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The Journal of Immunology, 1999, 162: 1566-1572.
Copyright © 1999 by The American Association of Immunologists

Atypical VH-D-JH Rearrangements in Newborn Autoimmune MRL Mice1

Kimberly D. Klonowski, Laura L. Primiano and Marc Monestier2

Department of Microbiology and Immunology, Temple University School of Medicine, Philadelphia, PA 19140


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Antinuclear Abs are the hallmark of the autoimmune disease systemic lupus erythematosus (SLE). The ability of self reactive autoantibodies to bind to DNA and nucleosomes is partly conferred by an increased number of arginine and asparagine residues in the heavy chain third complementarity determining region. This increased content of cationic residues is primarily the result of unusual VH-D-JH rearrangements, which include D-D fusions and D gene inversions. While self Ag-driven clonal expansion is a major contributor to the production of antinuclear Abs in lupus, we explore in this study the hypothesis that newly emerging B cells from autoimmune mice display more frequently these unusual VH-D-JH rearrangements. To this end, libraries of PCR-generated VH-D-JH junctions from MRL and C3H newborn livers were analyzed. When compared with the C3H controls, D and JH gene usage in MRL junctions suggests a greater frequency of secondary D-JH rearrangements in this strain. Furthermore, B cells from the autoimmune-prone MRL mice have significantly increased numbers of atypical VH-D-JH rearrangements (D-D fusions and D inversions). Therefore, B cells from MRL mice manifest intrinsic defects that could confer an increased propensity to produce unusual VH-D-JH rearrangements early in ontogeny.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Most exogenous and internal factors that contribute to the development of autoimmunity remain to be elucidated. One of the hallmarks of systemic lupus erythematosus (SLE) is the presence of autoantibodies specific for chromatin Ags such as DNA or native histone-DNA complexes (nucleosomes) (1, 2, 3, 4). Anti-DNA and anti-nucleosome Abs are characterized by the presence of cationic residues (mostly arginine) at specific positions (5, 6, 7). In anti-DNA Abs, these residues are contact points with negatively charged sites on the DNA helix (8). In anti-nucleosome Abs, these arginine side chains are presumably responsible for the interaction with the DNA moiety of the nucleosome (9, 10, 11, 12). Most of these cationic residues are located in the H chain complementarity determining region 3 (CDR3)3 and result from atypical VH-D-JH joints (4, 13). Indeed, anti-DNA and anti-nucleosome Abs display unusual rearrangements that include alternate D reading frames, inverted D segments, and D-D fusions (productive joining of two D segments in a single rearrangement). In contrast, these atypical VH-D-JH rearrangements are rarely encountered in the nonautoimmune murine repertoire (14).

A likely explanation for the presence of these otherwise infrequent VH-D-JH rearrangements in anti-chromatin Abs is that the rare B cells expressing them have been selectively expanded because of their reactivity with DNA or DNA-histone complexes. There is indeed a large body of evidence showing that self reactive B cells are clonally expanded during the autoimmune process (15). An additional mechanism may be that newly emerging B cells from autoimmune mice more frequently possess these unusual VH-D-JH rearrangements. If this hypothesis is correct, differences in VH-D-JH rearrangement patterns between normal and autoimmune mice could be apparent before the development of the autoimmune disease. In the present study, we therefore test the hypothesis that atypical rearrangements are more frequent in autoimmune-prone mice by comparing libraries of VH-D-JH joints that we have generated from newborn autoimmune and normal mice. The results suggest that, before the development of autoimmunity, atypical junctions (D-D fusions and inverted D segments) and secondary D-JH rearrangements occur more frequently in newborn MRL than in C3H animals.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Mice

Male and female animals from the autoimmune MRL/MpJ +/+ (MRL) and the Igh allotype-matched normal C3H/HeJ (C3H) strains were obtained from The Jackson Laboratory (Bar Harbor, ME) and bred in our animal facilities (16). Newborn animals (<24 h old) were sacrificed and their livers were collected and stored frozen at -80°C.

