scholarly journals Phospholipase Cγ2 Contributes to Light-Chain Gene Activation and Receptor Editing

2007 ◽  
Vol 27 (17) ◽  
pp. 5957-5967 ◽  
Author(s):  
Li Bai ◽  
Yuhong Chen ◽  
Yinghong He ◽  
Xuezhi Dai ◽  
Xueyan Lin ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Light Chain ◽  
Gene Activation ◽  
Chain Gene ◽  
Receptor Editing ◽  
Light Chain Gene ◽  
Transitional Type

ABSTRACT Phospholipase Cγ2 (PLCγ2) is critical for pre-B-cell receptor (pre-BCR) and BCR signaling. Current studies discovered that PLCγ2-deficient mice had reduced immunoglobulin λ (Igλ) light-chain usage throughout B-cell maturation stages, including transitional type 1 (T1), transitional type 2 (T2), and mature follicular B cells. The reduction of Igλ rearrangement by PLCγ2 deficiency was not due to specifically increased apoptosis or decreased proliferation of mutant Igλ+ B cells, as lack of PLCγ2 exerted a similar effect on apoptosis and proliferation of both Igλ+ and Igκ+ B cells. Moreover, PLCγ2-deficient IgHEL transgenic B cells exhibited an impairment of antigen-induced receptor editing among both the endogenous λ and κ loci in vitro and in vivo. Importantly, PLCγ2 deficiency impaired BCR-induced expression of IRF-4 and IRF-8, the two transcription factors critical for λ and κ light-chain rearrangements. Taken together, these data demonstrate that the PLCγ2 signaling pathway plays a role in activation of light-chain loci and contributes to receptor editing.

scholarly journals Receptor Editing Occurs Frequently during Normal B Cell Development

1998 ◽  
Vol 188 (7) ◽  
pp. 1231-1238 ◽  
Author(s):  
Marc W. Retter ◽  
David Nemazee
Keyword(s):  
B Cells ◽  
B Cell ◽  
Light Chain ◽  
Cell Receptor ◽  
Feedback Mechanism ◽  
Chain Gene ◽  
Allelic Exclusion ◽  
Receptor Editing ◽  
Light Chain Gene ◽  
Immature B Cells

Allelic exclusion is established in development through a feedback mechanism in which the assembled immunoglobulin (Ig) suppresses further V(D)J rearrangement. But Ig expression sometimes fails to prevent further rearrangement. In autoantibody transgenic mice, reactivity of immature B cells with autoantigen can induce receptor editing, in which allelic exclusion is transiently prevented or reversed through nested light chain gene rearrangement, often resulting in altered B cell receptor specificity. To determine the extent of receptor editing in a normal, non-Ig transgenic immune system, we took advantage of the fact that λ light chain genes usually rearrange after κ genes. This allowed us to analyze κ loci in IgMλ+ cells to determine how frequently in-frame κ genes fail to suppress λ gene rearrangements. To do this, we analyzed recombined VκJκ genes inactivated by subsequent recombining sequence (RS) rearrangement. RS rearrangements delete portions of the κ locus by a V(D)J recombinase-dependent mechanism, suggesting that they play a role in receptor editing. We show that RS recombination is frequently induced by, and inactivates, functionally rearranged κ loci, as nearly half (47%) of the RS-inactivated VκJκ joins were in-frame. These findings suggest that receptor editing occurs at a surprisingly high frequency in normal B cells.

Myosin light-chain 1 and 3 gene has two structurally distinct and differentially regulated promoters evolving at different rates

1985 ◽  
Vol 5 (11) ◽  
pp. 3168-3182
Author(s):  
E E Strehler ◽  
M Periasamy ◽  
M A Strehler-Page ◽  
B Nadal-Ginard
Keyword(s):  
Light Chain ◽  
Myosin Light Chain ◽  
Mammalian Species ◽  
In Vitro Transcription ◽  
Chain Gene ◽  
Promoter Regions ◽  
Light Chain Gene ◽  
Direct Initiation

