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Biomedical subjects

L Hood

Publications and source records attributed to L Hood.

At least 289 records · Page 16Linked to original sources
PubMed1985

L3T4 but not LFA-1 participates in antigen presentation by Ak-positive L-cell transformants.

We report that mouse L cells expressing Ak class II molecules on their surface after DNA-mediated gene transfer are capable of presenting the synthetic copolymer (Glu60 Ala30 Tyr10) to Ak-restricted long-term T-cell clones. Antigen-induced T-cell stimulation could be inhibited by monoclonal antibodies (mAb) directed at spatially distinct determinants of the alpha and/or beta subunits of the Ak molecule, and by the rat L3T4-specific mAb H129.19. In contrast, several rat mAb reactive with the mouse LFA-1 molecule failed to inhibit T-cell activation when L cells were used as antigen-presenting cells (APC), although these mAb strongly inhibited the same T-cell responses in the presence of leukocytic APC. Similarly, the cytolytic activity of the Ak-specific T-cell clone A15.1.17 was blocked by L3T4-specific and by LFA-1-specific mAb when tested on Ak-positive B-cell hybridomas, but only by L3T4-specific mAb and not by LFA-1-specific mAb when Ak-positive L-cell transformants were used as targets. These data support the notion that the LFA-1 molecule is not necessary for T-cell activation, and suggest that its functional role as an accessory molecule depends on the leukocytic nature of the APC tested.

Animals↗
PubMed1985

Rearranged beta T cell receptor genes in a helper T cell clone specific for lysozyme: no correlation between V beta and MHC restriction.

The helper T cell clone 3H.25 is specific for hen egg white lysozyme and the class II MHC molecule I-Ab. This TH cell has three rearrangements in the beta-chain gene family-a V beta-D beta-J beta 1 and a D beta 2-J beta 2 rearrangement on one homolog and a D beta 1-J beta 2 rearrangement on the other. These observations demonstrate that this functional T lymphocyte expresses only a single V beta gene segment and, accordingly, exhibits allelic exclusion of beta-chain gene expression. The rearranged 3H.25 V beta gene segment is the same as that expressed in a T helper cell specific for cytochrome c and an I-Ek MHC molecule. Thus, there is no simple correlation between the V beta gene segment and antigen specificity or MHC restriction.

Animals↗
PubMed1985

Cloning and characterization of the myelin basic protein gene from mouse: one gene can encode both 14 kd and 18.5 kd MBPs by alternate use of exons.

A mouse cosmid library was screened with a cDNA clone for rat small (M, 14,000) myelin basic protein (MBP). The entire MBP gene was isolated. Five exons were detected with the cDNA clone, which encode the Mr 14,000 MBP. A sixth exon, detected using a synthetic oligonucleotide, encodes the 41 amino acids specific to the Mr 18,500 form of MBP. Splicing together all six exons would give rise to Mr 18,500 MBP, while an mRNA omitting exon V would be translated to produce Mr 14,000 MBP. Further experiments indicate that there is probably a single MBP gene in the mouse genome, and that there is a single major 5' end for mouse MBP transcripts, 47 bp 5' of the initiator methionine codon.

Animals↗
PubMed1985

Chromosomal mapping of mouse myelin basic protein gene and structure and transcription of the partially deleted gene in shiverer mutant mice.

The gene for mouse myelin basic protein (MBP) was mapped to chromosome 18 by hybridization of cloned MBP probes to DNA from hamster-mouse hybrid cell lines, showing it to be linked to the shiverer mutation which causes abnormal CNS myelination. Genomic blotting experiments show that in the mutants five of six exons which constitute the wild-type gene have been deleted. In shiverer brains the steady state level of transcripts that initiate correctly at the 5' end of the remaining exon 1 is reduced 16-fold. These RNAs are not spliced correctly and are not efficiently polyadenylated. It is proposed that the partial deletion of the MBP gene is an important part of the shiverer lesion.

Animals↗
PubMed1985

Ia-transfected L-cell fibroblasts present a lysozyme peptide but not the native protein to lysozyme-specific T cells.

