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P D Gottlieb

Publications and source records attributed to P D Gottlieb.

At least 19 recordsLinked to original sources
PubMed1992

Inducible functions in hybrids of a Lyt-2+ BW5147 transfectant and the 2C CTL line.

Cytolytic activity and release of interleukin 2 (IL-2) were induced in Lyt-2-positive T-T cell hybrids by incubation with either concanavalin A or irradiated stimulator cells. Since hybrids of Lyt-2-positive class I-specific cytotoxic T lymphocytes (CTLs) with the fusable mouse thymoma cell line, BW5147, are invariably Lyt-2-negative, a derivative of BW5147 was produced by transfection which constitutively expresses surface Lyt-2.1. This cell line, 3B2, was fused with the H-2Ld-specific long term CTL line, 2C. Such hybrids expressed the transfected Lyt-2 gene but not the endogenous gene of the 2C fusion partner. That Lyt-2 plays a functional role in hybrids of 3B2 with 2C is shown by the observations that: 1) cytolysis by Lyt-2-positive hybrids was inhibited by Lyt-2-specific monoclonal antibody (mAb); 2) Lyt-2-positive but not Lyt-2-negative subclones of one such line develop specific cytotoxicity when incubated with stimulator cells; 3) Less IL-2 was released from Lyt-2-negative subclones incubated with stimulator cells than from Lyt-2-positive subclones; 4) Lyt-2-specific mAb inhibits release of IL-2 from Lyt-2-positive hybrids incubated with stimulator cells. All Lyt-2-positive hybrids expressed functional surface Lyt-3 encoded by the CTL fusion partner, demonstrating that expression of the Lyt-3 gene is not sensitive to the negative regulation which shuts off the endogenous Lyt-2 gene in hybrids of class I-specific CTLs with the 3B2 or BW5147 cell lines. The existence of inducible T-T cell hybrids expressing functional Lyt-2 and Lyt-3 provides a system for evaluation of the role(s) of Lyt-2 and Lyt-3 in the induction of function independent of cell growth.

Animals↗
PubMed1992

A self-reactive T cell population that is not subject to negative selection.

In male mice expressing a transgenic alpha beta TCR which recognizes a male antigen (HY), T cells which do not express normal levels of CD8 escape thymic deletion and appear in the periphery. These consist of two distinct populations, one which lacks expression of both CD4 and CD8, and one with low levels of CD8. Neither population has anti-HY reactivity, consistent with the known requirement of this TCR for CD8. We now describe the consequences of expression of both the anti-HY TCR transgene and a constitutive CD8.1 transgene on T cells of male mice. Peripheral T cells in these male 'double transgenic' mice express both the anti-HY TCR and normal levels of CD8, and can proliferate to male antigen in vitro. These cells do not express the endogenous allele of CD8 (CD8.2), suggesting that the increase in CD8 levels due to the CD8.1 transgene leads to the deletion of the CD8.2low population. In contrast, the CD8.1 transgene does not lead to the deletion of the CD8.2- population. This implies that, unlike the majority of alpha beta T cells, TCR+CD4-CD8- cells in TCR transgenic mice are not subject to deletion.

Animals↗
PubMed1991

Polymorphism in V kappa 10 genes encoding L chains of antibodies bearing the Ars-A and A48 cross-reactive idiotypes.