DNA fragments located 10 kilobases apart in the genome and containing, respectively, the first myosin light chain 1 (MLC1f) and the first myosin light chain 3 (MLC3f) specific exon of the rat myosin light chain 1 and 3 gene, together with several hundred base pairs of upstream flanking sequences, have been shown in runoff in vitro transcription assays to direct initiation of transcription at the cap sites of MLC1f and MLC3f mRNAs used in vivo. These results establish the presence of two separate, functional promoters within that gene. A comparison of the nucleotide sequence of the rat MLC1f/3f gene with the corresponding sequences from mouse and chicken shows that: the MLC1f promoter regions have been highly conserved up to position -150 from the cap site while the MLC3f promoter regions display a very poor degree of homology and even the absence or poor conservation of typical eucaryotic promoter elements such as TATA and CAT boxes; the exon/intron structure of this gene has been completely conserved in the three species; and corresponding exons, except for the regions encoding most of the 5' and 3' untranslated sequences, show greater than 75% homology while corresponding introns are similar in size but considerably divergent in sequence. The above findings indicate that the overall structure of the MLC1f/3f genes has been maintained between avian and mammalian species and that these genes contain two functional and widely spaced promoters. The fact that the structures of the alkali light chain gene from Drosophila melanogaster and of other related genes of the troponin C supergene family resemble a MLC3f gene without an upstream promoter and first exon strongly suggests that the present-day MLC1f/3f genes of higher vertebrates arose from a primordial alkali light chain gene through the addition of a far-upstream MLC1f-specific promoter and first exon. The two promoters have evolved at different rates, with the MLC1f promoter being more conserved than the MLC3f promoter. This discrepant evolutionary rate might reflect different mechanisms of promoter activation for the transcription of MLC1f and MLC3f RNA.

Ongoing diversification of the rearranged immunoglobulin light-chain gene in a bursal lymphoma cell line

1990 ◽  
Vol 10 (6) ◽  
pp. 3224-3231
Author(s):  
S Kim ◽  
E H Humphries ◽  
L Tjoelker ◽  
L Carlson ◽  
C B Thompson
Keyword(s):  
B Cell ◽  
Light Chain ◽  
Gene Segment ◽  
B Cell Development ◽  
Bursa Of Fabricius ◽  
Chain Gene ◽  
Lymphoma Cell Line ◽  
Immunoglobulin Light Chain ◽  
Light Chain Gene

The chicken immunoglobulin light-chain gene (IgL) encodes only a single variable gene segment capable of recombination. To generate an immune repertoire, chickens diversify this unique rearranged VL gene segment during B-cell development in the bursa of Fabricius. Sequence analysis of IgL cDNAs suggests that both gene conversion events derived from VL segment pseudogene templates (psi VL) and non-template-derived single-base-pair substitutions contribute to this diversity. To facilitate the study of postrecombinational mechanisms of immunoglobulin gene diversification, avian B-cell lines were examined for the ability to diversify their rearranged IgL gene during in vitro passage. One line that retains this ability, the avian leukosis virus-induced bursal lymphoma cell line DT40, has been identified. After passage for 1 year in culture, 39 of 51 randomly sequenced rearranged V-J segments from a DT40 population defined novel subclones of the parental tumor. All cloned V-J segments displayed the same V-J joint, confirming that the observed diversity arose after V-J rearrangement. Most sequence variations that we observed (203 of 220 base pairs) appeared to result from psi VL-derived gene conversion events; 16 of the 17 novel single nucleotide substitutions were transitions. Based on these data, it appears that immunoglobulin diversification during in vitro passage of DT40 cells is representative of the diversification that occurs during normal B-cell development in the bursa of Fabricius.

scholarly journals Myosin light-chain 1 and 3 gene has two structurally distinct and differentially regulated promoters evolving at different rates.

