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Augmented expression of a human gene for 8-oxoguanine DNA glycosylase (MutM) in B lymphocytes of the dark zone in lymph node germinal centers.

Kuo FC, Sklar J - J. Exp. Med. (1997)

Bottom Line: Northern blot analysis indicated that the human gene is expressed as two alternatively spliced messenger RNAs within GC B cells at levels greatly exceeding that found in other tissues.In situ hybridization studies revealed that expression of this gene is most abundant within the dark zones of GCs.Both the function and localized expression of this gene suggest that it may play a role in somatic hypermutation of immunoglobulin genes.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Oncology, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
B cells that mediate normal, T cell-dependent, humoral immune responses must first pass through germinal centers (GCs) within the cortex of antigenically stimulated lymph nodes. As they move through the dark zone and then the light zone in the GC, B cells are subjected to somatic hypermutation and switch recombination within their rearranged immunoglobulin genes and also participate in a number of other processes that control development into memory cells or cells specialized for antibody secretion. To investigate the molecular mechanisms that contribute to B cell development within GCs, we constructed a recombinant DNA library enriched for cDNAs derived from human genes expressed in B cells at this site. This library was found to contain a cDNA structurally and functionally related to genes in bacteria and yeast for the DNA repair enzyme 8-oxoguanine DNA glycosylase. Northern blot analysis indicated that the human gene is expressed as two alternatively spliced messenger RNAs within GC B cells at levels greatly exceeding that found in other tissues. In situ hybridization studies revealed that expression of this gene is most abundant within the dark zones of GCs. Both the function and localized expression of this gene suggest that it may play a role in somatic hypermutation of immunoglobulin genes.

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Comparison of amino acid sequences of GCN5, GCN6, and  yeast Ogg1 (YOGGI). The alignment was performed by the Bestfit program in the Wisconsin GCG package. Identical amino acids are marked  by a vertical line between amino acids in the respective sequences and  conserved changes by a colon. Dots represent gaps in the sequence generated by the program to allow better alignment. The overall identity is calculated to be 37%. The GCN5 is a partial cDNA that begins at codon 215  in GCN6. The aspartic acid conserved in the presumed active site among  DNA glycosylases is underlined. The DNA sequences reported in the paper are available from EMBL/Genbank/DDBJ under accession number  AF 026691.
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Figure 3: Comparison of amino acid sequences of GCN5, GCN6, and yeast Ogg1 (YOGGI). The alignment was performed by the Bestfit program in the Wisconsin GCG package. Identical amino acids are marked by a vertical line between amino acids in the respective sequences and conserved changes by a colon. Dots represent gaps in the sequence generated by the program to allow better alignment. The overall identity is calculated to be 37%. The GCN5 is a partial cDNA that begins at codon 215 in GCN6. The aspartic acid conserved in the presumed active site among DNA glycosylases is underlined. The DNA sequences reported in the paper are available from EMBL/Genbank/DDBJ under accession number AF 026691.

Mentions: The GCN6 cDNA contains a 1,023-bp open reading frame capable of encoding a protein having a molecular weight of 37 kD. A BLAST homology search against a database of known protein sequences revealed that the putative protein encoded by GCN6 has significant homology to a recently reported yeast base excision DNA repair protein, 8-oxoguanine DNA glycosylase (Ogg)1. Fig. 3 shows a comparison of the GCN6 coding sequence and Ogg1. Several regions of the two sequences are strongly homologous, including one toward the COOH-terminal end of the GCN6 open reading frame (codons 235–310), corresponding to a part of Ogg1 proposed to contain the active site of the enzyme. Within this region, the aspartic acid codon at position 269 of the GCN6 sequence is conserved among many of the DNA glycosylases and is thought to encode the amino acid that acts as the nucleophile during the enzymatic reaction. For as far as it extends, the coding sequence in GCN5 is identical to that in GCN6 before position 307 in the latter, after which GCN5 diverges completely and continues for another 66 codons. Examination of this region of the sequence in the GCN5 cDNA did not reveal significant homology to any protein in the database.


