Differential expression of interleukin 1 alpha by Thy-1+ and Thy-1- lung fibroblast subpopulations: enhancement of interleukin 1 alpha production by tumor necrosis factor-alpha.

The purpose of this investigation was to determine whether subpopulations of murine lung fibroblasts produced interleukin 1 (IL 1). We previously identified two major populations of pulmonary fibroblasts based on the presence or absence of Thy-1. Thy-1+ and Thy-1- subsets synthesize fibronectin and type I and III collagen, but only the Thy-1- population displays class II major histocompatibility complex antigens after stimulation with interferon-gamma and presents antigen to T helper clones.

CD43 (leukosialin, sialophorin, large sialoglycoprotein) can be expressed in both normal and Wiskott-Aldrich fibroblasts via transfection of a leukosialin cDNA.

Human leukosialin is among the most abundant sialoglycoproteins found on the surface of cells of the lympho-hematopoietic system. Leukosialin, also known as sialophorin, is involved in T cell proliferation, and its molecular isoform changes upon cellular activation. We show that human leukosialin is identical to the antigens described by the monoclonal antibodies (mAb) G10-2, G19-1 (CD43) and B1B6 (large sialoglycoprotein).

Mutations in the alpha 1 domain of a class I gene define residues important for specific allorecognition.

Our strategy to use saturation mutagenesis to produce an unbiased collection of major histocompatibility class I mutants has resulted in unpredicted mutant phenotypes. First, we have shown data supporting our earlier work of the Dp20(Y27N) mutant. Allorecognition is altered at the clonal level while no variation in lymphocytic choriomeningitis virus (LCMV)-restricted recognition is observed.

Alteration of the metastatic potential of line 1 lung carcinoma cells: opposite effects of class I antigen induction by interferons versus DMSO or gene transfection.

Class I antigens are necessary for the recognition of tumor cells by cytotoxic T lymphocytes (CTL). The line 1 lung carcinoma is a spontaneous murine tumor deficient in class I antigen expression. Consistent with this, line 1 cells are highly metastatic in vivo.

The transport of class I major histocompatibility complex antigens is determined by sequences in the alpha 1 and alpha 2 protein domains.

The transport of human-mouse hybrid class I histocompatibility antigens has been studied in a mutant human cell line, 174 X CEM.T2 (T2). T2, a somatic cell hybrid of human B- and T-lymphoblastoid cell lines (B-LCL and T-LCL, respectively), synthesized HLA-A2 and HLA-B5 glycoproteins, but expresses only low levels of A2 and undetectable levels of B5 at the cell surface.

Antibodies specific for Ig idiotype, but not isotype, can substitute for antigen to induce IgM secretion by a B cell clone.

Cells of the mouse B cell clone, CH12.LX, secrete IgM when cultured with nominal antigen (sheep erythrocytes, SRBC) and mAbs which bind their membrane Ek molecules. To determine whether anti-Ig antibodies can substitute for antigen in the induction of IgM secretion by CH12.

cDNA cloning and localization of the mouse leukosialin gene (Ly48) to chromosome 7.

Mouse leukosialin, previously known as the 3E8 antigen, is expressed primarily on cells of the hematopoietic and lymphoid lineages and is shown to be the mouse homologue to the human leukosialin/sialophorin and rat W3/13 molecules. A partial leukosialin cDNA clone was isolated via cross-species hybridization with a portion of a human leukosialin cDNA. This mouse cDNA clone was used to demonstrate that the leukosialin isoforms are encoded by a single mRNA species of approximately 4.

Haplotype-specific differences in signaling by transfected class II molecules to a Ly-1+ B-cell clone.

CH12.LX B cells are responsive to antigen-dependent differentiative signals transmitted through their surface Ek molecules. Although CH12.

Evidence for a subpopulation of antigen-presenting cells specific for the induction of the delayed-type hypersensitivity response.

Young adult SJL mice (8 weeks of age or younger) do not mount a delayed-type hypersensitivity (DTH) response due to the failure of a macrophage antigen-presenting cell (APC) to induce TDTH effector cells. SJL mice that are 10 weeks of age or older produce a normal DTH response. This genetic defect provides a model for the investigation of functional subpopulations of APC which interact with specific subpopulations of T cells.

Characterization of cDNAs encoding human leukosialin and localization of the leukosialin gene to chromosome 16.

We describe the isolation and characterization of cDNA clones encoding human leukosialin, a major sialoglycoprotein of human leukocytes. Leukosialin is very closely related or identical to the sialophorin molecule, which is involved in T-cell proliferation and whose expression is altered in Wiskott-Aldrich syndrome (WAS), an X chromosome-linked immunodeficiency disease. Using a rabbit anti-serum to leukosialin, a cDNA clone was isolated from a lambda gt11 cDNA library constructed from human peripheral blood cells.

Differential transport requirements of HLA and H-2 class I glycoproteins.

Transport of human and mouse major histocompatibility complex class I glycoproteins has been examined in a transport deficient B-lymphoblastoid cell line X T-lymphoblastoid cell line (B-LCL X T-LCL) hybrid, 174 X CEM.T2 (T2). This cell line expresses no detectable endogenous HLA-B5 and reduced levels of HLA-A2 on its surface although these molecules are synthesized.

Diminished expression of the T cell receptor on the expanded lymphocyte population in MRL/Mp-lpr/lpr mice.

