Limits...
Somatic hypermutation introduces insertions and deletions into immunoglobulin V genes.

Wilson PC, de Bouteiller O, Liu YJ, Potter K, Banchereau J, Capra JD, Pascual V - J. Exp. Med. (1998)

Bottom Line: Antigen-directed selection of B cell clones that generate high affinity surface Ig results in the affinity maturation of the antibody response.No germline genes that could have encoded these six cDNA clones were found after an extensive characterization of the genomic VH4 repertoire of the tonsil donor.The lack of similar instances in unmutated IgD+CD38- follicular mantle cDNA clones statistically associates these events to the somatic hypermutation process (P = 0.014).

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology Center, Department of Microbiology, University of Texas Southwestern Medical Center at Dallas, Texas 75235-9140, USA.

ABSTRACT
During a germinal center reaction, random mutations are introduced into immunoglobulin V genes to increase the affinity of antibody molecules and to further diversify the B cell repertoire. Antigen-directed selection of B cell clones that generate high affinity surface Ig results in the affinity maturation of the antibody response. The mutations of Ig genes are typically basepair substitutions, although DNA insertions and deletions have been reported to occur at a low frequency. In this study, we describe five insertion and four deletion events in otherwise somatically mutated VH gene cDNA molecules. Two of these insertions and all four deletions were obtained through the sequencing of 395 cDNA clones (approximately 110,000 nucleotides) from CD38+IgD- germinal center, and CD38-IgD- memory B cell populations from a single human tonsil. No germline genes that could have encoded these six cDNA clones were found after an extensive characterization of the genomic VH4 repertoire of the tonsil donor. These six insertions or deletions and three additional insertion events isolated from other sources occurred as triplets or multiples thereof, leaving the transcripts in frame. Additionally, 8 of 9 of these events occurred in the CDR1 or CDR2, following a pattern consistent with selection, and making it unlikely that these events were artifacts of the experimental system. The lack of similar instances in unmutated IgD+CD38- follicular mantle cDNA clones statistically associates these events to the somatic hypermutation process (P = 0.014). Close scrutiny of the 9 insertion/deletion events reported here, and of 25 additional insertions or deletions collected from the literature, suggest that secondary structural elements in the DNA sequences capable of producing loop intermediates may be a prerequisite in most instances. Furthermore, these events most frequently involve sequence motifs resembling known intrinsic hotspots of somatic hypermutation. These insertion/deletion events are consistent with models of somatic hypermutation involving an unstable polymerase enzyme complex lacking proofreading capabilities, and suggest a downregulation or alteration of DNA repair at the V locus during the hypermutation process.

Show MeSH

Related in: MedlinePlus

Proposed mechanism causing insertion/deletion events: polymerase slippage. This Figure is based on model a of Streisinger et al. (30)  and Ripley (31). The same model can account for both (A) insertions and  (B) deletions.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2199186&req=5

Figure 6: Proposed mechanism causing insertion/deletion events: polymerase slippage. This Figure is based on model a of Streisinger et al. (30) and Ripley (31). The same model can account for both (A) insertions and (B) deletions.

Mentions: The evidence for the involvement of DNA secondary structure in the production of insertion or deletion mutations during somatic hypermutation, as suggested in 1986 by Golding et al. (29), now seems unequivocal. The insertions and deletions described in our study, and those illustrated in Table 3, occur in a predictable fashion, involving sequence motifs that could form loop intermediates reminiscent of the replication slippage model of Streisinger et al. (30) and Ripley and Glickman (for review see 31) as presented in Fig. 6. Such mutations are postulated to occur when DNA polymerase slips or stutters and the newly synthesized strand shifts on the template and reanneals to an adjacent repetitive element, producing unpaired loop intermediates localized to one or the other strands. If this unpaired loop intermediate is not repaired then it will be perpetuated as an insertion of an instance of the repetitive element if in the daughter strand, or a deletion if in the template strand.


