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D Pious

Publications and source records attributed to D Pious.

53 records · Page 3Linked to original sources
PubMed1982

Gene dosage and gene expression in the HLA region: evidence from deletion variants.

Among variants selected in a human cell line for nonexpression of a single gene product in the HLA complex, most are single-gene variants, but several have been isolated that are cis-acting and abolish expression of a series of closely linked genes. The two most plausible mechanisms by which such variants could arise are mitotic crossing-over and chromosome deletion. In two HLA variants the presence of a visible chromosome deletion, a 50% reduction in activity of glyoxalase I (a closely linked marker), or both provided evidence for deletional origins. In a third variant these changes were not demonstrable. All three variants showed reductions in amount of cell surface HLA antigens: 40% for the Ia antigens (HLA-DR) and 20-25% for HLA-ABC antigens. The reductions in cell surface antigen in deletion variants have an important implication: in the case of the HLA-A, -B, and -C heterodimer, which consists of a subunit coded for within the major histocompatibility complex and another subunit (beta(2)-microglobulin) coded for on a different chromosome, it is the gene of the major histocompatibility complex that is limiting. The nonmutant haplotype includes A2; binding of an A2 monoclonal antibody in two of the mutants was found to be approximately equal to that in the wild-type cells. Thus, loss of one copy of HLA-ABC genes does not lead to gene dosage compensation-i.e., increased activity by the remaining ABC alleles. The results with the two types of antibodies support a deletional mechanism and are inconsistent with mitotic crossing-over. Of interest with respect to the potential use of deletion variants for purposes of mapping is the fact that each of these variants has distinctive breakpoints. The absence of mitotic crossing-over in 1.2 x 10(7) cells selected suggests that the event is rare in this autosomal region, if it occurs at all.

Cell Line↗
PubMed1980

Identification of a trans-acting function regulation HLA-DR expression in a DR-negative B cell variant.

Somatic cell hybridizations were performed between an HLA-DR negative variant of a human B lymphoid cell line (B-LCL) and normal unrelated B-LCLs. The HLA-DR codes for polymorphic determinants on a heterodimeric cell surface lymphocyte differentiation glycoprotein. A variant subline which was selected in a single step from a diploid heterozygous DR-1 DR-3 B-LCL had lost expression of both DR-1 and DR-3 and the heretodimer; it has been described earlier. In a fusion with a DR-2 B-LCL, the hybrids expressed DR-2 and reexpressed the DR-1 and DR-3 alleles. Similar results were seen in a fusion with a different normal B-LCL. Hybrid clones from both fusions were tested with a large number of alloantisera and essentially all informative sera showed reexpression. The results show that (1) the variant did not arise by mutations in the structural genes for DR-1 and DR-3; (2) the normal cells are supplying a missing gene product needed for expression of DR; (3) this gene product is capable of acting in trans. Chromosome counts showed that the apparent recessiveness of the variant in the hybrids was not due to chromosomal segregation.

B-Lymphocytes↗
PubMed1980

Linked marker analysis of spontaneous HLA variants of somatic cells.

Fourteen independent, spontaneously arising HLA-A1 and -B27 variant clones were isolated from the pseudodiploid B lymphoid line T5-1 by selection using A1 and B27 alloantiserum, respectively, and complement. T5-1 is heterozygous for the tightly linked loci HLA-DR, -B, and -A and probably for -C as well. Following recloning, each of the variants was tested for the HLA specificities of T5-1. None of these spontaneous variants had a genetic lesion which extended to the nearest flanking HLA marker, less than 1 cM distant. On the other hand, a variant isolated from mutagenized cells had a lesion which extended completely through one HLA region in cis, suggesting that haploid expression in the HLA region is compatible with viability, that there are no recessive lethals in the opposite HLA region, and that spontaneously arising variants with extended lesions could have been recovered had they occurred. From these results, we conclude that extended genetic lesions of 0.8 cM or longer contribute less than 10% to the overall rate of spontaneous variation for HLA alleles, which we previously estimated at about 5 x 10(-7) per cell per generation.

B-Lymphocytes↗
PubMed1979

Absence of allogeneic restriction in human T-cell-mediated cytotoxicity to Epstein-Barr virus-infected target cells. Demonstration of an HLA-linked control at the effector level.