1985 ◽  
Vol 5 (11) ◽  
pp. 3168-3182 ◽  
Author(s):  
E E Strehler ◽  
M Periasamy ◽  
M A Strehler-Page ◽  
B Nadal-Ginard
Keyword(s):  
Light Chain ◽  
Myosin Light Chain ◽  
Mammalian Species ◽  
In Vitro Transcription ◽  
Chain Gene ◽  
Promoter Regions ◽  
Light Chain Gene ◽  
Direct Initiation

DNA fragments located 10 kilobases apart in the genome and containing, respectively, the first myosin light chain 1 (MLC1f) and the first myosin light chain 3 (MLC3f) specific exon of the rat myosin light chain 1 and 3 gene, together with several hundred base pairs of upstream flanking sequences, have been shown in runoff in vitro transcription assays to direct initiation of transcription at the cap sites of MLC1f and MLC3f mRNAs used in vivo. These results establish the presence of two separate, functional promoters within that gene. A comparison of the nucleotide sequence of the rat MLC1f/3f gene with the corresponding sequences from mouse and chicken shows that: the MLC1f promoter regions have been highly conserved up to position -150 from the cap site while the MLC3f promoter regions display a very poor degree of homology and even the absence or poor conservation of typical eucaryotic promoter elements such as TATA and CAT boxes; the exon/intron structure of this gene has been completely conserved in the three species; and corresponding exons, except for the regions encoding most of the 5' and 3' untranslated sequences, show greater than 75% homology while corresponding introns are similar in size but considerably divergent in sequence. The above findings indicate that the overall structure of the MLC1f/3f genes has been maintained between avian and mammalian species and that these genes contain two functional and widely spaced promoters. The fact that the structures of the alkali light chain gene from Drosophila melanogaster and of other related genes of the troponin C supergene family resemble a MLC3f gene without an upstream promoter and first exon strongly suggests that the present-day MLC1f/3f genes of higher vertebrates arose from a primordial alkali light chain gene through the addition of a far-upstream MLC1f-specific promoter and first exon. The two promoters have evolved at different rates, with the MLC1f promoter being more conserved than the MLC3f promoter. This discrepant evolutionary rate might reflect different mechanisms of promoter activation for the transcription of MLC1f and MLC3f RNA.

scholarly journals Ongoing diversification of the rearranged immunoglobulin light-chain gene in a bursal lymphoma cell line.

1990 ◽  
Vol 10 (6) ◽  
pp. 3224-3231 ◽  
Author(s):  
S Kim ◽  
E H Humphries ◽  
L Tjoelker ◽  
L Carlson ◽  
C B Thompson
Keyword(s):  
B Cell ◽  
Light Chain ◽  
Gene Segment ◽  
B Cell Development ◽  
Bursa Of Fabricius ◽  
Chain Gene ◽  
Lymphoma Cell Line ◽  
Immunoglobulin Light Chain ◽  
Light Chain Gene

The chicken immunoglobulin light-chain gene (IgL) encodes only a single variable gene segment capable of recombination. To generate an immune repertoire, chickens diversify this unique rearranged VL gene segment during B-cell development in the bursa of Fabricius. Sequence analysis of IgL cDNAs suggests that both gene conversion events derived from VL segment pseudogene templates (psi VL) and non-template-derived single-base-pair substitutions contribute to this diversity. To facilitate the study of postrecombinational mechanisms of immunoglobulin gene diversification, avian B-cell lines were examined for the ability to diversify their rearranged IgL gene during in vitro passage. One line that retains this ability, the avian leukosis virus-induced bursal lymphoma cell line DT40, has been identified. After passage for 1 year in culture, 39 of 51 randomly sequenced rearranged V-J segments from a DT40 population defined novel subclones of the parental tumor. All cloned V-J segments displayed the same V-J joint, confirming that the observed diversity arose after V-J rearrangement. Most sequence variations that we observed (203 of 220 base pairs) appeared to result from psi VL-derived gene conversion events; 16 of the 17 novel single nucleotide substitutions were transitions. Based on these data, it appears that immunoglobulin diversification during in vitro passage of DT40 cells is representative of the diversification that occurs during normal B-cell development in the bursa of Fabricius.