Primers

DNA was extracted from pooled (five to six) newborn livers using a genomic DNA extraction kit (Promega, Madison, WI) and was then subjected to PCR amplification. The PCR primers and protocols were adapted from previously described methods (17, 18, 19) (Table IGo). Amplifications were conducted using nested PCR as follows. In the first PCR, DNA was amplified with a mixed set of antisense JH primers and either a sense J558 primer or a sense 7183 primer (corresponding to a consensus framework 1 region). In the second PCR, 10 µl of the first PCR were reamplified with primers that were internal to those used in the first PCR. All PCR reactions were conducted in a volume of 50 µl containing MgCl2 (1.5 mM), dNTPs (200 µM each), primers (0.4 µM each), and Taq DNA Polymerase (2 units). Cycling conditions were an initial 5 min denaturation at 94°C; 35 cycles (1 min at 94°C, 2 min at 50°C, 1 min at 72°C); and a final 5-min extension at 72°C. After the second (internal) PCR, amplification products were separated on a 1% agarose gel, and bands of the appropriate size were isolated with the Qiaex II agarose gel extraction kit (Qiagen, Chatsworth, CA). The purified products were directly ligated into the pGEM-T vector (Promega) and transformed into JM109 cells. After blue/white selection, positive colonies were grown in 5 ml of LB/Amp for plasmid purification. The inserts were sequenced using the fmol DNA sequencing system (Promega) with a 32P-labeled primer complementary to the T7 promoter (GTA ATA CGA CTC ACT ATA GGG C). Virtually all plasmids contained a VH-D-JH rearrangement, and their sequences were analyzed using the GCG program by comparison with known D and JH germline sequences (20, 21, 22). A minimum of four contiguous identical nucleotides was required for assignment to a germline D sequence.


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  		Table I. Primers used for amplification of murine VH-D-JH rearrangements1

 
Statistical analyses

All analyses were conducted with the Prism software (Release 2.01, GraphPad Software, San Diego, CA).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
VH-D-JH libraries

By nested PCR amplification, we generated sets of VH-D-JH sequences from newborn autoimmune MRL or control (allotype-matched) C3H mice. For each mouse strain, two sets of sequences were generated with sense primers corresponding to either J558 or 7183 VH families. We analyzed a total of 73 sequences (20 VHJ558 and 53 VH7183) from MRL mice and 68 sequences (13 VHJ558 and 55 VH7183) from C3H mice (available from GenBank under accession numbers AF071597 to AF071737). These sequences were characterized for multiple parameters such as D and JH usage, N and P nucleotide insertions, and sequence homology at the VH-D and D-JH junctions. Comparisons between the two strains are shown in Tables II to VI. The frequencies of nonproductive rearrangements were 0.38 (28/73) and 0.32 (22/68) for MRL and C3H, respectively. These values were not significantly different (using Fisher’s exact test) between both strains and were similar to those previously reported in other studies (23).

D and JH gene segment usage

The two strains differed significantly in their JH gene usage (Table IIGo, p < 0.0001). The MRL VH-D-JH rearrangements used JH4 at a greater frequency than C3H (47% vs 7%). The usage of the four D gene families (SP2, FL16, ST4, and Q52) by VH-D-JH rearrangements from MRL and C3H mice is listed in Table IIIGo. The distribution of these families significantly differed between both strains (p = 0.008), with a greater usage of Q52 in C3H mice. Within each strain, there was no significant difference in D or JH usage between productive and nonproductive rearrangements. While Q52 and ST4 are single gene families, FL16 has two members, and SP2 comprises at least nine germline genes. In previous repertoire studies conducted in BALB/c mice, the most frequently used D gene was DFL16.1 (14, 17). Although it was used more often than its DFL16.2 counterpart, DFL16.1 was found in only 4 of 68 C3H and 7 of 73 MRL sequences in the present study. Of all the VH-D-JH rearrangements using an SP2 gene, the most frequently used sequence (18 MRL and 8 C3H) was not one of the usually recognized germline genes (DSP2–2 to DSP2–10), but a sequence (cctactatAgtaactac) that differs by only one nucleotide from the DSP2.7 germline sequence (cctactatGgtaactac). This sequence is probably a bona fide germline D gene since a GenBank database search detected it in several unrelated mAbs (not shown) and in D-JH joints (24, 25). This putative germline D gene was actually named DSP2.x by Gu et al. (25). Whether DSP2.x represents an additional germline gene or an allelic variant of one of the previously identified germline genes remains to be determined. The reason for the frequent usage of DSP2.x is not known, but the nucleotide difference in this new sequence results in a serine codon (instead of a glycine) in the most frequently used reading frame. Since none of the other members of the DSP2 family possesses a serine codon in this reading frame, DSP2.x may be preferentially selected because of the versatile binding properties conferred by the serine side chain in the CDR3 (26). The relationship between DSP2.x usage and the autoimmune process is unknown, but it is worth noting that our GenBank search detected full-length DSP2.x sequences in several independently derived anti-DNA Abs (27, 28).


