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The transcriptional promoter regulates hypermutation of the antibody heavy chain locus.

Tumas-Brundage K, Manser T - J. Exp. Med. (1997)

Bottom Line: However, while the distribution of mutation in such loci appears normal, the frequency of mutation does not.Conversely, moving the VH promoter 750 bp upstream of its normal location results in a commensurate change in the site specificity of hypermutation in H chain loci, and the foreign DNA inserted into the VH leader intron to produce this promoter displacement is hypermutated in a manner indistinguishable from natural Ig DNA.These data establish a direct mechanistic link between the IgH transcription and hypermutation processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Kimmel Cancer Institute, Thomas Jefferson Medical College, Philadelphia, Pennsylvania 19107, USA.

ABSTRACT
A somatic process introduces mutations into antibody variable (V) region genes at a high rate in many vertebrates, and is a major source of antibody diversity. The mechanism of this hypermutation process remains enigmatic, although retrospective studies and transgenic experiments have recently suggested a role for transcriptional regulatory elements. Here, we demonstrate that mouse heavy (H) chain loci in which the natural VH promoter has been replaced by a heterologous promoter undergo hypermutation. However, while the distribution of mutation in such loci appears normal, the frequency of mutation does not. Conversely, moving the VH promoter 750 bp upstream of its normal location results in a commensurate change in the site specificity of hypermutation in H chain loci, and the foreign DNA inserted into the VH leader intron to produce this promoter displacement is hypermutated in a manner indistinguishable from natural Ig DNA. These data establish a direct mechanistic link between the IgH transcription and hypermutation processes.

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The total number of hybridomas expressing BP2 (B29 promoter-driven) and CPM (enlarged leader intron) hybrid loci with a given  number of VH somatic mutations. Values for wild-type 36-65VH hybrid  loci were obtained from previously published data (18). All hybridomas  from CPM transgenic mice and wild-type 36-65VH transgenic mice were  generated using the hyperimmunized primary protocol and 8–10-wk-old  mice. For BP2 mice, primary, secondary, and tertiary immunization protocols were used (see Materials and Methods). Those hybrid locus expressing hybridomas from the B29 line that had the largest number of  mutations (>2) were from secondary and tertiary immunizations.
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Figure 2: The total number of hybridomas expressing BP2 (B29 promoter-driven) and CPM (enlarged leader intron) hybrid loci with a given number of VH somatic mutations. Values for wild-type 36-65VH hybrid loci were obtained from previously published data (18). All hybridomas from CPM transgenic mice and wild-type 36-65VH transgenic mice were generated using the hyperimmunized primary protocol and 8–10-wk-old mice. For BP2 mice, primary, secondary, and tertiary immunization protocols were used (see Materials and Methods). Those hybrid locus expressing hybridomas from the B29 line that had the largest number of mutations (>2) were from secondary and tertiary immunizations.

Mentions: Low levels of VH mutation were also observed on average in enlarged leader intron hybrid loci expressed by late primary response hybridomas, and many such loci contained no VH mutations. As was the case for the hybridomas containing B29 promoter-driven hybrid loci, the canonical Vκ mutation frequency in hybridomas containing enlarged leader intron hybrid loci appeared normal (data from seven representative hybridomas are shown in Table 1). As summarized in Fig. 2, the mutation frequencies in the VH genes in both types of hybrid loci are well below that previously determined for wild-type 36-65VH hybrid loci at times late in the primary response.


The transcriptional promoter regulates hypermutation of the antibody heavy chain locus.

Tumas-Brundage K, Manser T - J. Exp. Med. (1997)

The total number of hybridomas expressing BP2 (B29 promoter-driven) and CPM (enlarged leader intron) hybrid loci with a given  number of VH somatic mutations. Values for wild-type 36-65VH hybrid  loci were obtained from previously published data (18). All hybridomas  from CPM transgenic mice and wild-type 36-65VH transgenic mice were  generated using the hyperimmunized primary protocol and 8–10-wk-old  mice. For BP2 mice, primary, secondary, and tertiary immunization protocols were used (see Materials and Methods). Those hybrid locus expressing hybridomas from the B29 line that had the largest number of  mutations (>2) were from secondary and tertiary immunizations.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2196128&req=5

Figure 2: The total number of hybridomas expressing BP2 (B29 promoter-driven) and CPM (enlarged leader intron) hybrid loci with a given number of VH somatic mutations. Values for wild-type 36-65VH hybrid loci were obtained from previously published data (18). All hybridomas from CPM transgenic mice and wild-type 36-65VH transgenic mice were generated using the hyperimmunized primary protocol and 8–10-wk-old mice. For BP2 mice, primary, secondary, and tertiary immunization protocols were used (see Materials and Methods). Those hybrid locus expressing hybridomas from the B29 line that had the largest number of mutations (>2) were from secondary and tertiary immunizations.
Mentions: Low levels of VH mutation were also observed on average in enlarged leader intron hybrid loci expressed by late primary response hybridomas, and many such loci contained no VH mutations. As was the case for the hybridomas containing B29 promoter-driven hybrid loci, the canonical Vκ mutation frequency in hybridomas containing enlarged leader intron hybrid loci appeared normal (data from seven representative hybridomas are shown in Table 1). As summarized in Fig. 2, the mutation frequencies in the VH genes in both types of hybrid loci are well below that previously determined for wild-type 36-65VH hybrid loci at times late in the primary response.

Bottom Line: However, while the distribution of mutation in such loci appears normal, the frequency of mutation does not.Conversely, moving the VH promoter 750 bp upstream of its normal location results in a commensurate change in the site specificity of hypermutation in H chain loci, and the foreign DNA inserted into the VH leader intron to produce this promoter displacement is hypermutated in a manner indistinguishable from natural Ig DNA.These data establish a direct mechanistic link between the IgH transcription and hypermutation processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Kimmel Cancer Institute, Thomas Jefferson Medical College, Philadelphia, Pennsylvania 19107, USA.

ABSTRACT
A somatic process introduces mutations into antibody variable (V) region genes at a high rate in many vertebrates, and is a major source of antibody diversity. The mechanism of this hypermutation process remains enigmatic, although retrospective studies and transgenic experiments have recently suggested a role for transcriptional regulatory elements. Here, we demonstrate that mouse heavy (H) chain loci in which the natural VH promoter has been replaced by a heterologous promoter undergo hypermutation. However, while the distribution of mutation in such loci appears normal, the frequency of mutation does not. Conversely, moving the VH promoter 750 bp upstream of its normal location results in a commensurate change in the site specificity of hypermutation in H chain loci, and the foreign DNA inserted into the VH leader intron to produce this promoter displacement is hypermutated in a manner indistinguishable from natural Ig DNA. These data establish a direct mechanistic link between the IgH transcription and hypermutation processes.

Show MeSH