p-azophenylarsonate-specific antibodies of A/J mice which bear the Ars-A crossreactive idiotype utilize the V kappa-Ars-A gene segment, a member of the V kappa 10 family. Southern hybridization of genomic DNA from several inbred strains using a probe from the 5' flanking region of the V kappa-Ars-A gene demonstrated three patterns of restriction fragment length polymorphisms (RFLP). Six genes corresponding to hybridizing bands were obtained from DNA libraries of C.AKR, PERU and A/J mice, and nucleotide sequence comparisons revealed two allelic groups: AKR1 (Igk-V10.1a), AJ1 (Igk-V10.1b) and PERU1 (Igk-V10.1c); AKR2 (Igk-V10.2a), AJ2 (Igk-V10.2b), and PERU2 (Igk-V10.2c). The Igk-V10.1b gene of the A/J strain is the V kappa-Ars-A gene used in Ars-A idiotype-positive antibodies. The product of the C.AKR allele (Igk-V10.1a) contained four amino acid substitutions in CDR3 as compared with Igk-V10.1b. These substitutions probably explain the failure of AKR mice and other strains with the same V kappa 10 RFLP pattern to provide in genetic crosses a L chain which, together with the A/J VH-ArsA gene product, form Ars-A idiotype-positive antibodies. Also, the nucleotide sequence identity between the Igk-V10.1c and Igk-V10.1b alleles and the Igk-V10.2c and Igk-V10.2b alleles is significantly greater than that seen in comparisons with the Igk-V10.1a and Igk-V10.2a alleles, respectively, suggesting an evolutionary pathway similar to that of the linked Igk-J locus. BALB/c antibodies bearing the A48 regulatory idiotype contain L chains encoded by the BALB/c Igk-V10.1b and Igk-V10.2b alleles. Strongly A48 idiotype-positive antibodies utilize the Igk-V10.1b chain, and weakly A48-positive antibodies use the Igk-V10.2b L chain. The possible effects of amino acid substitutions specified by the Igk-V10.1a, Igk-V10.1c, Igk-V10.2a, and Igk-V10.2c alleles on their ability to provide L chains used in A48 idiotype-positive antibodies are discussed.

Alleles↗
PubMed1989

Lymphomas with acquired mouse mammary tumor virus proviruses resemble distinct prethymic and intrathymic phenotypes defined in vivo.

A number of murine T cell lymphomas expressing the T cell Ag Thy-1 contain acquired mouse mammary tumor (MMTV) proviruses. These lymphomas all express detectable levels of MMTV RNA, yet the majority of the tumors fail to produce MMTV particles. To determine if the ability of lymphomas to produce MMTV is a reflection of the differentiation state of the tumor, we examined eight lymphomas for expression of surface B and T cell Ag as well as for rearrangements and expression of TCR genes. All tumors could be grouped into three categories observed in vivo, including early lymphoid, nonmature intrathymic T cells, and immature intrathymic T cells. Cell lines corresponding to all three phenotypes produced MMTV particles, suggesting that production of virus is not linked to the differentiation state of lymphoid cells. These studies highlight the potential advantage of studying T cell lymphomas vs mixed primary populations or T cell hybridomas for evaluation of both phenotypic and molecular markers in clonal T cells.

Animals↗
PubMed1989

Structural and evolutionary comparisons of four alleles of the mouse Igk-J locus which encodes immunoglobulin kappa light chain joining (J kappa) segments.

The Igk-J locus of the mouse encodes the immunoglobulin kappa light chain joining (J) segments. Four Igk-J alleles have been described on the basis of restriction enzyme length polymorphisms. The nucleotide sequences of the Igk-Ja allele (type strain, C.C58), Igk-Jc allele (type strain, SJL/J), and Igk-Jd allele (type strain, SK/CamRk) have been determined and are compared with the previously reported Igk-Jb allele sequence (type strain, BALB/c). The mouse sequences are also compared with published sequences for rat and human J kappa sequences. Far more differences were found between the Igk-Ja allele and the other mouse alleles than between any two of the latter. These result in two amino acid substitutions which distinguish the J2 and J3' segments of the Igk-Ja allele from the other three alleles. Use of the Phylogenetic Analysis Using Parsimony program to generate a phylogenetic tree strongly indicates that after divergence from the rat ancestor, there appears to have been an early split between the Igk-Ja allele and the evolutionary precursor of the other mouse alleles. There also appears to have been far less divergence from the ancestral condition in the Igk-Ja allele than in the other alleles. Also, the presence of only one convergent mutation among the four mouse alleles provides strong evidence against any crossing over within the Igk-J locus during the history of these alleles. Finally, the differences in rates of evolution of the Igk-J alleles are in marked contrast to the relatively uniform rates of divergence of four alleles of a mouse V kappa gene, Igk-VSer.