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  		Table II. JH germline gene usage in MRL and C3H VH-D-JH rearrangements1

 

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  		Table III. D family usage in MRL and C3H VH-D-JH rearrangements1

 
The D and JH gene usages in the present report differ somewhat from the results of previous VH-D-JH repertoire investigations (14, 17). These discrepancies result most likely from methodological differences among studies. These earlier studies have used BALB/c mice, which are of the Igh a allotype, while C3H and MRL mice are both Igh j. This allotype difference is important, as attested by the frequent usage of DSP2.x in both MRL and C3H, whereas this sequence is absent from the BALB/c repertoire. Previous studies have used different sets of PCR primers in establishing their libraries, or even generated libraries from RNA, while we utilized genomic DNA. Nevertheless, several earlier findings parallel those seen in the nonautoimmune C3H mice in the present study. Chukwuocha and Feeney examined BALB/c newborn pre-B cell genomic DNA and detected a high incidence of JH1 usage when compared with that of JH4 (19). Likewise, Gu and colleagues observed an overutilization of JH1 and DQ52 in amplified D-JH rearrangements from neonatal CB.20 pre-B cells (29).

D segments can be transcribed in all three possible reading frames, but, in genes from the SP2 and FL16 families, one of the reading frames (termed RF1) is more frequently utilized. Counterselection against the other RFs is due to the presence of stop codons in RF3 and of the transcription of a Dµ protein in RF2 leading to the premature termination of rearrangements at the H chain locus (25). RF usage in our panel of VH-D-JH rearrangements is listed in Table IVGo, showing that RF1 is the most frequently used RF in both MRL (77%) and C3H (80%). There was no significant difference in the distribution of RF usage between both strains.


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  		Table IV. Reading frame usage in MRL and C3H VH-D-JH rearrangements1

 
Because of the symmetric structure of their flanking recombination signal sequences, D segments can occasionally rearrange in an inverted orientation. The presence of two D sequences in a single VH-D-JH rearrangement (termed D-D fusion) is also possible, albeit rare. These two unusual rearrangements are often combined, in that one of the two D segments involved in a D-D fusion can also be inverted (30). Fig. 1Go depicts the D-D fusions and inverted D segments that were identified in our MRL and C3H libraries of VH-D-JH rearrangements. As stated above, the impetus for this study was to test whether newly emerging B cells from autoimmune mice more frequently possess atypical VH-D-JH rearrangements such as inverted D segments or D-D fusions. Therefore, the null hypothesis to be disproved is that the libraries of MRL and C3H have the same frequency of atypical rearrangements. The MRL library contained 11 inverted D or D-D fusions (out of 73 sequences) vs only 3 of 68 sequences in the C3H library. This difference is statistically significant (p = 0.03 with a one-sided Fisher’s exact test). Within each strain, there is no significant difference in the frequency of atypical rearrangements between productive and nonproductive rearrangements.



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  		FIGURE 1. Nucleotide sequences of atypical VH-D-JH rearrangements from neonate MRL and C3H mice. Atypical rearrangements are defined as inverted D and/or D-D fusions. Bars beneath the sequences indicate homology with known germline D genes with mismatches indicated by interruptions in these horizontal bars. (INV), Indicates a reverse complement sequence. Lower case letters, Indicate regions of homology between the D segment and the VH or JH genes. Italics, Indicate P nucleotides. The distribution of atypical rearrangements was significantly different (p = 0.03 with Fisher’s exact test) between MRL (11/73) and C3H (3/68) strains.

 
Many atypical rearrangements observed in MRL and C3H mice in this study involve the inversion of the DSP2.x gene. Since the coding sequences of the DSP2 family display partial internal symmetry, we considered the possibility that these rearrangements may not result from an actual inversion but that this sequence is a hitherto unknown DSP2 allele present in the MRL and C3H strains. To test this possibility, we searched the GenBank database for the inverted DSP2.x sequence, and we found only two matches in VH-D-JH rearrangements obtained from B-less mice that express a human {lambda} transgene (31). Both rearrangements were D-D fusions where one of the D segments is frequently rearranged in inverted orientation. Our GenBank search did not find this sequence in any conventional VH-D-JH rearrangement, nor in any reported D-JH joint. In contrast, a similar GenBank search with the DSP2.x sequence in standard orientation discovered many uses of this sequence in D-JH joints and VH-D-JH rearrangements from independently derived mAbs (24, 25, 27, 28, 32, 33, 34, 35, 36). Therefore, the most logical interpretation of our findings is that these sequences do indeed contain the DSP2.x gene in inverted orientation and that atypical rearrangements are more frequent in the MRL neonatal repertoire than in C3H mice.