CD19 regulates positive selection and maturation in B lymphopoiesis: lack of CD19 imposes developmental arrest of immature B cells and consequential stimulation of receptor editing

Blood ◽  
2005 ◽  
Vol 105 (8) ◽  
pp. 3247-3254 ◽  
Author(s):  
Eran Diamant ◽  
Zohar Keren ◽  
Doron Melamed
Keyword(s):  
B Cells ◽  
Positive Selection ◽  
B Cell ◽  
Receptor Editing ◽  
B Lymphopoiesis ◽  
Developmental Arrest ◽  
Surface Marker Expression ◽  
Immature B Cells

AbstractLigand-independent signals that are produced by the B-cell antigen receptor (BCR) confer an important positive selection checkpoint for immature B cells. Generation of inappropriate signals imposes developmental arrest of immature B cells, though the fate of these cells has not been investigated. Studies have shown that the lack of CD19 results in inappropriate signaling. In immunoglobulin transgenic mice, this inappropriate signaling impairs positive selection and stimulates receptor editing. Here, we studied the extent and significance of receptor editing in CD19-regulated positive selection of normal, nontransgenic B lymphopoiesis, using our bone marrow culture system. We found that the lack of CD19 resulted in elevated tonic signaling and impaired maturation, as revealed by surface marker expression and by functional assays. Immature CD19-/- B cells did not suppress RAG and underwent intensive receptor editing attempts in culture. Finally, in vivo analysis of light-chain isotype expression and Jκ use in CD19-/- mice validated our in vitro observations. Our results suggest that CD19 has an important function in regulating positive selection and maturation of nontransgenic B-cell precursors and that receptor editing is an important salvage mechanism for immature B cells that fail positive selection. (Blood. 2005;105:3247-3254)

scholarly journals BCL6 is critical for the development of a diverse primary B cell repertoire

2010 ◽  
Vol 207 (6) ◽  
pp. 1209-1221 ◽  
Author(s):  
Cihangir Duy ◽  
J. Jessica Yu ◽  
Rahul Nahar ◽  
Srividya Swaminathan ◽  
Soo-Mi Kweon ◽  
...  
Keyword(s):  
Dna Damage ◽  
B Cells ◽  
B Cell ◽  
Light Chain ◽  
Gene Rearrangement ◽  
Chain Gene ◽  
Cell Repertoire ◽  
Light Chain Gene ◽  
B Cell Repertoire ◽  
Induced Apoptosis

BCL6 protects germinal center (GC) B cells against DNA damage–induced apoptosis during somatic hypermutation and class-switch recombination. Although expression of BCL6 was not found in early IL-7–dependent B cell precursors, we report that IL-7Rα–Stat5 signaling negatively regulates BCL6. Upon productive VH-DJH gene rearrangement and expression of a μ heavy chain, however, activation of pre–B cell receptor signaling strongly induces BCL6 expression, whereas IL-7Rα–Stat5 signaling is attenuated. At the transition from IL-7–dependent to –independent stages of B cell development, BCL6 is activated, reaches expression levels resembling those in GC B cells, and protects pre–B cells from DNA damage–induced apoptosis during immunoglobulin (Ig) light chain gene recombination. In the absence of BCL6, DNA breaks during Ig light chain gene rearrangement lead to excessive up-regulation of Arf and p53. As a consequence, the pool of new bone marrow immature B cells is markedly reduced in size and clonal diversity. We conclude that negative regulation of Arf by BCL6 is required for pre–B cell self-renewal and the formation of a diverse polyclonal B cell repertoire.

Differential surrogate light chain expression governs B-cell differentiation

Blood ◽  
2002 ◽  
Vol 99 (7) ◽  
pp. 2459-2467 ◽  
Author(s):  
Yui-Hsi Wang ◽  
Robert P. Stephan ◽  
Alexander Scheffold ◽  
Désirée Kunkel ◽  
Hajime Karasuyama ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
B Cells ◽  
B Cell ◽  
Light Chain ◽  
B Cell Receptors ◽  
Chain Gene ◽  
B Cell Differentiation ◽  
Light Chain Gene ◽  
Surrogate Light Chain ◽  
Rag 1

Surrogate light chain expression during B lineage differentiation was examined by using indicator fluorochrome-filled liposomes in an enhanced immunofluorescence assay. Pro-B cells bearing surrogate light chain components were found in mice, but not in humans. A limited subpopulation of relatively large pre-B cells in both species expressed pre-B cell receptors. These cells had reduced expression of the recombinase activating genes, RAG-1 and RAG-2. Their receptor-negative pre-B cell progeny were relatively small, expressed RAG-1 and RAG-2, and exhibited selective down-regulation of VpreB and λ5expression. Comparative analysis of the 2 pre-B cell subpopulations indicated that loss of the pre-B cell receptors from surrogate light chain gene silencing was linked with exit from the cell cycle and light chain gene rearrangement to achieve B-cell differentiation.

scholarly journals Normal human B cells display ordered light chain gene rearrangements and deletions.

