-coded by time point. (h,i) Two-dimensional plots depicting Env-specific sequences from animal DCF1 at the indicated time points. Sequences are plotted by their VH divergence from germline and their identity to the 4594 HC (h); or by their CDR H3 length (i). Red triangle indicates sequence 4594; magenta triangles indicate sequences related to 4594; shaded boxes indicate reads with long CDR H3 regions.
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f2: Next-generation sequencing of antigen-specific B cells after SHIVAD8 infection.Data are shown from four good neutralizers, red; and four poor neutralizers, blue. (a,b) SHM for each animal over time; each symbol represents the average percent divergence from germline for Env-specific (gp120+) (a) or nonspecific (gp120−) (b) sequences from a given animal; error bars indicate s.e.m. (c) Histogram representation of SHM distribution from all sequences from 90–110 weeks post infection; binning averaged in 2% increments. (d) CDR H3 length distribution of Env-specific sequences from individual animals for all time points combined. Colour/symbol scheme as in a; binning averaged in 3-aa increments; arrow indicates population of sequences with long CDR H3 regions. (e) Proportion of unique sequences with long CDR H3 regions from DCF1 at the indicated time points. χ2-test with Yates correction was used to calculate P values between proportions from Env+ or − samples at each time point; n.s., not significant; *P<0.0001. (f) Long CDR H3 regions from DCF1 may be derived in at least two unique ways. Example 1 depicts the CDR H3 region of sequence 4594, arising from V(DD)J recombination. Example 2 depicts the CDR H3 region of sequence 107, arising from N-addition. Bold text indicates mature antibody sequence; red stars indicate predicted tyrosine sulfation; CDR H3 charge is shown. (g) Divergence over time of long CDR H3 antibodies from the parent lineage that includes sequence 4594. A phylogenic tree was constructed by maximum likelihood and rooted to the IGHV4D*01 allele and is colour-coded by time point. (h,i) Two-dimensional plots depicting Env-specific sequences from animal DCF1 at the indicated time points. Sequences are plotted by their VH divergence from germline and their identity to the 4594 HC (h); or by their CDR H3 length (i). Red triangle indicates sequence 4594; magenta triangles indicate sequences related to 4594; shaded boxes indicate reads with long CDR H3 regions.

Mentions: To determine how SHIVAD8 infection influences Ig maturation, AD8 gp120 Env-specific and nonspecific memory B cells from infected animals were isolated at various time points post infection, sequenced and analysed (Supplementary Table 2). A total of 5,526 unique HC sequences were derived from 28,000 Env-specific cells, and 311,274 HC sequences were derived from ∼1,313,000 nonspecific cells. Env-specific B cells accumulated mutations over the course of infection and two of the four good neutralizers had higher divergence levels than the poor neutralizers (Fig. 2a). As a control, sequences from nonspecific memory B cells did not significantly accumulate mutations over the course of the infection (Fig. 2b). Indeed, by 110 weeks post infection Env-specific B cells had accumulated more mutations from germline than had nonspecific cells (Fig. 2c). These data show that SHIVAD8 infection results in the accumulation of SHM that is HIV-specific and correlates with serum neutralization.


Analysis of immunoglobulin transcripts and hypermutation following SHIV(AD8) infection and protein-plus-adjuvant immunization.

Francica JR, Sheng Z, Zhang Z, Nishimura Y, Shingai M, Ramesh A, Keele BF, Schmidt SD, Flynn BJ, Darko S, Lynch RM, Yamamoto T, Matus-Nicodemos R, Wolinsky D, NISC Comparative Sequencing ProgramNason M, Valiante NM, Malyala P, De Gregorio E, Barnett SW, Singh M, O'Hagan DT, Koup RA, Mascola JR, Martin MA, Kepler TB, Douek DC, Shapiro L, Seder RA - Nat Commun (2015)

Next-generation sequencing of antigen-specific B cells after SHIVAD8 infection.Data are shown from four good neutralizers, red; and four poor neutralizers, blue. (a,b) SHM for each animal over time; each symbol represents the average percent divergence from germline for Env-specific (gp120+) (a) or nonspecific (gp120−) (b) sequences from a given animal; error bars indicate s.e.m. (c) Histogram representation of SHM distribution from all sequences from 90–110 weeks post infection; binning averaged in 2% increments. (d) CDR H3 length distribution of Env-specific sequences from individual animals for all time points combined. Colour/symbol scheme as in a; binning averaged in 3-aa increments; arrow indicates population of sequences with long CDR H3 regions. (e) Proportion of unique sequences with long CDR H3 regions from DCF1 at the indicated time points. χ2-test with Yates correction was used to calculate P values between proportions from Env+ or − samples at each time point; n.s., not significant; *P<0.0001. (f) Long CDR H3 regions from DCF1 may be derived in at least two unique ways. Example 1 depicts the CDR H3 region of sequence 4594, arising from V(DD)J recombination. Example 2 depicts the CDR H3 region of sequence 107, arising from N-addition. Bold text indicates mature antibody sequence; red stars indicate predicted tyrosine sulfation; CDR H3 charge is shown. (g) Divergence over time of long CDR H3 antibodies from the parent lineage that includes sequence 4594. A phylogenic tree was constructed by maximum likelihood and rooted to the IGHV4D*01 allele and is colour-coded by time point. (h,i) Two-dimensional plots depicting Env-specific sequences from animal DCF1 at the indicated time points. Sequences are plotted by their VH divergence from germline and their identity to the 4594 HC (h); or by their CDR H3 length (i). Red triangle indicates sequence 4594; magenta triangles indicate sequences related to 4594; shaded boxes indicate reads with long CDR H3 regions.
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Related In: Results  -  Collection

