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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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Diagram of the  GCN5 and GCN6 cDNAs. The  labeled arrows above each cDNA  represent PCR primers used to  generate region-specific probes.  The two striped regions represent  coding exon in the calcium-calmodulin–dependent protein kinase I transcript.
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Figure 2: Diagram of the GCN5 and GCN6 cDNAs. The labeled arrows above each cDNA represent PCR primers used to generate region-specific probes. The two striped regions represent coding exon in the calcium-calmodulin–dependent protein kinase I transcript.

Mentions: The relationship between GCN5 and 6 is illustrated in Fig. 2. Amplification by PCR of DNA from the unsubtracted GC B cell cDNA library using a 5′ primer complementary to sequence in the middle of GCN6 (Fig. 2, S4) and a 3′ primer complementary to sequence in the first part of GCN5 (Fig. 2, A5) yielded a fragment of the size and sequence expected from a continuous stretch of DNA (data not shown). It therefore appears that GCN5 is an incomplete cDNA that shares a 5′ and coding sequence with GCN6. However, GCN5 and 6 differ from each other in exonic sequences at their 3′ ends, and consequently seem to represent alternatively spliced transcripts of the same gene. This interpretation was supported by the isolation of genomic DNA fragments containing these sequences and the finding within the genomic fragments of apparent intronic sequences adjacent to the exons present in the two cDNAs.


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)

Diagram of the  GCN5 and GCN6 cDNAs. The  labeled arrows above each cDNA  represent PCR primers used to  generate region-specific probes.  The two striped regions represent  coding exon in the calcium-calmodulin–dependent protein kinase I transcript.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199107&req=5

Figure 2: Diagram of the GCN5 and GCN6 cDNAs. The labeled arrows above each cDNA represent PCR primers used to generate region-specific probes. The two striped regions represent coding exon in the calcium-calmodulin–dependent protein kinase I transcript.
Mentions: The relationship between GCN5 and 6 is illustrated in Fig. 2. Amplification by PCR of DNA from the unsubtracted GC B cell cDNA library using a 5′ primer complementary to sequence in the middle of GCN6 (Fig. 2, S4) and a 3′ primer complementary to sequence in the first part of GCN5 (Fig. 2, A5) yielded a fragment of the size and sequence expected from a continuous stretch of DNA (data not shown). It therefore appears that GCN5 is an incomplete cDNA that shares a 5′ and coding sequence with GCN6. However, GCN5 and 6 differ from each other in exonic sequences at their 3′ ends, and consequently seem to represent alternatively spliced transcripts of the same gene. This interpretation was supported by the isolation of genomic DNA fragments containing these sequences and the finding within the genomic fragments of apparent intronic sequences adjacent to the exons present in the two cDNAs.

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