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Restricted immunoglobulin variable region (Ig V) gene expression accompanies secondary rearrangements of light chain Ig V genes in mouse plasmacytomas.

Diaw L, Siwarski D, Coleman A, Kim J, Jones GM, Dighiero G, Huppi K - J. Exp. Med. (1999)

Bottom Line: The many binding studies of monoclonal immunoglobulin (Ig) produced by plasmacytomas have found no universally common binding properties, but instead, groups of plasmacytomas with specific antigen-binding activities to haptens such as phosphorylcholine, dextrans, fructofuranans, or dinitrophenyl.Subsequently, it was found that plasmacytomas with similar binding chain specificities not only expressed the same idiotype, but rearranged the same light (V(L)) and heavy (V(H)) variable region genes to express a characteristic monoclonal antibody.These results suggest that plasmacytomas use a restricted population of B cells that may still be undergoing rearrangement, thereby bypassing the allelic exclusion normally associated with expression of antibody genes.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The many binding studies of monoclonal immunoglobulin (Ig) produced by plasmacytomas have found no universally common binding properties, but instead, groups of plasmacytomas with specific antigen-binding activities to haptens such as phosphorylcholine, dextrans, fructofuranans, or dinitrophenyl. Subsequently, it was found that plasmacytomas with similar binding chain specificities not only expressed the same idiotype, but rearranged the same light (V(L)) and heavy (V(H)) variable region genes to express a characteristic monoclonal antibody. In this study, we have examined by enzyme-linked immunosorbent assay five antibodies secreted by silicone-induced mouse plasmacytomas using a broader panel of antigens including actin, myosin, tubulin, single-stranded DNA, and double-stranded DNA. We have determined the Ig heavy and light chain V gene usage in these same plasmacytomas at the DNA and RNA level. Our studies reveal: (a) antibodies secreted by plasmacytomas bind to different antigens in a manner similar to that observed for natural autoantibodies; (b) the expressed Ig heavy genes are restricted in V gene usage to the V(H)-J558 family; and (c) secondary rearrangements occur at the light chain level with at least three plasmacytomas expressing both kappa and lambda light chain genes. These results suggest that plasmacytomas use a restricted population of B cells that may still be undergoing rearrangement, thereby bypassing the allelic exclusion normally associated with expression of antibody genes.

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Transcript levels of Igκ, Igλ, and RAG-2 in SIPC tumors. (Top) Equal amounts of RNA from mouse liver (lane 1), spleen (lane 2), SIPC3301 (lane 3), SIPC3308 (lane 4), SIPC3282 (lane 5), SIPC3336 (lane 6), and SIPC3385 (lane 7) were loaded into slots and hybridized to nick-translated probes from Vκ24 (row 1), Vκ1 (row 2), Cκ (row 3), or Vλ (row 4). (Bottom) RNA from mouse spleen (lane 1), thymus (lane 2), SIPC3282 (lane 3), SIPC3301 (lane 4), SIPC3308 (lane 5), SIPC3336 (lane 6), SIPC3385 (lane 7), ABPC18 (lane 8), and MOPC104E (lane 9) was reverse transcribed (RT) and PCR amplified using primers specific for RAG-2.
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Figure 9: Transcript levels of Igκ, Igλ, and RAG-2 in SIPC tumors. (Top) Equal amounts of RNA from mouse liver (lane 1), spleen (lane 2), SIPC3301 (lane 3), SIPC3308 (lane 4), SIPC3282 (lane 5), SIPC3336 (lane 6), and SIPC3385 (lane 7) were loaded into slots and hybridized to nick-translated probes from Vκ24 (row 1), Vκ1 (row 2), Cκ (row 3), or Vλ (row 4). (Bottom) RNA from mouse spleen (lane 1), thymus (lane 2), SIPC3282 (lane 3), SIPC3301 (lane 4), SIPC3308 (lane 5), SIPC3336 (lane 6), SIPC3385 (lane 7), ABPC18 (lane 8), and MOPC104E (lane 9) was reverse transcribed (RT) and PCR amplified using primers specific for RAG-2.

Mentions: Levels of Igκ and Igλ were compared by slot blot hybridization using specific probes for Vκ1, Vκ24, Vλ, and Cκ (Fig. 9). Interestingly, SIPC3301 expressed high levels of Igλ, whereas SIPC3385 expressed high levels of Vκ1. Consistently, both SIPC3282 and SIPC3336 expressed high levels of both Vκ1 and Vκ24. Since there is evidence that peritoneal cavity B cells are enriched in B-1 cells, we also tested for Ly1 expression by RT-PCR amplification of total RNA from thymus, spleen, two PCs, (ABPC18, MOPC104E) and the SIPC tumors. Although expression of Ly1 was found in thymus, spleen, and the two conventional PCs, no expression of Ly1 was evident in the SIPC tumors (data not shown). RAG-1 (data not shown) and RAG-2 (Fig. 9) activity, both of which have recently been found in the peripheral B-1 population 23, were also independently assayed by RT-PCR amplification among similar panels of RNAs. Both RAG-1 and RAG-2 expression were found in thymus, spleen, SIPC3301, SIPC3308, SIPC3385, ABPC18, and MOPC104E, but not in SIPC3282 or SIPC3336.