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f2: Next-generation sequencing of antigen-specific B cells after SHIVAD8 infection.Data are shown from four good neutralizers, red; and four poor neutralizers, blue. (a,b) SHM for each animal over time; each symbol represents the average percent divergence from germline for Env-specific (gp120+) (a) or nonspecific (gp120−) (b) sequences from a given animal; error bars indicate s.e.m. (c) Histogram representation of SHM distribution from all sequences from 90–110 weeks post infection; binning averaged in 2% increments. (d) CDR H3 length distribution of Env-specific sequences from individual animals for all time points combined. Colour/symbol scheme as in a; binning averaged in 3-aa increments; arrow indicates population of sequences with long CDR H3 regions. (e) Proportion of unique sequences with long CDR H3 regions from DCF1 at the indicated time points. χ2-test with Yates correction was used to calculate P values between proportions from Env+ or − samples at each time point; n.s., not significant; *P<0.0001. (f) Long CDR H3 regions from DCF1 may be derived in at least two unique ways. Example 1 depicts the CDR H3 region of sequence 4594, arising from V(DD)J recombination. Example 2 depicts the CDR H3 region of sequence 107, arising from N-addition. Bold text indicates mature antibody sequence; red stars indicate predicted tyrosine sulfation; CDR H3 charge is shown. (g) Divergence over time of long CDR H3 antibodies from the parent lineage that includes sequence 4594. A phylogenic tree was constructed by maximum likelihood and rooted to the IGHV4D*01 allele and is colour-coded by time point. (h,i) Two-dimensional plots depicting Env-specific sequences from animal DCF1 at the indicated time points. Sequences are plotted by their VH divergence from germline and their identity to the 4594 HC (h); or by their CDR H3 length (i). Red triangle indicates sequence 4594; magenta triangles indicate sequences related to 4594; shaded boxes indicate reads with long CDR H3 regions.
Mentions: To determine how SHIVAD8 infection influences Ig maturation, AD8 gp120 Env-specific and nonspecific memory B cells from infected animals were isolated at various time points post infection, sequenced and analysed (Supplementary Table 2). A total of 5,526 unique HC sequences were derived from 28,000 Env-specific cells, and 311,274 HC sequences were derived from ∼1,313,000 nonspecific cells. Env-specific B cells accumulated mutations over the course of infection and two of the four good neutralizers had higher divergence levels than the poor neutralizers (Fig. 2a). As a control, sequences from nonspecific memory B cells did not significantly accumulate mutations over the course of the infection (Fig. 2b). Indeed, by 110 weeks post infection Env-specific B cells had accumulated more mutations from germline than had nonspecific cells (Fig. 2c). These data show that SHIVAD8 infection results in the accumulation of SHM that is HIV-specific and correlates with serum neutralization.

Bottom Line: Here we use two nonhuman primate models to compare the roles of antigen persistence, diversity and innate immunity.The use of adjuvants results in increased ELISA titres but does not affect the mean SHM levels or CDR H3 lengths.Our study shows how the ontogeny of Env-specific B cells can be tracked, and provides insights into the requirements for developing neutralizing antibodies that should facilitate translation to human vaccine studies.

View Article: PubMed Central - PubMed

Affiliation: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
Developing predictive animal models to assess how candidate vaccines and infection influence the ontogenies of Envelope (Env)-specific antibodies is critical for the development of an HIV vaccine. Here we use two nonhuman primate models to compare the roles of antigen persistence, diversity and innate immunity. We perform longitudinal analyses of HIV Env-specific B-cell receptor responses to SHIV(AD8) infection and Env protein vaccination with eight different adjuvants. A subset of the SHIV(AD8)-infected animals with higher viral loads and greater Env diversity show increased neutralization associated with increasing somatic hypermutation (SHM) levels over time. The use of adjuvants results in increased ELISA titres but does not affect the mean SHM levels or CDR H3 lengths. Our study shows how the ontogeny of Env-specific B cells can be tracked, and provides insights into the requirements for developing neutralizing antibodies that should facilitate translation to human vaccine studies.

No MeSH data available.


Related in: MedlinePlus