Somatic hypermutation introduces insertions and deletions into immunoglobulin V genes.

Wilson PC, de Bouteiller O, Liu YJ, Potter K, Banchereau J, Capra JD, Pascual V - J. Exp. Med. (1998)

Proposed mechanism causing insertion/deletion events: polymerase slippage. This Figure is based on model a of Streisinger et al. (30)  and Ripley (31). The same model can account for both (A) insertions and  (B) deletions.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199186&req=5

Figure 6: Proposed mechanism causing insertion/deletion events: polymerase slippage. This Figure is based on model a of Streisinger et al. (30) and Ripley (31). The same model can account for both (A) insertions and (B) deletions.
Mentions: The evidence for the involvement of DNA secondary structure in the production of insertion or deletion mutations during somatic hypermutation, as suggested in 1986 by Golding et al. (29), now seems unequivocal. The insertions and deletions described in our study, and those illustrated in Table 3, occur in a predictable fashion, involving sequence motifs that could form loop intermediates reminiscent of the replication slippage model of Streisinger et al. (30) and Ripley and Glickman (for review see 31) as presented in Fig. 6. Such mutations are postulated to occur when DNA polymerase slips or stutters and the newly synthesized strand shifts on the template and reanneals to an adjacent repetitive element, producing unpaired loop intermediates localized to one or the other strands. If this unpaired loop intermediate is not repaired then it will be perpetuated as an insertion of an instance of the repetitive element if in the daughter strand, or a deletion if in the template strand.

Bottom Line: Antigen-directed selection of B cell clones that generate high affinity surface Ig results in the affinity maturation of the antibody response.No germline genes that could have encoded these six cDNA clones were found after an extensive characterization of the genomic VH4 repertoire of the tonsil donor.The lack of similar instances in unmutated IgD+CD38- follicular mantle cDNA clones statistically associates these events to the somatic hypermutation process (P = 0.014).

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology Center, Department of Microbiology, University of Texas Southwestern Medical Center at Dallas, Texas 75235-9140, USA.

ABSTRACT
During a germinal center reaction, random mutations are introduced into immunoglobulin V genes to increase the affinity of antibody molecules and to further diversify the B cell repertoire. Antigen-directed selection of B cell clones that generate high affinity surface Ig results in the affinity maturation of the antibody response. The mutations of Ig genes are typically basepair substitutions, although DNA insertions and deletions have been reported to occur at a low frequency. In this study, we describe five insertion and four deletion events in otherwise somatically mutated VH gene cDNA molecules. Two of these insertions and all four deletions were obtained through the sequencing of 395 cDNA clones (approximately 110,000 nucleotides) from CD38+IgD- germinal center, and CD38-IgD- memory B cell populations from a single human tonsil. No germline genes that could have encoded these six cDNA clones were found after an extensive characterization of the genomic VH4 repertoire of the tonsil donor. These six insertions or deletions and three additional insertion events isolated from other sources occurred as triplets or multiples thereof, leaving the transcripts in frame. Additionally, 8 of 9 of these events occurred in the CDR1 or CDR2, following a pattern consistent with selection, and making it unlikely that these events were artifacts of the experimental system. The lack of similar instances in unmutated IgD+CD38- follicular mantle cDNA clones statistically associates these events to the somatic hypermutation process (P = 0.014). Close scrutiny of the 9 insertion/deletion events reported here, and of 25 additional insertions or deletions collected from the literature, suggest that secondary structural elements in the DNA sequences capable of producing loop intermediates may be a prerequisite in most instances. Furthermore, these events most frequently involve sequence motifs resembling known intrinsic hotspots of somatic hypermutation. These insertion/deletion events are consistent with models of somatic hypermutation involving an unstable polymerase enzyme complex lacking proofreading capabilities, and suggest a downregulation or alteration of DNA repair at the V locus during the hypermutation process.

Show MeSH
Related in: MedlinePlus