Peripheral T lymphocytes from patients with infectious mononucleosis (IM) are sensitized in vivo against the Epstein-Barr virus (EBV). The expression of HLA-A, B, or C molecules at the target cell surface is necessary for the cytotoxic reaction because (a) EBV-positive Daudi cells lacking HLA-A, B, and C determinants are resistant to anti-EBV T-cell lysis, (b) cytolysis of EBV-positive target cells can be consistently inhibited by anti-HLA-A, B, and C and anti-beta 2 microglobulin antibodies. However, no evidence for allogeneic restriction in this system was apparent as (a) cytotoxic T lymphocytes (CTL) from one given individual could exert a cytotoxicity of a similar magnitude on different EBV-positive target cells, regardless of the number of HLA-A or B specificities shared by the effectors and targets; (b) CTL from IM patients were able to kill target cells without any HLA-A or B antigen in common; and (c) T5-1 variants lacking one or two HLA antigens at the A, B, or D locus are killed to the same extent as the parental cells. 7 of the 9 IM patients with detectable circulating anti-EBV CTL carried the HLA-A1 antigen, whereas none of the 16 IM patients lacking detectable peripheral CTL were HLA-A1 positive (mean specific lysis of T5-1 target cells by T cells from HLA-A1 positive patients: 29.3 vs. 0.6% in HLA-A1-negative patients) (P less than 10(-9)). These data suggest an HLA-A1-linked gene control of the magnitude of the anti-EBV CTL response. Thus, the HLA region appears to act at two different level sin the T-cell-mediated lysis of EBV-infected cells by controlling first, the development of anti-EBV and second, the expression of HLA-A, B, and C molecules involved as recognition structures at the target cell surface.

Antigen-Antibody Reactions↗
PubMed1977

HLA variants of cultured human lymphoid cells: evidence for mutational origin and estimation of mutation rate.

Variants of a diploid lymphoid cell line that show a loss of HLA-B27 antigen occur randomly in time and independently of exposure to the alloantiserum used for their isolation. From these and previous findings of variant stability, inducibility by mutagens, and the absence of linked variation, we colclude that most HLA variants arise by mutation. The mutation rate for HLA-B27 loss is 8 x 10(-7) per cell per generation.

B-Lymphocytes↗
PubMed1976

Isolation of stable and unstable ("suppressed") Beta2-microglobulin-deficient clones from a lymphoid cell line.

After selection with anti-Beta2m and C against a human lymphoid line, about 10% of surviving clones manifested decreased cytotoxic sensitivity to anti-Beta2m("suppression") which gradually reverted to normal after 35 to 55 cell doublings. Antiserum alone induced suppression. Exposure of suppressed clones to DMSO substantially reversed suppression. One clone, surviving selection, had a stable decrease in anti-Beta2m sensitivity; it possibly represents a Beta2m mutation.

Beta-Globulins↗
PubMed1973

Isolation and characterization of HL-A variants in cultured human lymphoid cells.

We have developed a procedure for immune selection in an established human lymphoid cell line based on HL-A, the major human histocompatibility locus. After a single brief exposure to selective conditions, HL-A2 variant clones were isolated from an HL-A2/HL-A3 heterozygous line. The variant clones occurred at a frequency of about 1 x 10(-6). The variant phenotype was stable during prolonged growth in the absence of antiserum after isolation. The variant sublines bound [unk] 1/1000 the HL-A2 antibody per cell as the parent line. Variation was specific in that expression of antigens not selected against was unimpaired. Loss of the chromosome bearing HL-A2 was excluded as the cause of variation because the variants, like the parent line, were heterozygous for phosphoglucomutase (EC 2.7.5.1) determined by structural locus PGM(3), which is linked to HL-A.

Animals↗
PubMedDate not supplied

A gene in the human major histocompatibility complex class II region controlling the class I antigen presentation pathway.

Major histocompatibility complex (MHC) class I molecules export peptides to the cell surface for surveillance by cytotoxic T lymphocytes. Intracellular peptide binding is critical for the proper assembly and transport of class I molecules. This mechanism is impaired as a result of a non-functional peptide supply factor gene (PSF) in several human mutant cell lines with genomic lesions in the MHC. We have now identified PSF in the MHC class II region by deletion mapping in mutants and chromosome-walking. PSF is homologous to mammalian and bacterial ATP-dependent transport proteins, suggesting that it operates in the intracellular transport of peptides.

Amino Acid Sequence↗