Augmented expression of a human gene for 8-oxoguanine DNA glycosylase (MutM) in B lymphocytes of the dark zone in lymph node germinal centers.

Kuo FC, Sklar J - J. Exp. Med. (1997)

Comparison of amino acid sequences of GCN5, GCN6, and  yeast Ogg1 (YOGGI). The alignment was performed by the Bestfit program in the Wisconsin GCG package. Identical amino acids are marked  by a vertical line between amino acids in the respective sequences and  conserved changes by a colon. Dots represent gaps in the sequence generated by the program to allow better alignment. The overall identity is calculated to be 37%. The GCN5 is a partial cDNA that begins at codon 215  in GCN6. The aspartic acid conserved in the presumed active site among  DNA glycosylases is underlined. The DNA sequences reported in the paper are available from EMBL/Genbank/DDBJ under accession number  AF 026691.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2199107&req=5

Figure 3: Comparison of amino acid sequences of GCN5, GCN6, and yeast Ogg1 (YOGGI). The alignment was performed by the Bestfit program in the Wisconsin GCG package. Identical amino acids are marked by a vertical line between amino acids in the respective sequences and conserved changes by a colon. Dots represent gaps in the sequence generated by the program to allow better alignment. The overall identity is calculated to be 37%. The GCN5 is a partial cDNA that begins at codon 215 in GCN6. The aspartic acid conserved in the presumed active site among DNA glycosylases is underlined. The DNA sequences reported in the paper are available from EMBL/Genbank/DDBJ under accession number AF 026691.
Mentions: The GCN6 cDNA contains a 1,023-bp open reading frame capable of encoding a protein having a molecular weight of 37 kD. A BLAST homology search against a database of known protein sequences revealed that the putative protein encoded by GCN6 has significant homology to a recently reported yeast base excision DNA repair protein, 8-oxoguanine DNA glycosylase (Ogg)1. Fig. 3 shows a comparison of the GCN6 coding sequence and Ogg1. Several regions of the two sequences are strongly homologous, including one toward the COOH-terminal end of the GCN6 open reading frame (codons 235–310), corresponding to a part of Ogg1 proposed to contain the active site of the enzyme. Within this region, the aspartic acid codon at position 269 of the GCN6 sequence is conserved among many of the DNA glycosylases and is thought to encode the amino acid that acts as the nucleophile during the enzymatic reaction. For as far as it extends, the coding sequence in GCN5 is identical to that in GCN6 before position 307 in the latter, after which GCN5 diverges completely and continues for another 66 codons. Examination of this region of the sequence in the GCN5 cDNA did not reveal significant homology to any protein in the database.

Bottom Line: Northern blot analysis indicated that the human gene is expressed as two alternatively spliced messenger RNAs within GC B cells at levels greatly exceeding that found in other tissues.In situ hybridization studies revealed that expression of this gene is most abundant within the dark zones of GCs.Both the function and localized expression of this gene suggest that it may play a role in somatic hypermutation of immunoglobulin genes.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Oncology, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
B cells that mediate normal, T cell-dependent, humoral immune responses must first pass through germinal centers (GCs) within the cortex of antigenically stimulated lymph nodes. As they move through the dark zone and then the light zone in the GC, B cells are subjected to somatic hypermutation and switch recombination within their rearranged immunoglobulin genes and also participate in a number of other processes that control development into memory cells or cells specialized for antibody secretion. To investigate the molecular mechanisms that contribute to B cell development within GCs, we constructed a recombinant DNA library enriched for cDNAs derived from human genes expressed in B cells at this site. This library was found to contain a cDNA structurally and functionally related to genes in bacteria and yeast for the DNA repair enzyme 8-oxoguanine DNA glycosylase. Northern blot analysis indicated that the human gene is expressed as two alternatively spliced messenger RNAs within GC B cells at levels greatly exceeding that found in other tissues. In situ hybridization studies revealed that expression of this gene is most abundant within the dark zones of GCs. Both the function and localized expression of this gene suggest that it may play a role in somatic hypermutation of immunoglobulin genes.

Show MeSH