We studied the antigen-presenting capacity of mouse L fibroblasts transfected with genes encoding Ia polypeptides of the major histocompatibility complex (MHC). These cells function as efficient antigen-presenting cells (APC) in stimulating peptide antigen-specific MHC-restricted proliferation of long-term T-cell lines, thus establishing the capacity of Ia-expressing L-cell transfectants to present antigens to apparently normal T cells. However, in contrast to splenic APC, L-cell transfectants fail to present native hen egg-white lysozyme to the same T cells. Since this result is similar to that obtained with physiologic APC pretreated to prevent antigen degradation, it suggests that L-cell transfectants, without such pretreatments, may be compromised in their ability to process native lysozyme. However, since such transfectant cells have been shown to present other complex polypeptides such as keyhole limpet hemocyanin, a random copolymer of glutamic acid, alanine, and tyrosine, and influenza virus neuraminidase, this observation suggests that protein antigens differ in the stringency of processing requirements.

Animals↗
PubMed1985

Expression of hybrid class I genes of the major histocompatibility complex in mouse L cells.

The class I genes of the major histocompatibility complex of the mouse can be divided into two categories: those encoding the transplantation antigens and those encoding the Qa and Tla antigens. The inbred BALB/c mouse has 28 potential Qa/Tla genes. The sites of tissue expression, developmental regulation, and functions of these genes are virtually unknown. We have used the technique of exon shuffling to construct hybrid genes between each of three Qa region genes (Q5, Q7, and Q8) and two other class I genes (H-2Ld and Q6). The hybrid genes have been transfected into mouse L cells, in which intact transplantation antigen genes generally are expressed and in which intact Qa genes generally are not expressed. Analysis of expression of the hybrid gene constructs indicates that the 5' half of two of the Qa genes (Q5 and Q8) can readily be expressed in the context of a hybrid molecule, whereas the 3' half prevents cell-surface expression. The exon shuffling approach described here will be useful in characterizing Qa/Tla genes and in identifying or producing new reagents to study the Qa/Tla gene products, their tissue distribution, their developmental stages of expression, and, ultimately, their functions.

Animals↗
PubMed1985

Localization of the human myelin basic protein gene (MBP) to region 18q22----qter by in situ hybridization.

A restriction endonuclease fragment derived from a cloned portion of human genomic DNA corresponding to the myelin basic protein gene has been used to map the position of this gene by in situ hybridization to human metaphase chromosomes. Ten percent of the radioactively labeled sites observed were on chromosome 18. Eighty-four percent of the grains on chromosome 18 were located within the region corresponding to 18q22----qter. This represents a greater than 10-fold increase in labeling at this position over the background grain distribution found along all of the other chromosomes.

Animals↗
PubMed1985

Interaction of alpha 1 with alpha 2 region in class I MHC proteins contributes determinants recognized by antibodies and cytotoxic T cells.

The structure-function relationship of individual coding regions of class I mouse major histocompatibility complex proteins was studied by a combination of recombinant DNA, gene transfer techniques, and serologic and functional characterization. To examine the role of alpha 1 and alpha 2 regions in antibody and CTL recognition, the third exon of H-2Dd, Kd, and Ld transplantation antigen genes was replaced by the homologous coding region of the Qa-2-coded class I gene, Q6. We have chosen to carry out the exon shuffling experiments between these two different types of class I genes, because they are structurally similar and did not evolve to carry out identical functions. Therefore, it is less likely that the hybrid proteins will fortuitously recreate alpha 1-alpha 2 controlled functionally important determinants. The replacement of H-2 alpha 2 coding region with its Q6 counterpart had different effects on the expression of the three genes. The mutant H-2Dd gene transfected into L cells was expressed at high levels and retained several of the serologic determinants found on parental H-2Dd and Q6 domains. The serologic epitopes on the mutant H-2Kd-transfected cells were detectable at very low levels, whereas the product of the mutant H-2Ld gene could not be identified at all. Analysis of cells transfected with mutant H-2Dd gene with alloreactive and minor antigen(s)-restricted cytotoxic T cells indicated that the hybrid proteins lost the ability to be recognized by T cells. Our data suggest that cytotoxic T cells recognize conformational determinants composed of amino acids from alpha 1 and alpha 2 regions. Alternatively, it could be proposed that T cell recognition sites located in a single alpha 1 or alpha 2 protein region are susceptible to distortion upon alpha 1-alpha 2 interactions. Such susceptibility to conformational changes of the amino-terminal domain of transplantation antigens could be of functional importance for H-2-restricted antigen presentation.