Alleles↗
PubMed1989

Structural and evolutionary comparisons of four alleles of the mouse immunoglobulin kappa chain gene, Igk-VSer.

The mouse Igk-VSer gene encodes an immunoglobulin kappa light chain variable region which gives rise to two phenotypic polymorphisms of mouse kappa chains. The nucleotide sequences of coding and flanking regions of the Igk-VSerc and Igk-VSerd alleles found in recently inbred strains of wild mice are compared with those of the Igk-VSera and Igk-VSerb alleles described previously. Results suggest that the gene is evolving randomly and that framework 2 and complementarity determining region 2 are preserved, presumably for overall light chain structure. Results indicate that all four alleles have an octamer motif upstream of the gene which should be functional and allow prediction of whether or not the product of the germ line gene will be detectable as either the IB-peptide or Ef1a phenotypic polymorphism. Southern hybridization of genomic DNA using as probe a 1-kb Xba I-Xba I fragment located approximately 4 kb upstream of the BALB/c Igk-VSerb coding region demonstrated the presence of homologous DNA in mice bearing the Igk-VSera allele and absence from mice bearing the Igk-VSerc and Igk-VSerd alleles. Nucleotide sequence comparison of BALB/c and SK/CamRk (Igk-VSerd) DNA in this region demonstrated that BALB/c contained an insertion 2.4 kb in length which was absent from SK/CamRk. Both strains contain DNA homologous to the reverse complement of the mouse Bam5 repetitive element at the point of the insertion, with BALB/c containing approximately 70 nucleotides more of the element than SK/CamRk. Surprisingly, the strains containing DNA related to the Xba I-Xba I probe are not those determined to be the most similar by nucleotide sequence comparisons and by the Phylogenetic Analysis Using Parsimony program. The evolutionary relationship of the alleles and a possible basis for the inconsistency presented by the Xba I-Xba I fragment-related DNA are discussed.

Alleles↗
PubMed1988

Analysis of the effect of a strain-specific alteration in the upstream octamer region on transcription of a mouse V kappa Ser light chain gene.

Previous studies had shown that several phenotypic markers expressed by several strains of mice (C58, AKR, PL, RF) involve a light chain group, called V kappa Ser, encoded by a single germ-line gene (Igk-VSera). Most other inbred strains (e.g., BALB/c) contain an allele (Igk-VSerb) which differs from Igk-VSera both in coding regions and in an upstream octamer region known to be important for transcription. Since no evidence for expression of the Igk-VSerb gene product has been observed, experiments were undertaken to determine whether the alteration in the regulatory octamer region of BALB/c might have rendered it defective for transcription. The upstream octamer-containing region of a cloned functional V kappa Ser gene expressed by the C.C58 myeloma M75 was replaced by the corresponding region from BALB/c or deleted entirely. Constructs were transfected into J558L cells and quantity of transcription and site of transcription initiation were compared. Results suggest that the BALB/c octamer (CTTTGCGT), which differs at two of eight nucleotides from the consensus octamer sequence (ATTTGCAT), is fully functional in transcription initiation. This is consistent with results of S1 nuclease protection experiments which indicate the presence of small amounts of correctly initiated V kappa Ser-related RNA in BALB/c spleen.

Animals↗
PubMed1988

Nucleotide sequence analysis of the C.AKR Lyt-2a gene: structural polymorphism in alleles encoding the Lyt-2.1 T-cell surface alloantigen.