Restricted immunoglobulin variable region (Ig V) gene expression accompanies secondary rearrangements of light chain Ig V genes in mouse plasmacytomas.

Diaw L, Siwarski D, Coleman A, Kim J, Jones GM, Dighiero G, Huppi K - J. Exp. Med. (1999)

Transcript levels of Igκ, Igλ, and RAG-2 in SIPC tumors. (Top) Equal amounts of RNA from mouse liver (lane 1), spleen (lane 2), SIPC3301 (lane 3), SIPC3308 (lane 4), SIPC3282 (lane 5), SIPC3336 (lane 6), and SIPC3385 (lane 7) were loaded into slots and hybridized to nick-translated probes from Vκ24 (row 1), Vκ1 (row 2), Cκ (row 3), or Vλ (row 4). (Bottom) RNA from mouse spleen (lane 1), thymus (lane 2), SIPC3282 (lane 3), SIPC3301 (lane 4), SIPC3308 (lane 5), SIPC3336 (lane 6), SIPC3385 (lane 7), ABPC18 (lane 8), and MOPC104E (lane 9) was reverse transcribed (RT) and PCR amplified using primers specific for RAG-2.
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Related In: Results  -  Collection

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Figure 9: Transcript levels of Igκ, Igλ, and RAG-2 in SIPC tumors. (Top) Equal amounts of RNA from mouse liver (lane 1), spleen (lane 2), SIPC3301 (lane 3), SIPC3308 (lane 4), SIPC3282 (lane 5), SIPC3336 (lane 6), and SIPC3385 (lane 7) were loaded into slots and hybridized to nick-translated probes from Vκ24 (row 1), Vκ1 (row 2), Cκ (row 3), or Vλ (row 4). (Bottom) RNA from mouse spleen (lane 1), thymus (lane 2), SIPC3282 (lane 3), SIPC3301 (lane 4), SIPC3308 (lane 5), SIPC3336 (lane 6), SIPC3385 (lane 7), ABPC18 (lane 8), and MOPC104E (lane 9) was reverse transcribed (RT) and PCR amplified using primers specific for RAG-2.
Mentions: Levels of Igκ and Igλ were compared by slot blot hybridization using specific probes for Vκ1, Vκ24, Vλ, and Cκ (Fig. 9). Interestingly, SIPC3301 expressed high levels of Igλ, whereas SIPC3385 expressed high levels of Vκ1. Consistently, both SIPC3282 and SIPC3336 expressed high levels of both Vκ1 and Vκ24. Since there is evidence that peritoneal cavity B cells are enriched in B-1 cells, we also tested for Ly1 expression by RT-PCR amplification of total RNA from thymus, spleen, two PCs, (ABPC18, MOPC104E) and the SIPC tumors. Although expression of Ly1 was found in thymus, spleen, and the two conventional PCs, no expression of Ly1 was evident in the SIPC tumors (data not shown). RAG-1 (data not shown) and RAG-2 (Fig. 9) activity, both of which have recently been found in the peripheral B-1 population 23, were also independently assayed by RT-PCR amplification among similar panels of RNAs. Both RAG-1 and RAG-2 expression were found in thymus, spleen, SIPC3301, SIPC3308, SIPC3385, ABPC18, and MOPC104E, but not in SIPC3282 or SIPC3336.

Bottom Line: The many binding studies of monoclonal immunoglobulin (Ig) produced by plasmacytomas have found no universally common binding properties, but instead, groups of plasmacytomas with specific antigen-binding activities to haptens such as phosphorylcholine, dextrans, fructofuranans, or dinitrophenyl.Subsequently, it was found that plasmacytomas with similar binding chain specificities not only expressed the same idiotype, but rearranged the same light (V(L)) and heavy (V(H)) variable region genes to express a characteristic monoclonal antibody.These results suggest that plasmacytomas use a restricted population of B cells that may still be undergoing rearrangement, thereby bypassing the allelic exclusion normally associated with expression of antibody genes.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Genetics, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The many binding studies of monoclonal immunoglobulin (Ig) produced by plasmacytomas have found no universally common binding properties, but instead, groups of plasmacytomas with specific antigen-binding activities to haptens such as phosphorylcholine, dextrans, fructofuranans, or dinitrophenyl. Subsequently, it was found that plasmacytomas with similar binding chain specificities not only expressed the same idiotype, but rearranged the same light (V(L)) and heavy (V(H)) variable region genes to express a characteristic monoclonal antibody. In this study, we have examined by enzyme-linked immunosorbent assay five antibodies secreted by silicone-induced mouse plasmacytomas using a broader panel of antigens including actin, myosin, tubulin, single-stranded DNA, and double-stranded DNA. We have determined the Ig heavy and light chain V gene usage in these same plasmacytomas at the DNA and RNA level. Our studies reveal: (a) antibodies secreted by plasmacytomas bind to different antigens in a manner similar to that observed for natural autoantibodies; (b) the expressed Ig heavy genes are restricted in V gene usage to the V(H)-J558 family; and (c) secondary rearrangements occur at the light chain level with at least three plasmacytomas expressing both kappa and lambda light chain genes. These results suggest that plasmacytomas use a restricted population of B cells that may still be undergoing rearrangement, thereby bypassing the allelic exclusion normally associated with expression of antibody genes.

Show MeSH
Related in: MedlinePlus