MRL mice homozygous for the recessive lpr gene develop an accelerated autoimmune syndrome and massive lymphadenopathy. Because the function of the expanded lymph node population is unclear, we have studied the subunits of the T cell receptor for antigen (TcR). DNA and RNA were prepared from MRL/Mp-lpr/lpr (lpr) and congenic MRL/Mp(-)+/+ (+/+) mice by standard techniques and studied by Southern blot, northern blot, and dot blot analysis using the cDNAs TT11, specific for the TcR alpha chain; 86T5, specific for the TcR beta chain; and T3 delta; specific for the subunit of the T3 molecule.

High-level expression of a bioengineered, cysteine-free hepatocyte-stimulating factor (interleukin 6)-like protein.

Hepatocyte-stimulating factor, interferon-beta 2, B-cell stimulation factor 2, and hybridoma/plasmacytoma growth factor are identical proteins presently referred to as interleukin 6 (IL-6). Through the use of synthetic oligonucleotide technology, we have constructed a biologically active recombinant IL-6 (rIL-6) gene based on the sequence of a human IL-6 cDNA. The synthetic gene encodes a cysteine-free, bioengineered rIL-6 protein that is expressed at high levels in Escherichia coli as a tripartite fusion protein.

Random oligonucleotide mutagenesis: application to a large protein coding sequence of a major histocompatibility complex class I gene, H-2DP.

We have used random oligonucleotide mutagenesis (or saturation mutagenesis) to create a library of point mutations in the alpha 1 protein domain of a Major Histocompatibility Complex (MHC) molecule. This protein domain is critical for T cell and B cell recognition. We altered the MHC class I H-2DP gene sequence such that synthetic mutant alpha 1 exons (270 bp of coding sequence), which contain mutations identified by sequence analysis, can replace the wild type alpha 1 exon.

Cytotoxic T lymphocyte recognition of hybrid MHC class I molecules.

We have utilized a group of MHC class I genes produced by in vitro recombination between Dp and Dd to study recognition of MHC class I molecules by cytolytic T cells (CTLs). Both polyclonal allo-specific and H-2-restricted CTLs require that alpha 1 and alpha 2 of the target class I molecule be derived from the same haplotype for efficient killing. By using T-cell lines we showed that within the bulk population there must exist a fraction of T cells which can recognize epitopes in alpha 1 or alpha 2.

Mouse genetic locus Lps influences susceptibility to Neisseria meningitidis infection.

We surveyed a number of inbred mouse strains for susceptibility to meningococcemia. Mice of all strains became bacteremic after intraperitoneal injection of a serogroup C, serotype 2a human disease isolate, but the strains differed in levels of bacteremia, indicating influences of the host genome on susceptibility. There was no significant correlation between level of bacteremia and differences at major histocompatibility or immunoglobulin loci; the Salmonella susceptibility locus, Ity; the complement C5 locus, Hc; the antibody response locus, xid; or the transferrin locus, Trf.

Molecular analysis of deficient class I H-2 antigen expression by mouse lung carcinoma cells.

We have continued our investigations of line lung carcinoma cells to understand the molecular basis of decreased expression of class I H-2 Ag and class I Ag induction with DMSO. We show that line 1, a murine lung carcinoma cell line, has low levels of class I Ag (H-2K, D, and L) because it is deficient in both class I and beta 2-microglobulin (B2M) RNA, and that these mRNA can be coordinately induced with DMSO. Evidence presented herein also shows that IFN-gamma can induce surface expression of class I Ag and suggests that it may act through a different mechanism than DMSO in inducing class I Ag.

Saturation mutagenesis of a major histocompatibility complex protein domain: identification of a single conserved amino acid important for allorecognition.

The interactive association between T lymphocytes and their target cells is an important system of cell-cell interactions. Major histocompatibility complex class I molecules are the cell surface structures recognized by cytolytic T lymphocytes. To define the molecular structures recognized by cytotoxic T lymphocytes, we have saturated the 270-base-pair alpha 1 exon of the H-2Dp gene with point mutations, rapidly producing a "library" of 2.

Lysis of a lung carcinoma by poly I:C-induced natural killer cells is independent of the expression of class I histocompatibility antigens.

Cells from the line 1 murine carcinoma express little if any H-2d when grown in normal medium. These cells are susceptible to splenic cell populations with NK activity, stimulated by prior injection of poly I:C, but are not lysed by NK-deficient splenocytes from homozygous beige mice treated with anti-asialo GM1. Incubation of line 1 cells in medium containing DMSO leads to a dramatic stimulation of H-2d expression but no change in lytic susceptibility to splenic NK cells.

Identification and characterization of a mouse cell surface antigen with alternative molecular forms.

We present the characterization of a new mouse cell surface protein, recognized by the 3E8-specific monoclonal antibody. The expression of this antigen is predominantly restricted to the hematopoietic and lymphoid tissues: bone marrow, spleen, lymph node, and thymus. Immunoblot analyses show that the 3E8 determinant is present on molecules with different apparent relative masses.

Signaling to a B-cell clone by Ek, but not Ak, does not reflect alteration of Ak genes.

The mouse B-cell clone, CH12.LX (Iak, Ly-1+, mu+, delta+), can be induced to differentiate and secrete antibody in an antigen-specific, H-2-restricted manner. Induction requires two signals.