Junctional diversity

Homology-based VH-D or D-JH recombinations, i.e., junctions that are identical to both the 3' end of the upstream and the 5' end of the downstream germline sequences, are frequent in the fetal and neonatal repertoires (18). We indeed observed that the majority of our sequences contain regions of junctional homology and that the frequency of sequences possessing homologies is not significantly different between the MRL (71%) and C3H (74%) strains (Table VGo).


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  		Table V. Frequency of MRL and C3H VH-D-JH rearrangements with homologous junctions and N or P additions1

 
Because of the lack of terminal deoxynucleotidyl transferase expression during early development, N nucleotide additions are less frequent in the immature than in the adult repertoire (14, 17). As expected for libraries of neonatal sequences, only a minority of our VH-D-JH sequences (29% for MRL and 34% for C3H) contained N nucleotide insertions. The average number of N nucleotides per VH-D-JH sequence was similar for both strains, 0.74 for MRL and 0.81 for C3H. Few sequences from either strain (7 for MRL and 9 for C3H) contained P nucleotide insertions. None of these values were significantly different between both strains.

Deduced amino acid sequences from productive rearrangements

We examined the deduced protein sequences of the 45 MRL and 46 C3H productive rearrangements. These sequences were overall similar in both strains and are listed in Fig. 2Go. The average residue lengths of the CDR3 were 9.87 for MRL and 9.61 for C3H (not significantly different). VH-D-JH rearrangements from both strains contained arginine and asparagine residues at similar frequencies: 0.04 Arg and 0.31 Asn/CDR3 for MRL and 0.04 arg and 0.26 Asn/CDR3 for C3H. These values were not statistically different between both strains (Table VIGo).



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  		FIGURE 2. Deduced amino acid sequences from productive VH-D-JH rearrangements in MRL (A) and C3H (B) mice. Sequences are displayed from the second invariant VH cysteine through the first residue after the invariant JH tryptophan. Nonproductive joints are not listed.

 

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  		Table VI. Characteristics of the amino acid sequences deduced from productive VH-D-JH rearrangements1

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
We have analyzed libraries of VH-D-JH junctions from autoimmune MRL and Igh allotype-matched control C3H newborn mice. Our results indicate that rearrangements generated from MRL mice possess some of the characteristics that are observed in the adult autoimmune repertoire. In adult autoimmunity, features such as atypical VH-D-JH rearrangements (D-D fusions and inverted D segments) are associated with an increase in the frequency of arginine residues in the CDR3 of anti-DNA and anti-nucleosome Abs (5, 6, 7, 9, 10, 11, 12). As expected and in support of the legitimacy of these differences between C3H and MRL mice, most of the other parameters that we analyzed (such as N nucleotide frequency or frequency of homology-containing sequences) were not significantly different between both strains. Therefore, our results indicate that atypical VH-D-JH rearrangements occur more frequently in the early repertoire of autoimmune MRL mice than in normal animals. These neonatal B cells with atypical VH-D-JH rearrangements are, however, unlikely to be the precursors of autoantibody-producing cells in autoimmune animals. Most anti-DNA or anti-nucleosome Abs possess features such as N nucleotide additions or arginine residues in their H chain CDR3s that are not present in our library of VH-D-JH rearrangements. Thus, although most autoreactive B cells originate from the adult repertoire, the propensity to produce atypical VH-D-JH junctions is already present in newborn autoimmune mice.

C3H and MRL VH-D-JH rearrangements differ in their D segment and JH gene usage. C3H junctions utilize DQ52 in 19 of 65 conventional rearrangements, vs 5 of 62 for MRL. Conversely, MRL junctions use JH4 in 30 of 62 conventional rearrangements, vs 5 of 65 for C3H. DQ52, the most 3' D segment, is overrepresented in primary D-JH rearrangements, presumably because of its proximity to the JH locus (14). The lesser use of DQ52 and the increased utilization of JH4 in MRL mice suggest a greater frequency of secondary D-JH rearrangements in this strain. Such rearrangements use an upstream D sequence and a downstream JH gene to delete a previous D-JH rearrangement. Secondary D-JH rearrangements are frequent in transformed pre-B cell lines (37), but are thought to be uncommon in normal mouse repertoire (17, 38). If the dissimilar use of DQ52 and JH4 between MRL and C3H is due to an increased frequency of secondary D-JH rearrangements in MRL mice, then DQ52-JH4 joints must be rare. This is indeed the case since, in C3H mice, none of the 19 DQ52-containing rearrangements involved JH4, whereas, in MRL mice, only one of the 30 JH4+ joints involved DQ52. Therefore, in both strains, DQ52 is more likely to rearrange with one of the JH genes that is located upstream of JH4 and JH4 is more likely to be associated with one of the D genes that are 5' of DQ52.