The Lyt-2a allele of the C.AKR strain of mice (genotype Lyt-2a, Lyt-3a) was cloned, and its complete nucleotide sequence as well as that of 2 kb of 5' flanking DNA was determined. The sequence was compared with the partial sequence of the Lyt-2a allele of DBA/2 (genotype Lyt-2a, Lyt-3b) and the nearly complete sequence of the B10.CAS2 Lyt-2b allele reported by Liaw and coworkers (1986). The coding regions of the two Lyt-2a alleles differ from each other by two nucleotide substitutions in the three exons over which they could be compared, resulting in two amino acid substitutions in the leader and transmembrane segments. The coding region of the C.AKR Lyt-2a allele differs from the Lyt-2b allele by two nucleotide substitutions in the extracellular V-like domain, one of which is silent and the second of which leads to substitution of valine for methionine at amino acid position 78 giving rise to the Lyt-2.1 allotypic specificity. The coding region of the DBA/2 Lyt-2a allele shares with C.AKR the allotypic substitution at position 78 and differs from Lyt-2b by three additional nucleotide substitutions in the coding regions, two of which lead to amino acid substitutions in the leader and transmembrane segments. It would therefore appear that the Lyt-2 alleles of the three strains analyzed are distinct, and the nomenclature Lyt-2a1 and Lyt-2a2 is suggested to distinguish the alleles of C.AKR and DBA/2, respectively. These alleles share a common difference from the Lyt-2b gene product at position 78, and since the amino acid substitutions which distinguish them from each other are in the leader and transmembrane segments, their mature Lyt-2 gene products appear antigenically identical.

Alleles↗
PubMed1988

Structure and expression of the Lyt-3a gene of C.AKR mice.

The mouse Lyt-3a gene, which encodes the Lyt-3.1 T-cell surface alloantigen of the C.AKR strain, has been cloned, and the nucleotide sequence of its exons and more than 2 kb of 5' flanking sequence have been determined. The gene extends over approximately 16 kb of DNA and consists of six exons encoding leader, leader plus V-like domain, membrane-proximal, transmembrane, and cytoplasmic domains. The only difference between the coding region of the Lyt-3a gene and the cDNA sequences reported for Lyt-3b (Nakauchi et al. 1987. Panaccio et al. 1987) is at position 77 of the mature protein where Lyt-3a encodes serine and Lyt-3b encodes arginine. This substitution must therefore be the basis for the serological distinction between the Lyt-3.1 and Lyt-3.2 alloantigens. Potential TATA and CAAT sequences, two Sp1 protein binding sites, two extended repeats of the dinucleotide, CA, a number of short inverted repeats, and an inverted segment of the mouse B1 repetitive sequence are found 5' to the Lyt-3a gene. Two consensus poly-A addition signals and a complete copy of the mouse B1 sequence are found 3' to the gene. Both B1-related regions are flanked by short direct repeats suggesting that they arose by an insertional mechanism. Cotransfection of the Lyt-3a gene together with a cloned Lyt-2a gene resulted in expression of both Lyt-2 and Lyt-3.1 on the surface of Ltk- and BW5147 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Alleles↗
PubMed1987

The endogenous retrovirus Mtv-8 on mouse chromosome 6 maps near several kappa light chain markers.

Endogenous retroviruses are known to affect expression of cellular genes in the vicinity of their integration sites. The endogenous mouse mammary tumor provirus (Mtv-8) previously has been reported to reside on mouse chromosome 6 near the immunoglobulin kappa chain locus. Using pairs of mouse strains on the BALB/c (Mtv-8 positive) and C58 (Mtv-8 negative) backgrounds which are congenic for chromosome 6 genetic markers, we have confirmed the chromosome assignment of this provirus. Moreover, we have analyzed the N1 progeny of a (B6 X C58) X C58 backcross to determine the segregation of the Mtv-8 provirus with respect to polymorphisms in the Igk-VSer and Igk-J loci. The results with congenic and backcross mice together with results of others suggest that Mtv-8 is located approximately 0.52 cM from several closely linked kappa markers on chromosome 6.

Animals↗
PubMed1986

Genetic polymorphism at the mouse immunoglobulin J kappa locus (Igk-J) as demonstrated by Southern hybridization and nucleotide sequence analysis.

