Expression of error-prone polymerases in BL2 cells activated for Ig somatic hypermutation

Vladimir Poltoratsky, Caroline J. Woo, Brigette Tippin, Alberto Martin, Myron F. Goodman, Matthew D. Scharff

Research output: Contribution to journalArticle

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Abstract

High affinity antibodies are generated in mice and humans by means of somatic hypermutation (SHM) of variable (V) regions of Ig genes. Mutations with rates of 10-5-10-3 per base pair per generation, about 106-fold above normal, are targeted primarily at V-region hot spots by unknown mechanisms. We have measured mRNA expression of DNA polymerases ι, η, and ζ by using cultured Burkitt's lymphoma (BL)2 cells. These cells exhibit 5-10-fold increases in heavy-chain V-region mutations targeted only predominantly to RGYW (R = A or G, Y = C or T, W = T or A) hot spots if costimulated with T cells and IgM crosslinking, the presumed in vivo requirements for SHM. An ∼4-fold increase pol ι mRNA occurs within 12 h when cocultured with T cells and surface IgM crosslinking. Induction of pols η and ζ occur with T cells, IgM crosslinking, or both stimuli. The fidelity of pol ι was measured at RGYW hot-and non-hot-spot sequences situated at nicks, gaps, and double-strand breaks. Pol ι formed T·G mispairs at a frequency of 10-2, consistent with SHM-generated C to T transitions, with a 3-fold increased error rate in hot- vs. non-hot-spot sequences for the single-nucleotide overhang. The T cell and IgM crosslinking-dependent induction of pol ι at 12 h may indicate an SHM "triggering" event has occurred. However, pols ι, η, and ζ are present under all conditions, suggesting that their presence is not sufficient to generate mutations because both T cell and IgM stimuli are required for SHM induction.

Original languageEnglish (US)
Pages (from-to)7976-7981
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume98
Issue number14
DOIs
StatePublished - 2001

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Immunoglobulin Somatic Hypermutation
Immunoglobulin M
T-Lymphocytes
Messenger RNA
Immunoglobulin Genes
Mutation
Antibody Affinity
Burkitt Lymphoma
Mutation Rate
DNA-Directed DNA Polymerase
Base Pairing

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Expression of error-prone polymerases in BL2 cells activated for Ig somatic hypermutation. / Poltoratsky, Vladimir; Woo, Caroline J.; Tippin, Brigette; Martin, Alberto; Goodman, Myron F.; Scharff, Matthew D.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 98, No. 14, 2001, p. 7976-7981.

Research output: Contribution to journalArticle

Poltoratsky, Vladimir ; Woo, Caroline J. ; Tippin, Brigette ; Martin, Alberto ; Goodman, Myron F. ; Scharff, Matthew D. / Expression of error-prone polymerases in BL2 cells activated for Ig somatic hypermutation. In: Proceedings of the National Academy of Sciences of the United States of America. 2001 ; Vol. 98, No. 14. pp. 7976-7981.
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