Our observation supports the view that B cells from autoimmune individuals or mice manifest intrinsic defects that contribute to the self reactive process (39, 40). There is additional evidence that VH-D-JH rearrangements are intrinsically different in autoimmune mice. Alarcón-Riquelme and Fernández have examined the sequences of the H chain CDR3s from a panel of small resting C57BL6/lpr B cells (41). They observed that a majority of these "preimmune" rearrangements involved D segments read in RF2 and RF3, and they proposed that this was the result of a defect in the selection mechanisms acting independently of, and earlier than, Ag selection (41). Nevertheless, the high density B cells used by these authors were obtained from 5-mo old animals, and Ag selection could not be entirely ruled out. We have characterized a panel of several polyreactive IgM mAbs from mice belonging to various spontaneously autoimmune strains (42). In contrast to most conventional Abs that use RF1, the D segments of these polyreactive Abs made use of all three possible RFs. Since none of the polyreactive mAbs in our panel reacted with DNA, this unusual RF usage was not selected by stimulation with DNA as Ag. In our present study, RF usage was similar in autoimmune and control newborn mice, suggesting that these RF differences may manifest themselves only in adult animals.

The differences observed between MRL and C3H are not the mere result of a defect in Fas-mediated selection since we used MRL +/+ mice that possess an intact Fas gene contrary to their MRL-lpr/lpr counterparts. The mechanisms that favor the presence of atypical VH-D-JH rearrangements can only be speculative at this stage. These could result from differences in the recombination machinery itself or from selection events that take place after the association of the H chain with the surrogate light chain or with a functionally rearranged {kappa} or {lambda} light chain (43, 44). Other investigators have suggested that the properties of H chain rearrangements influence B cell development (39). In birds, Pandey et al. suggested that B cells undergo prenatal developmental selection before exposure to Ag through a mechanism involving interaction between the protein product resulting from V(D)J recombination and endogenous ligands (45). If indeed V(D)J products are selected before Ag exposure, it is conceivable that an anomaly in this selection process may be one of the defects responsible for the development of systemic autoimmunity.

The precise sequence of events that result in a VH-D-D-JH rearrangement (D-D fusion) has yet to be established with certainty. Since recombination signal sequences with 12-bp spacers flank D segments on both sides, D-D fusions violate the 12/23 rule. They can presumably occur because of the recognition of an alternate recombination signal sequence (with a 24-bp spacer) created by a heptamer-like sequence within the coding region of certain D genes (46). It has been shown that two unrearranged D segments can directly join (30), but it is also possible for an upstream D to rearrange with a preexisting D-JH joint. The latter process is similar to a partial secondary D-JH rearrangement, where the recombinase machinery joins an upstream D to a previous D-JH rearrangement, rather than to a downstream JH. That this phenomenon is relevant to autoimmunity is suggested by the frequent usage of JH4 in MRL autoantibodies, such as anti-chromatin Abs and rheumatoid factors (11, 12, 47, 48). It is unlikely that JH4+ clones are selected for their binding properties since all four germline JH genes are similar in sequence and their contribution to Ab diversity and specificity is limited. Therefore, in MRL mice, the same mechanisms may result in an increased frequency of atypical VH-D-JH joints and of secondary D-JH rearrangements.

Although atypical VH-D-JH are present in many anti-DNA Abs characterized from lupus-prone mice (13), they are not a universal feature of induced anti-DNA Abs. For instance, a panel of high affinity IgG anti-DNA mAbs obtained from BALB/c mice after various regimens of protein Ag immunizations possessed conventional VH-D-JH junctions (49). Recently, Ash-Lerner et al. compared the structures of anti-DNA mAbs that use the BW-16 VH gene, a germline gene associated with the autoimmune response in lupus-prone mice (50). Whereas BW-16+ anti-DNA mAbs from NZB x NZW/F1 mice displayed complex rearrangement patterns with D-D fusions and alternate RFs, BW-16+ anti-DNA mAbs from C3H mice immunized with DNA complexed to an immunogenic peptide possessed "standard" VH-D-JH rearrangements (50). Therefore, the immune system is capable of generating anti-DNA Abs from a variety of B cell precursors. That it elects to use B cells with atypical VH-D-JH junctions in autoimmune mice may simply result from the increased availability of such rearrangements in lupus-prone strains.