Comparison of the nucleotide sequences of the C.C58 M75 myeloma kappa chain gene and the BALB/c germ-line J kappa segments suggested that the J kappa regions of C.C58 and BALB/c might be distinguished by restriction enzyme polymorphisms. This was shown to be the case in Southern hybridizations of Hinf I and Acc I digests of liver DNA from these and other strains with a J kappa-specific probe. Tests of a wide variety of inbred, congenic, recombinant, and recombinant-inbred strains provided evidence for three alleles, Igk-Ja, Igk-Jb, and Igk-Jc, the type strains for which are C58/J, BALB/c, and SJL/J, respectively. Analysis of the B6.PL(85NS) congenic strain suggests that the Igk-J locus lies in the neighborhood of the Lyt-2/Lyt-3 loci, approximately 0.30 cM from the V gene segment determining the Igk-VSer and Igk-Efl polymorphisms. Finally, nucleotide substitutions lead to amino acid sequence differences between the C.C58 M 75 kappa gene and the BALB/c germ line in J kappa 2 and J kappa 4. Two of these substitutions reflect true germ-line differences, raising the possibility that idiotype differences observed among strains could reflect J kappa as well as V kappa differences.

Amino Acid Sequence↗
PubMed1986

Structural differences in a single gene encoding the V kappa Ser group of light chains explain the existence of two mouse light-chain genetic markers.

Two phenotypic markers of mouse immunoglobulin kappa light chains, the IB-peptide marker and the Ef1a isoelectric focusing marker, are expressed by the C58/J, AKR/J, RF/J, and PL/J strains (called expressor strains) but not by BALB/c and most inbred strains. Expression is linked to the kappa light-chain locus and the Lyt-2/Lyt-3 genes on chromosome 6. Light chains bearing these markers belong to a group of variable region kappa chain (V kappa) regions called V kappa Ser, which has a serine amino terminus and a framework 1 region not observed to date among BALB/c light chains. Southern hybridization of genomic DNA with a V kappa Ser-specific cDNA probe has demonstrated a single strongly hybridizing DNA fragment in all strains of mice tested. Characteristic restriction enzyme polymorphisms define the V kappa Ser alleles of expressor (Igk-VSera) and nonexpressor (Igk-VSerb) strains. In the present study, the unrearranged V kappa Ser gene and its flanking regions from an expressor (C.C58) and nonexpressor (BALB/c) strain have been cloned and their nucleotide sequences determined. The C.C58 V kappa Ser gene isolated (the Igk-VSera allele) was shown to code for the two phenotypic markers described. While the nucleotide sequence of the BALB/c coding region (the Igk-VSerb allele) shows 97% identity with the C.C58 gene, single nucleotide substitutions lead to structural changes in the encoded protein which render it IB-negative and Ef1a-negative. These differences alone can explain the failure of strains containing the BALB/c allele to express these kappa-chain phenotypic markers. Also, the BALB/c gene contains a single substitution in a conserved octamer sequence approximately equal to 100 nucleotides upstream of the coding region, which could affect its expression. Finally, the C.C58 allele contains a BAM5/R repetitive DNA element approximately equal to 1200 nucleotides upstream of the coding regions that is not present in BALB/c. This element gives rise to the EcoRI and BamHI restriction enzyme polymorphisms, which distinguish the Igk-VSera and Igk-VSerb alleles.

Amino Acid Sequence↗
PubMed1985

Molecular genetic analysis of the V kappa Ser group associated with two mouse light chain genetic markers. Complementary DNA cloning and southern hybridization analysis.

Previous studies (21) have shown that two mouse kappa light (L) chain variable (V) region polymorphisms, the IB-peptide and Efla markers, reflect expression of a characteristic group of V kappa regions, called V kappa Ser, by some inbred strains and not others. Expression of V kappa Ser is controlled by a locus on chromosome 6, the chromosome that contains the kappa locus. To further characterize this V kappa group and begin to analyze the basis for its strain-specific expression, full-length complementary DNA (cDNA) copies were produced of L chain mRNA from the M75 myeloma that had been induced in the C.C58 strain of mice, and which produces a V kappa Ser L chain. The C.C58 strain is congenic with BALB/cAn, differing in the region of chromosome 6 that controls expression of the V kappa polymorphisms and the Lyt-2 and Lyt-3 T cell alloantigens. The complete nucleotide sequence of this cloned cDNA was determined and compared with the nucleotide sequences the most closely related BALB/c myeloma L chains known. Results indicated significant differences throughout the variable region, but particularly toward the 5' portion of the sequence. A probe corresponding to 200 bp of the 5' end of the cloned V kappa Ser cDNA was used in Southern hybridizations of restriction digests of liver DNA from a number of inbred, recombinant, and recombinant inbred strains. Under stringent hybridization conditions, one strongly-hybridizing fragment was observed in Bam HI, Hind III, and Eco RI digests, and based on the size of the fragments, strains could be organized into two groups. The presence of strongly hybridizing Bam HI, Hind III, and Eco RI fragments of 3.2, 2.8, and 2.1 kb, respectively, was found to correlate completely with expression by the strain of the IB-peptide and Efla markers. All nonexpressor strains yielded hybridizing fragments of 7.8, 8.4, and 2.8 kb, respectively. Possible explanations for strain-specific expression of V kappa Ser-associated phenotypic markers are discussed.