Animals↗
PubMed1984

Tracts of high or low sequence divergence in the mouse major histocompatibility complex.

The K, I and S regions of the mouse major histocompatibility complex (MHC) are composed of long tracts of DNA which differ in sequence divergence. A correlation exists between the location of an MHC gene in a variable or conserved chromosomal tract and the degree of polymorphism and diversity of the proteins encoded by its alleles. Variable tracts appear to be the result of mechanisms which mutate certain coding and non-coding sequences to the same extent and selective pressures operating on the genes.

Animals↗
PubMed1984

Multiple VH gene segments encode murine antistreptococcal antibodies.

Most mouse strains are able to mount a diverse antibody response against group A streptococcal carbohydrate (GAC). We have previously reported that murine anti-GAC antibodies are for the most part restricted to IgM and IgG3 subclasses. In addition, despite extensive heterogeneity in their isoelectric focusing patterns, greater than 50% of A/J anti-GAC antibodies share a common light chain defined by spectrotypic and idiotypic (VK1GAC) criteria. We have used protein and DNA sequencing strategies to examine the genetic basis of diversity in murine anti-GAC antibodies. In particular, we report that, (a) multiple, closely homologous VH gene segments contribute to the generation of anti-GAC antibodies, (b) a common framework sequence, related to the VK27 subgroup, probably defines VK1GAC, and (c) the A/J anti-GAC VH regions and BALB/c anti-inulin VH sequences are 95% homologous at the protein level and are likely encoded by overlapping VH gene families. Lastly, we discuss the genetic mechanisms that might permit the evolution of multiple, closely homologous germline VH gene segments in the context of highly divergent flanking region sequences.

Alleles↗
PubMed1984

Cytotoxic T lymphocytes recognize determinants on the BALB/c-H-2Ld molecule controlled by alpha 1 and alpha 2 but not alpha 3 external domains.

We have shown that cytotoxic T lymphocytes (CTL) raised in H-2d mice use H-2Ld but not H-2Dd or H-2Kd antigens as restricting elements in lymphocytic choriomeningitis virus (LCMV) and vesicular stomatis virus (VSV) infections. To localize the regions of H-2Ld protein recognized by CTL, we constructed a recombinant H-2Ld/Dd gene encoding a hybrid antigen with alpha 1 and alpha 2 external domains of H-2Ld and alpha 3, transmembrane and cytoplasmic domains of H-2Dd. The recombinant gene was transfected into mouse cells and the hybrid molecules were characterized serologically, biochemically and functionally. In all assays, H-2Ld/Dd molecules were recognized by LCMV-and VSV-specific H-2Ld-restricted CTL in a manner similar to that of wild-type H-2Ld antigens. Analogous results were obtained with alloreactive CTL. Hybrid antigens containing the alpha 3 domain of H-2Ld fused to alpha 1 and alpha 2 domains of a Qa-2,3 region-encoded antigen were not used as restricting elements by LCMV-specific CTL. These results suggest that H-2Ld-restricted CTL directed against LCMV and VSV recognize determinants controlled by the alpha 1 and/or alpha 2 domains of the H-2Ld molecule.

Animals↗
PubMed1984

Cloning and identification of the H-2Dp gene.

We have cloned six different class I genes from a B10.P sperm library. After cotransfection with the herpes simplex tk gene, one L-cell line was found to react with six H-2Dp-specific monoclonal antibodies. The cell line L12a did not react with Kp-specific monoclonal antibodies. This identification was confirmed by mapping a 2.5 kb Bam H1 restriction fragment present in the lambda 12a DNA clone to the D-TL region of H-2p. Only a single 8.8 kb Bam H1 fragment can be assigned to Kp by restriction fragment length polymorphism, while many others map to the D-TL interval. A restriction map of lambda 12a is presented.

Animals↗
PubMed1984

Gene transfer of H-2 class II genes: antigen presentation by mouse fibroblast and hamster B-cell lines.