One may therefore wonder whether a difference in the number of B cell precursors could affect the level of the Ab response or if this difference would be irrelevant because of the magnitude of Ag-driven clonal expansion. There is experimental evidence that B cell precursor frequency affects the amplitude of the Ab response against the corresponding self Ag. BALB/c mice have a greater frequency of preimmune B cells self reactive to mouse cytochrome c than C57BL/6 mice (51). This difference persists during the secondary response to mouse cytochrome c-OVA where BALB/c still have three times more B cells specific for murine cytochrome c than do C57BL/6 mice (51).

In summary, our data suggest that lupus-prone MRL mice possess an intrinsic defect that favors the production of atypical VH-D-JH joints and secondary D-JH rearrangements in newborn animals. It will be critical to assess in future experiments whether similar differences exist between autoimmune and normal mice in the adult pre-B cell repertoire.


   Acknowledgments
 
We are grateful to Dr. Martin Weigert for his insightful comments and to Mike Fowler for his technical assistance.


   Footnotes
 
1 This work was supported by National Institutes of Health Grant AI-26665 and by a Sheryl N. Hirsch Award from the Lupus Foundation of Philadelphia. Back

2 Address correspondence and reprint requests to Dr. Marc Monestier, Department of Microbiology and Immunology, Temple University School of Medicine, 3400 North Broad Street, Philadelphia, PA 19140. E-mail address: Back

3 Abbreviations used in this paper: CDR, complementarity determining region; MRL, MRL/MpJ +/+; C3H, C3H/HeJ; RF, reading frame. Back