Amino Acid Sequence↗
PubMed1985

Linkage of a 7S RNA sequence and kappa light chain genes in the mouse.

A mouse 7S RNA cDNA plasmid clone was employed to identify and map DNA restriction fragment variants using recombinant inbred (RI) and congenic mouse strains. More than a dozen such restriction variants were identified and mapped to different regions of the mouse genome. One such variant, designated Rn7s-6, showed close linkage to the Ly-2,3-Igk-V (T lymphocyte antigens 2 and 3, kappa immunoglobulin variable region) cluster of markers on chromosome 6. No recombinants were detected among three of these markers in 59 RI strains. On the basis of these data, the Rn7s-6 sequence may be placed within 1.3 centimorgans of Ly-3 and one of the Igk-V-region markers, Igk-Ef1. Two mouse stocks with previously identified crossovers within the Ly-2,3-Igk-V region were used to sublocalize Rn7s-6. The results are consistent with the gene order (Ly-2, Ly-3)-(Rn7s-6, Igk-Ef1)-Igk-Ef2. Several mouse plasmacytomas, known to have various parts of the kappa chain complex deleted, retain the Rn7s-6 sequence. The Rn7s-6 variant is a plus/minus variant; no sequence allelic to Rn7s-6 is found in inbred strains that share the Ly-3a-Igk-Ef1a haplotype.

Animals↗
PubMed1984

BALB/c antiarsonate idiotypes: gene complementation necessary for expression.

The expression of two idiotype (id) families (5AF6 and 3C6) associated with the BALB/c p-azophenylarsonate-specific antibody response was examined in 11 mouse strains. Eight strains produced some of one or the other of these two id families with the mean percent expression in the anti-Ar responses of id(+) strains ranging from 8 to 43% for the 5AF6 and from 2 to 10% for the 3C6 idiotype. Four strains of mice (C58, AKR, PL, and RF) thought to have Lyt-3.1-linked VL repertoire differences from other mouse strains (Lyt-3.2) were tested for their capacity to contribute to 5AF6 and 3C6 id expression. The RF strain was capable of producing 5AF6 id and small amounts of 3C6 id. Tests of Lyt-3.1 congenic strains C.AKR (AKR Lyt-3.1 on a BALB/c background) and C.C58 (C58 Lyt-3.1 on a BALB/c background) showed that C.AKR could produce 5AF6 id while C.C58 could not. 3C6 id expression was present but depressed in C.C58 mice compared with the high 3C6 id expression in C.AKR. Breeding studies mating C.C58 (bearing the required Igh-Ca-linked VH genes) to other 5AF6(-) strains showed that gene complementation could result in 5AF6 expression in F1 offspring. 5AF6(-) strains capable of complementation included CBA/J, C57BL/6J, AKR/J, and PL/J. C58/J (from which C.C58 were derived) was the only tested strain that failed to complement for 5AF6 id expression. Additional matings between C58/J[5AF6(-)] and CBA/J[5AF6(-)] showed F1 offspring could produce 5AF6 id, indicating that C58/J can contribute functional VH genes necessary for 5AF6 id expression. Depressed expression of 5AF6 and 3C6 id was noted in mice where the C58/J-derived Lyt-3.1 genotype was present. The possibility that the depression of 5AF6 and 3C6 id expression derived from C58/J mice was due to regulatory influences rather than a lack of the VL structural genes is discussed.

Animals↗