1982 ◽  
Vol 156 (4) ◽  
pp. 975-985 ◽  
Author(s):  
S J Korsmeyer ◽  
P A Hieter ◽  
S O Sharrow ◽  
C K Goldman ◽  
P Leder ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Light Chain ◽  
Developmental Process ◽  
Germ Line ◽  
Chain Gene ◽  
Gene Rearrangements ◽  
Lambda Light Chain ◽  
Light Chain Gene

Human kappa-producing B cell lines and leukemias retain their excluded lambda light chain genes in the germ line configuration, whereas transformed lambda-producing B cells uniformly rearrange or delete their kappa genes (12). Whether the unexpected lambda gene recombinations within malignant lambda-producing B cells reflect a normal developmental process or are secondary to transformation and specific chromosomal translocations was uncertain. To resolve this issue, we purified circulating lambda-bearing B cells from a normal individual to 97% purity by using a series of negative selection steps and a final positive selection on a cell sorter. Over 95% of the collective kappa genes in these lambda B cells were no longer in their germ line form, with the majority (60%) deleted and the remainder present but in a rearranged state. The chromosomal loss of the germ line kappa genes included the joining (J kappa) segments as well as the constant (C kappa) region, yet the particular variable (V kappa) gene family studied was spared. In addition, the incidence of kappa gene deletions was higher in long-term than in freshly transformed lambda B cell lines. This implies that the deletion of aberrantly rearranged kappa genes may occur as a second event. Such a mechanism would serve to eliminate aberrant transcripts and light chain fragments that might interfere with the synthesis and assembly of effective immunoglobulin molecules. Thus, despite the nearly equal usage of kappa and lambda light chain genes in man, there appears to be a sequential order to their expression during normal B cell ontogeny in which kappa gene rearrangements precede those of lambda.

scholarly journals NF-κB activity marks cells engaged in receptor editing

2009 ◽  
Vol 206 (8) ◽  
pp. 1803-1816 ◽  
Author(s):  
Emily J. Cadera ◽  
Fengyi Wan ◽  
Rupesh H. Amin ◽  
Hector Nolla ◽  
Michael J. Lenardo ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
Chain Gene ◽  
Receptor Editing ◽  
Light Chain Gene ◽  
Reporter Mice ◽  
Bone Marrow Cultures ◽  
Chain Gene Rearrangement ◽  
Primary Bone

Because of the extreme diversity in immunoglobulin genes, tolerance mechanisms are necessary to ensure that B cells do not respond to self-antigens. One such tolerance mechanism is called receptor editing. If the B cell receptor (BCR) on an immature B cell recognizes self-antigen, it is down-regulated from the cell surface, and light chain gene rearrangement continues in an attempt to edit the autoreactive specificity. Analysis of a heterozygous mutant mouse in which the NF-κB–dependent IκBα gene was replaced with a lacZ (β-gal) reporter complementary DNA (cDNA; IκBα+/lacZ) suggests a potential role for NF-κB in receptor editing. Sorted β-gal+ pre–B cells showed increased levels of various markers of receptor editing. In IκBα+/lacZ reporter mice expressing either innocuous or self-specific knocked in BCRs, β-gal was preferentially expressed in pre–B cells from the mice with self-specific BCRs. Retroviral-mediated expression of a cDNA encoding an IκBα superrepressor in primary bone marrow cultures resulted in diminished germline κ and rearranged λ transcripts but similar levels of RAG expression as compared with controls. We found that IRF4 transcripts were up-regulated in β-gal+ pre–B cells. Because IRF4 is a target of NF-κB and is required for receptor editing, we suggest that NF-κB could be acting through IRF4 to regulate receptor editing.

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