Received for publication July 13, 1998. Accepted for publication October 27, 1998.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Pisetsky, D. S.. 1992. Anti-DNA antibodies in systemic lupus erythematosus. Rheum. Dis. Clin. North Am. 18:437.[Medline]
  2. Stollar, B. D.. 1994. Molecular analysis of anti-DNA antibodies. FASEB J. 8:337.[Abstract]
  3. Radic, M. Z., M. Weigert. 1994. Genetic and structural evidence for antigen selection of anti-DNA antibodies. Annu. Rev. Immunol. 12:487.[Medline]
  4. Monestier, M.. 1997. Autoantibodies to nucleosomes and histone-DNA complexes. Methods 11:36.[Medline]
  5. Eilat, D., R. Fischel. 1991. Recurrent utilization of genetic elements in V regions of antinucleic acid antibodies from autoimmune mice. J. Immunol. 147:361.[Abstract]
  6. Tillman, D. M., N. T. Jou, R. J. Hill, T. N. Marion. 1992. Both IgM and IgG anti-DNA antibodies are the products of clonally selective B cell stimulation in (NZB x NZW)F1 mice. J. Exp. Med. 176:761.[Abstract/Free Full Text]
  7. Shlomchik, M., M. Mascelli, H. Shan, M. Z. Radic, D. Pisetsky, A. Marshak-Rothstein, M. Weigert. 1990. Anti-DNA antibodies from autoimmune mice arise by clonal expansion and somatic mutation. J. Exp. Med. 171:265.[Abstract/Free Full Text]
  8. Radic, M. Z., J. Mackle, J. Erikson, C. Mol, W. F. Anderson, M. Weigert. 1993. Residues that mediate DNA binding of autoimmune antibodies. J. Immunol. 150:4966.[Abstract]
  9. Monestier, M., K. E. Novick. 1996. Specificities and genetic characteristics of nucleosome-reactive antibodies from autoimmune mice. Mol. Immunol. 33:89.[Medline]
  10. Monestier, M., K. E. Novick, M. J. Losman. 1994. D-penicillamine- and quinidine-induced antinuclear antibodies in A.SW (H-2S) mice: similarities with autoantibodies in spontaneous and heavy metal-induced autoimmunity. Eur. J. Immunol. 24:723.[Medline]
  11. Losman, M. J., T. M. Fasy, K. E. Novick, M. Monestier. 1993. Relationships among antinuclear antibodies from autoimmune MRL mice reacting with histone H2A-H2B dimers and DNA. Int. Immunol. 5:513.[Abstract/Free Full Text]
  12. Losman, M. J., T. M. Fasy, K. E. Novick, M. Monestier. 1992. Monoclonal autoantibodies to subnucleosomes from a MRL/Mp +/+ mouse: oligoclonality of the antibody response and recognition of a determinant composed of histones H2A, H2B, and DNA. J. Immunol. 148:1561.[Abstract]
  13. Radic, M. Z., M. A. Mascelli, J. Erikson, H. Shan, M. Shlomchik, M. Weigert. 1989. Structural patterns in anti-DNA antibodies from MRL/lpr mice. Cold Spring Harbor Symp. Quant. Biol. 54:(Pt 2):933.
  14. Bangs, L. A., I. E. Sanz, J. M. Teale. 1991. Comparison of D, JH, and junctional diversity in the fetal, adult, and aged B cell repertoires. J. Immunol. 146:1996.[Abstract]
  15. Shlomchik, M. J., A. Marshak-Rothstein, C. B. Wolfowicz, T. L. Rothstein, M. G. Weigert. 1987. The role of clonal selection and somatic mutation in autoimmunity. Nature 328:805.[Medline]
  16. Theofilopoulos, A. N., R. Kofler, P. Singer, F. J. Dixon. 1989. Molecular genetics of murine lupus models. Adv. Immunol. 46:61.[Medline]
  17. Feeney, A. J.. 1990. Lack of N regions in fetal and neonatal mouse immunoglobulin V-D-J junctional sequences. J. Exp. Med. 172:1377.[Abstract/Free Full Text]
  18. Feeney, A. J.. 1992. Predominance of VH-D-JH junctions occurring at sites of short sequence homology results in limited junctional diversity in neonatal antibodies. J. Immunol. 149:222.[Abstract]
  19. Chukwuocha, R. U., A. J. Feeney. 1993. Role of homology-directed recombination: predominantly productive rearrangements of Vh81X in newborns but not in adults. Mol. Immunol. 30:1473.[Medline]
  20. Devereux, J., P. Haeberli, O. Smithies. 1984. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 12:387.
  21. Kabat, E. A., T. T. Wu, H. M. Perry, K. S. Gottesman, C. Foeller. 1991. Sequences of Proteins of Immunological Interest 1. U.S. Government Printing Office, Bethesda, MD.
  22. Feeney, A. J., R. Riblet. 1993. DST4: a new, and probably the last, functional DH gene in the BALB/c mouse. Immunogenetics 37:217.[Medline]
  23. Carlsson, L., C. Overmo, D. Holmberg. 1992. Selection against N-region diversity in immunoglobulin heavy chain variable regions during the development of pre-immune B cell repertoires. Int. Immunol. 4:549.[Abstract/Free Full Text]
  24. Tarlinton, D., A. M. Stall, L. A. Herzenberg. 1988. Repetitive usage of immunoglobulin VH and D gene segments in CD5+ Ly-1 B clones of (NZB x NZW)F1 mice. EMBO J. 7:3705.[Medline]
  25. Gu, H., D. Kitamura, K. Rajewsky. 1991. B cell development regulated by gene rearrangement: arrest of maturation by membrane-bound Dm protein and selection of DH element reading frames. Cell 65:47.[Medline]
  26. Lea, S., D. Stuart. 1995. Analysis of antigenic surfaces of proteins. FASEB J. 9:87.[Abstract]
  27. Staines, N. A., F. J. Ward, A. N. Denbury, J. Mitchiner, O. Hartley, D. Eilat, D. A. Isenberg, S. Bansal. 1993. Primary sequence and location of the idiotopes of V-88, a DNA- binding monoclonal autoantibody, determined by idiotope scanning with synthetic peptides on pins. Immunology 78:371.[Medline]
