1 alpha,25-dihydroxy vitamin D3 modulation of HLA-DR mRNA induced by gamma-interferon in cultured epithelial tumor cell lines.

Using three cultured epithelial tumor cell lines, we investigated and analyzed the effects of gamma-interferon (gamma-IFN) and 1 alpha,25-dihydroxy vitamin D3 (1,25-(OH)2D3) on the levels of HLA-DR (alpha) mRNA and HLA-DR (beta) mRNA by Northern blot analysis. After treatment with gamma-IFN alone, the levels of the mRNA increased. Treatment with both gamma-IFN and 1,25-(OH)2D3 at the same time resulted in a significant decrease in the levels of mRNA in K-TL-1, IK-TL-2, and M-TL cells as compared to those induced by gamma-IFN alone.

Platelet taurine concentration and uptake in gestosis patients with edema, proteinuria and hypertension.

The taurine concentration and uptake in platelets obtained from normal pregnant women and gestosis patients with edema, proteinuria and hypertension (EPH gestosis) were investigated. The taurine concentration in platelets showed a marked increase in severe EPH gestosis compared with normal pregnancy or mild and moderate EPH gestosis, while the plasma taurine concentration did not change significantly. Taurine uptake in platelets paralleled the severity of EPH gestosis. The Vmax of the uptake in severe EPH gestosis was about 2.4 times higher than that in normal pregnancy or mild and moderate EPH gestosis, but no significant difference was seen in the Km value among these groups.

Sequence analysis and oligonucleotide genotyping of HLA-DR"JX6", a DR"blank" haplotype found in the Japanese population.

We analyzed one of the HLA-DR"blank" haplotypes found in the Japanese population using serologic studies, sequence determination, and genotyping with sequence-specific oligonucleotide (SSO) probes. The DR"blank" haplotype, designated DR"JX6", segregated in a family in association with the DRw52 and the DQw7 specificities. The cDNA and genomic DNA of the DRB1 gene originating from the DR"JX6" haplotype were amplified enzymatically and sequenced after cloning into a plasmid vector. The amino acid sequence of the first domain in the DR beta 1 chain of the DR"JX6" haplotype was different from those of other DR haplotypes sequenced so far, but in the first hypervariable region, the sequence was identical to those of the DRw11, DRw13, DRw14, and DRw17 haplotypes. SSO probes were synthesized on the basis of the DR"JX6" haplotype sequence as well as known sequences of the DRB1, DRB3, and DRB4 genes of other DR haplotypes. These SSO probes were used for the genotyping of Japanese donors whose DRB genes were amplified enzymatically and found to show a hybridization profile that was consistent with the results of serologic studies on the DR"JX6" haplotype.

Effects of adenine nucleotides on choriogonadotropin alpha and beta subunit synthesis.

The alpha subunit of human chorionic gonadotropin (CG) contains a discrete cAMP response element in the 5' flanking region of the gene. Since cAMP also stimulates the synthesis of the CG beta subunit the presence of a cAMP cis element in the CG beta gene was examined. Deletion mutants bearing various lengths of CG beta 5' region in front of the chloramphenicol acetyl transferase (CAT) gene were transfected in placental tumor cells. No discrete cAMP response element could be identified. Unexpectedly we also observed that AMP and adenosine not only stimulated CAT activity driven by CG beta promoter sequences but also enhanced synthesis of CG alpha and beta subunits in cultured choriocarcinoma cells. GMP, CMP, guanosine, and cytosine were inactive at comparable concentrations. These data suggest that the response of the CG alpha and beta genes to the non-cyclic adenine derivatives occurs by a mechanism that differs from cAMP.

Molecular characterization of the A/J J558 family of heavy chain variable region gene segments.

The nucleotide sequences of the coding as well as the flanking regions of 11 A/J J558 heavy chain variable region (VH) gene segments are presented. Among these J558 VH segments was the unrearranged germline VH gene segment recruited in the predominant A strain-specific anti-arsonate response. Three other VH gene segments that are greater than 92% related to the p-azophenylarsenate (Ars) A VH gene segment were also isolated. Detailed analysis of the nucleotide sequences of these as well as the remaining seven J558 VH gene segments reveal that the J558 VH gene family is composed of distinct, but related, J558 VH subfamilies. Deletion mapping analyses were used to position the Ars A VH gene segment proximally with respect to the DH-JH clusters within the J558 VH gene family and distally with respect to its own J558 subfamily. The documentation of J558 VH subfamilies is discussed in the context of J558 VH family evolution and diversification.

Identification of a promoter region in the CG beta gene cluster.

To identify the promoter sequence(s) of the CG beta gene, genomic fragments derived from a cosmid containing the CG beta gene family were transfected into mouse Y1 adrenal cortical cells. Using this system, we showed that the CG beta genes 5, 3, and 8 have functional promoters, the basal element of which in the case of CG beta 5, was within 78 base pairs 5' ward from the CAP site. The size of the CG beta transcripts and identity of the transcription start site was the same for CG beta mRNA synthesized in Y1 cells as in first trimester placenta. The promoter region identified by this system was also capable of driving the chloramphenicol acetyltransferase gene when transfected stably into choriocarcinoma cells. Chloramphenicol acetyltransferase constructs bearing variable length of 5'-flanking sequences from CG beta 5 and transfected into trophoblast cells suggest the presence of regulatory sequences within 700 base pairs from the CAP site. The information obtained here provide a foundation for studies of analyzing trans-acting placental and pituitary proteins to the defined CG beta promoter region.

Partial N-terminal sequence analysis of human class II molecules expressing the DQw3 determinant.

HLA-DQ molecules were isolated from DRw9-homozygous and DR4-homozygous cell lines by using a monoclonal antibody HU-18, which recognizes class II molecules carrying the conventional DQw3 determinant. The partial N-terminal sequence analysis of the DQw3 molecules revealed that they have sequences homologous to those of murine I-A molecules. Within the limits of our sequence analysis, the DQw3 molecules from the two cell lines are identical to each other in both the alpha and beta chains. The DQ alpha as well as DQ beta chains were found to have amino acid substitutions when compared to other I-A-like molecules whose sequences have been reported. These differences may contribute to the DQw supertypic specificity. The polymorphic nature of DQ molecules is in marked contrast to that of DR molecules where DR alpha chains are highly conserved while DR beta chains have easily detectable amino acid substitutions.