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Editing anti-DNA B cells by Vlambdax.

Li Y, Louzoun Y, Weigert M - J. Exp. Med. (2004)

Bottom Line: However, the lambda locus, either of man or mouse, does not allow V gene replacement.We have found that in anti-DNA heavy chain transgenic mice (tgs) VH3H9/56R, B cells that express anti-DNA receptors comprised of lambda1 in association with an anti-DNA heavy chain often coexpress a kappa chain that prevents DNA binding.Here we describe a mechanism of preventing DNA binding by expression of a rarely used member of the Vlambda family, Vlambdax.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, NJ 08544, USA. yijinli@uchicago.edu

ABSTRACT
Receptor editing is performed by replacement of Vkappa genes that contribute to autoreactivity. In addition, the Ckappa locus can be deleted by Vkappa rearrangement to intronic or 3' of Ckappa RS sequences (also referred to as kappa deletion elements). B cells that delete the Ckappa can then express lambda light chains. However, the lambda locus, either of man or mouse, does not allow V gene replacement. Nor does it appear to be deleted. Therefore, editing of autoreactive lambda B cells may require alternative pathways. We have found that in anti-DNA heavy chain transgenic mice (tgs) VH3H9/56R, B cells that express anti-DNA receptors comprised of lambda1 in association with an anti-DNA heavy chain often coexpress a kappa chain that prevents DNA binding. We speculate that such isotypically included cells may have low anti-DNA receptor densities, a feature that may lead to self-tolerance. Here we describe a mechanism of preventing DNA binding by expression of a rarely used member of the Vlambda family, Vlambdax. The lambdax B cells of the tgs also express CD25 and may represent B cells that have exhausted light chain editing possibilities.

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CD25+ B cells express the Vλx-Jλ2 mRNA. CD25+/CD19+ and the CD25−/CD19+ B cells were isolated from the spleens of VH3H9/56R mice using anti-CD19 and CD25 antibodies and a FACS Vantage™ flow cytometer. cDNAs were synthesized from total RNA extracted from both fractions. Vλx-Jλ2 PRC was performed on serial dilutes of cDNA from each sample. The bottom panel shows the actin PCR.
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fig3: CD25+ B cells express the Vλx-Jλ2 mRNA. CD25+/CD19+ and the CD25−/CD19+ B cells were isolated from the spleens of VH3H9/56R mice using anti-CD19 and CD25 antibodies and a FACS Vantage™ flow cytometer. cDNAs were synthesized from total RNA extracted from both fractions. Vλx-Jλ2 PRC was performed on serial dilutes of cDNA from each sample. The bottom panel shows the actin PCR.

Mentions: To confirm that conventional L chains are replaced by Vλx, we sorted the CD25+ cells from the spleen of VH3H9/56R tg and performed a Vλx RT-PCR. We used a Jλ2 primer and a Vλx primer corresponding to a sequence spanning the FW2 and CDR2 region, which has no homology to Vλ1, Vλ2, or Vκ. A stronger signal is detected in the CD25+ fraction than the CD25− B cells sorted from the same mice (Fig. 3). PCR using genomic DNA from sorted cells shows similar results (see Fig. 6, middle).


Editing anti-DNA B cells by Vlambdax.

Li Y, Louzoun Y, Weigert M - J. Exp. Med. (2004)

CD25+ B cells express the Vλx-Jλ2 mRNA. CD25+/CD19+ and the CD25−/CD19+ B cells were isolated from the spleens of VH3H9/56R mice using anti-CD19 and CD25 antibodies and a FACS Vantage™ flow cytometer. cDNAs were synthesized from total RNA extracted from both fractions. Vλx-Jλ2 PRC was performed on serial dilutes of cDNA from each sample. The bottom panel shows the actin PCR.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2211795&req=5

fig3: CD25+ B cells express the Vλx-Jλ2 mRNA. CD25+/CD19+ and the CD25−/CD19+ B cells were isolated from the spleens of VH3H9/56R mice using anti-CD19 and CD25 antibodies and a FACS Vantage™ flow cytometer. cDNAs were synthesized from total RNA extracted from both fractions. Vλx-Jλ2 PRC was performed on serial dilutes of cDNA from each sample. The bottom panel shows the actin PCR.
Mentions: To confirm that conventional L chains are replaced by Vλx, we sorted the CD25+ cells from the spleen of VH3H9/56R tg and performed a Vλx RT-PCR. We used a Jλ2 primer and a Vλx primer corresponding to a sequence spanning the FW2 and CDR2 region, which has no homology to Vλ1, Vλ2, or Vκ. A stronger signal is detected in the CD25+ fraction than the CD25− B cells sorted from the same mice (Fig. 3). PCR using genomic DNA from sorted cells shows similar results (see Fig. 6, middle).

Bottom Line: However, the lambda locus, either of man or mouse, does not allow V gene replacement.We have found that in anti-DNA heavy chain transgenic mice (tgs) VH3H9/56R, B cells that express anti-DNA receptors comprised of lambda1 in association with an anti-DNA heavy chain often coexpress a kappa chain that prevents DNA binding.Here we describe a mechanism of preventing DNA binding by expression of a rarely used member of the Vlambda family, Vlambdax.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, NJ 08544, USA. yijinli@uchicago.edu

ABSTRACT
Receptor editing is performed by replacement of Vkappa genes that contribute to autoreactivity. In addition, the Ckappa locus can be deleted by Vkappa rearrangement to intronic or 3' of Ckappa RS sequences (also referred to as kappa deletion elements). B cells that delete the Ckappa can then express lambda light chains. However, the lambda locus, either of man or mouse, does not allow V gene replacement. Nor does it appear to be deleted. Therefore, editing of autoreactive lambda B cells may require alternative pathways. We have found that in anti-DNA heavy chain transgenic mice (tgs) VH3H9/56R, B cells that express anti-DNA receptors comprised of lambda1 in association with an anti-DNA heavy chain often coexpress a kappa chain that prevents DNA binding. We speculate that such isotypically included cells may have low anti-DNA receptor densities, a feature that may lead to self-tolerance. Here we describe a mechanism of preventing DNA binding by expression of a rarely used member of the Vlambda family, Vlambdax. The lambdax B cells of the tgs also express CD25 and may represent B cells that have exhausted light chain editing possibilities.

Show MeSH
Related in: MedlinePlus