  28. Krishnan, M. R., N. T. Jou, T. N. Marion. 1996. Correlation between the amino acid position of arginine in VH-CDR3 and specificity for native DNA among autoimmune antibodies. J. Immunol. 157:2430.[Abstract]
  29. Gu, H., I. Förster, K. Rajewsky. 1990. Sequence homologies, N sequence insertion and JH gene utilization in VHDJH joining: implications for the joining mechanism and the ontogenic timing of Ly 1 B cell and B-CLL progenitor generation. EMBO J. 9:2133.[Medline]
  30. Meek, K. D., C. A. Hasemann, J. D. Capra. 1989. Novel rearrangements at the immunoglobulin D locus: inversions and fusions add to IgH somatic diversity. J. Exp. Med. 170:39.[Abstract/Free Full Text]
  31. Levinson, D. A., J. Campos-Torres, P. Leder. 1993. Molecular characterization of transgene-induced immunodeficiency in B-less mice using a novel quantitative limiting dilution polymerase chain reaction method. J. Exp. Med. 178:317.[Abstract/Free Full Text]
  32. Perlmutter, R. M., J. F. Kearney, S. P. Chang, L. E. Hood. 1985. Developmentally controlled expression of immunoglobulin VH genes. Science 227:1597.[Abstract/Free Full Text]
  33. Mo, J. A., R. Holmdahl. 1996. The B cell response to autologous type II collagen: biased V gene repertoire with V gene sharing and epitope shift. J. Immunol. 157:2440.[Abstract]
  34. Weller, S., C. Conde, A. M. Knapp, H. Levallois, S. Gilfillan, J. L. Pasquali, T. Martin. 1997. Autoantibodies in mice lacking terminal deoxynucleotidyl transferase: evidence for a role of N region addition in the polyreactivity and in the affinities of anti-DNA antibodies. J. Immunol. 159:3890.[Abstract]
  35. Takahashi, Y., P. R. Dutta, D. M. Cerasoli, G. Kelsoe. 1998. In situ studies of the primary immune response to (4-hydroxy-3-nitrophenyl)acetyl. V. Affinity maturation develops in two stages of clonal selection. J. Exp. Med. 187:885.[Abstract/Free Full Text]
  36. Schittek, B., K. Rajewsky. 1992. Natural occurrence and origin of somatically mutated memory B cells in mice. J. Exp. Med. 176:427.[Abstract/Free Full Text]
  37. Reth, M. G., S. Jackson, F. W. Alt. 1986. VHDJH formation and DJH replacement during pre-B differentiation: non-random usage of gene segments. EMBO J. 5:2131.[Medline]
  38. Chang, Y., C. J. Paige, G. E. Wu. 1992. Enumeration and characterization of DJH structures in mouse fetal liver. EMBO J. 11:1891.[Medline]
  39. Schwartz, R. S., B. D. Stollar. 1994. Heavy-chain directed B-cell maturation: continuous clonal selection beginning at the pre-B cell stage. Immunol. Today 15:27.[Medline]
  40. Kakkanaiah, V. N., E. S. Sobel, G. C. MacDonald, R. L. Cheek, P. L. Cohen, R. A. Eisenberg. 1997. B cell genotype determines the fine specificity of autoantibody in lpr mice. J. Immunol. 159:1027.[Abstract]
  41. Alarcon-Riquelme, M. E., C. Fernandez. 1995. CDR3 regions in the preimmune VH B cell repertoire of lpr mice. Clin. Exp. Immunol. 101:73.[Medline]
  42. Novick, K. E., T. M. Fasy, M. J. Losman, M. Monestier. 1992. Polyreactive IgM antibodies generated from autoimmune mice and selected for histone-binding activity. Int. Immunol. 4:1103.[Abstract/Free Full Text]
  43. Decker, D. J., G. H. Kline, T. A. Hayden, S. N. Zaharevitz, N. R. Klinman. 1995. Heavy chain V gene-specific elimination of B cells during the pre-B cell to B cell transition. J. Immunol. 154:4924.[Abstract]
  44. Wasserman, R., Y.-S. Li, S. A. Shinton, C. E. Carmack, T. Manser, D. L. Wiest, K. Hayakawa, R. R. Hardy. 1998. A novel mechanism for B cell repertoire maturation based on response by B cell precursors to pre-B receptor assembly. J. Exp. Med. 187:264.
  45. Pandey, A., L. W. Tjoelker, C. B. Thompson. 1993. Restricted immunoglobulin junctional diversity in neonatal B cells results from developmental selection rather than homology-based V(D)J joining. J. Exp. Med. 177:329.[Abstract/Free Full Text]
  46. Kurosawa, Y., S. Tonegawa. 1982. Organization, structure, and assembly of immunoglobulin heavy chain diversity DNA segments. J. Exp. Med. 155:201.[Abstract/Free Full Text]
  47. Monestier, M.. 1991. Variable region genes of anti-histone autoantibodies from a MRL/Mp-lpr/lpr mouse. Eur. J. Immunol. 21:1725.[Medline]
  48. Duchosal, M. A., R. Kofler, R. S. Balderas, M. T. Aguado, F. J. Dixon, A. N. Theofilopoulos. 1989. Genetic diversity of murine rheumatoid factors. J. Immunol. 142:1737.[Abstract]
  49. Shefner, R., G. Kleiner, A. Turken, L. Papazian, B. Diamond. 1991. A novel class of anti-DNA antibodies identified in BALB/c mice. J. Exp. Med. 173:287.[Abstract/Free Full Text]
  50. Ash-Lerner, A., M. Ginsberg-Strauss, Y. Pewner-Jung, D. D. Desai, T. N. Marion, D. Eilat. 1997. Expression of an anti-DNA associated VH gene in immunized and autoimmune mice. J. Immunol. 159:1508.[Abstract]
  51. Mueller, C. M., J. M. Minnerath, R. Jemmerson. 1997. B lymphocyte recognition of the self antigen mouse cytochrome c in different mouse strains: targeting of the same dominant epitope by naturally-occurring cells expressing distinct VH genes. Mol. Immunol. 34:843.[Medline]



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