We have transferred the mouse Ak alpha and Ak beta genes, which encode the class II I-Ak molecule, into mouse L-cell fibroblasts and hamster B cells. I-Ak molecules are expressed on the surface of both cell types. The L-cell and hamster B-cell I-Ak molecules appear normal by serological analyses and two-dimensional gel electrophoresis. Furthermore, the I-Ak molecules on L cells can act as targets for the allogenic T-cell killing of the transformed L cells. The I-Ak molecules in both mouse fibroblasts and hamster B cells can present certain antigens to T-cell helper hybridomas. Thus only class II molecules are required to convert the nonantigen-presenting cell. Accordingly, it will be possible to dissect the structure-function relationships existing between Ia molecules, foreign antigen, and T-cell receptor molecules by in vitro site-directed mutagenesis and gene transfer.

Animals↗
PubMed1984

The human T cell antigen receptor is encoded by variable, diversity, and joining gene segments that rearrange to generate a complete V gene.

A cDNA clone YT35 , synthesized from poly(A)+ RNA of the human T cell tumor Molt 3, exhibits homology to the variable (V), joining (J), and constant (C) regions of immunoglobulin genes. We have isolated and sequenced the germ-line V and J gene segment counterparts to YT35 from a human cosmid library, and these failed to encode 14 nucleotides of the cDNA clone between the V and J regions. We postulate that these 14 nucleotides are encoded by a third gene segment analogous to the diversity (D) gene segments of immunoglobulin heavy chain genes. This T cell antigen receptor V gene appears to be assembled from three gene segments, V, D, and J, and accordingly most closely resembles immunoglobulin heavy chain V genes.

Amino Acid Sequence↗
PubMed1984

Genomic constitution of an H-2:Tla variant leukemia.

A TL+ leukemia of a (B6 X A)F1 hybrid mouse (H-2b/H-2a) was previously subjected to immunoselection against H-2a by passage in (B6 X A.SW)F1 mice (H-2b/H-2s). A variant leukemia line was obtained that serologically lacked not only the H-2a phenotype but also the TL phenotype determined by the linked cis Tlaa allele of strain A. The H-2b phenotype and the TL phenotype of the Tlab allele of the B6 strain, which is expressed only by leukemia cells, were retained by the variant. Southern blotting with an H-2 cDNA probe that identifies restriction fragment polymorphisms distinguishing alleles of the H-2 and Tla regions of the B6 and A strains indicates that both the H-2a and Tlaa alleles are missing from the genome of this H-2a:Tlaa negative variant. Since the variant has two apparently unaltered chromosomes 17, where the H-2:Tla complex is situated, and since the intensity of bands in Southern blotting is suggestive of H-2b homozygosity, it is considered that loss of the H-2a:Tlaa haplotype by the variant was accompanied by duplication of the H-2b:Tlab haplotype. The implied change from heterozygosity to homozygosity that the variant has undergone with respect to H-2:Tla was not paralleled by a similar change at the three other loci tested, since the variant retained heterozygosity for Pep-3 (chromosome 1), Gpi-1 (chromosome 7), and Es-1 (chromosome 8).

Animals↗
PubMed1984

Cell-type-specific cDNA probes and the murine I region: the localization and orientation of Ad alpha.

Labeled cDNA probes highly enriched for B-cell-specific (B-T) and T-suppressor-cell-specific (Ts-B) sequences were used to screen a set of genomic cosmid clones spanning 230 kilobases of the murine immune response (I) region. With the B-cell derived probe, four I-region genes were detectable (A beta, E beta, E beta 2, and E alpha), as well as an additional (fifth) region of hybridization. The T-cell probe, prepared from a putative I-J positive suppressor cell hybridoma, was negative in a parallel experiment. A genomic fragment corresponding to the new region of hybridization seen with the B-cell cDNA probe identified a discrete mRNA species in RNA blotting analysis that had a pattern of expression strikingly similar to E alpha mRNA in a variety of lymphoid tumor lines. This fragment was also used to isolate cDNA clones from a library highly enriched (20-40 times) for B-cell-specific sequences (selected cDNA cloning). DNA sequence analysis of one of these cDNA clones indicates that this gene encodes the A alpha molecule of the d haplotype (Ad alpha). These findings establish that the order of the class II genes in the Ia complex is as follows: centromere-A beta-A alpha-E beta-E beta 2-E alpha.

Animals↗