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Biomedical subjects

N H Jones

Publications and source records attributed to N H Jones.

9 recordsLinked to original sources

Expression and function of a CD5 cDNA in human and murine T cells.

In order to define the function of the CD 5 (T1, Leu 1, Tp 67 in the human; Ly-1 in the mouse) molecule, a cDNA clone of human CD 5 was expressed in a CD5-deficient Jurkat cell line and in a murine T cell hybridoma. A Jurkat subclone (Jurkat 9.9) produced interleukin 2 (IL 2) in response to anti-CD 3 monoclonal antibody (mAb) cross-linked to solid support. IL 2 production was enhanced by co-culture with the anti-CD 5 mAb OKT 1. A CD 5-deficient mutant clone Jurkat 1.15 was generated by treatment with ethyl methanesulfonate followed by selection with anti-CD 5 mAb plus complement. Jurkat 1.15 did not demonstrate enhancement of IL 2 production by OKT 1 in the presence of cross-linked anti-CD 3 mAb. A cDNA encoding human CD 5 was introduced into a defective retrovirus which was used to infect Jurkat 1.15. A Jurkat clone stably expressing CD 5 was established. In response to OKT 1, a rise in intracellular calcium was observed in both the parent Jurkat 9.9 and the CD 5+ infectant but not in the CD 5- mutant or a G 418- resistant control. Furthermore, expression of CD 5 restored the augmentation of IL 2 production by OKT 1 in response to cross-linked anti-CD 3 mAb. A murine T cell hybridoma By 155.16 which produces IL 2 in response to HLA-DR antigens was also infected with the CD 5-recombinant retrovirus and three stable CD 5+ infectants were generated. These hybridomas showed enhancement of IL 2 production by stimulation with OKT 1 in the presence of suboptimal concentrations of soluble anti-murine CD 3 mAb. These results provide further evidence that CD 5 provides a co-stimulatory signal for T cell activation.

Animals

Isolation and structural characterization of the human 4F2 heavy-chain gene, an inducible gene involved in T-lymphocyte activation.

The human 4F2 cell surface antigen is a 120-kilodalton (kDa) disulfide-linked heterodimer which is composed of an 80- to 90-kDa glycosylated heavy chain (4F2HC) and a 35- to 40-kDa nonglycosylated light chain (4F2LC). 4F2 belongs to a family of inducible cell surface molecules which are involved in T-lymphocyte activation and growth. To better understand the molecular mechanism(s) that controls 4F2HC gene expression in both resting and activated T cells, a 4F2HC human genomic clone was isolated and structurally characterized. The 4F2HC gene spans 8 kilobases of chromosome 11 and is composed of nine exons. The 5' upstream region of the gene displays several properties which are characteristic of housekeeping genes. It is G+C rich and hypomethylated in peripheral blood lymphocyte DNA and contains multiple binding sites for the Sp1 transcription factor while lacking TATA or CCAAT sequences. This region of the gene also displays sequence homologies with several other inducible T-cell genes, including the interleukin-2, interleukin-2 receptor alpha chain, dihydrofolate reductase, thymidine kinase, and transferrin receptor genes. A 255-base-pair fragment of the 4F2HC gene which contains 154 base pairs of the 5' flanking sequence was able to efficiently promote expression of the bacterial chloramphenicol acetyltransferase gene in human Jurkat T cells, indicating that it contains promoter or enhancer (or both) sequences. Analyses of chromatin structure in resting and lectin-activated T cells revealed the presence of stable DNase I-hypersensitive sites within both the 5' flanking and intron 1 regions of the 4F2HC gene. Although the 4F2HC gene displayed many of the structural features characteristic of a constitutively expressed gene, lectin-mediated activation of resting peripheral blood T lymphocytes resulted in a dramatic increase in steady-state levels of 4F2HC mRNA.

Amino Acid Sequence

Molecular cloning of Ly-1, a membrane glycoprotein of mouse T lymphocytes and a subset of B cells: molecular homology to its human counterpart Leu-1/T1 (CD5).

We report the isolation of cDNA clones of the mouse lymphocyte differentiation antigen Ly-1. One of these cDNA clones was confirmed to be full-length by DNA sequencing and by expression of Ly-1 by L cells transfected with this clone. Analysis of the predicted amino acid sequence indicated that the Ly-1 polypeptide is synthesized with a 23 amino acid leader and that the mature protein consists of an amino-terminal region of 347 amino acids, a transmembrane sequence of 30 residues, and a carboxyl-terminal region of 94 amino acids. The amino-terminal region appears to be divided into two subregions by a threonine- and proline-rich sequence of 23 amino acids that is highly conserved between Ly-1 and its human homologue Leu-1 (CD5) in position and amino acid composition. The first amino-terminal subregion of 111 amino acids is predicted to be arranged in a beta-pleated sheet structure of six strands. The entire amino-terminal region is rich in cysteine, with all of its 22 cysteine residues conserved between Ly-1 and Leu-1. The carboxyl-terminal region has no cysteines. Ly-1 and Leu-1 are 63% identical, with a gradient of identical residues from 43% for the first amino-terminal subregion to 58% for the second amino-terminal subregion and 90% for the carboxyl-terminal region. The predicted secondary structure of the first amino-terminal subregion and identities of certain conserved residues among most members of the immunoglobulin gene superfamily suggest that Ly-1 and Leu-1 are distant members of this family.

Amino Acid Sequence

Molecular cloning of complementary DNAs encoding the heavy chain of the human 4F2 cell-surface antigen: a type II membrane glycoprotein involved in normal and neoplastic cell growth.

Complementary DNA (cDNA) clones encoding the heavy chain of the heterodimeric human membrane glycoprotein 4F2 have been isolated by immunoscreening of a lambda gt11 expression library. The identity of these clones has been confirmed by hybridization to RNA and DNA prepared from mouse L-cell transfectants, which were produced by whole cell gene transfer and selected for cell-surface expression of the human 4F2 heavy chain. DNA sequence analysis suggests that the 4F2 heavy-chain cDNAs encode an approximately 526-amino acid type II membrane glycoprotein, which is composed of a large C-terminal extracellular domain, a single potential transmembrane region, and a 50-81 amino acid N-terminal intracytoplasmic domain. Southern blotting experiments have shown that the 4F2 heavy-chain cDNAs are derived from a single-copy gene that has been highly conserved during mammalian evolution.

Amino Acid Sequence

An evaluation of an enzymatic choline determination for the identification of semen in casework samples.

This study compares the detection of choline in seminal stains by both an enzymatic method and by the standard Florence crystal test. The tests were conducted on 293 actual casework samples. In those samples identified as containing semen, choline was detected twice as often by the enzymatic method compared to the Florence method (84.6 versus 40.3%). The choline results were correlated with spermatozoa and acid phosphatase tests. The enzymatic detection of choline in seminal stains was found to be a fast, easy, sensitive, and reliable test.

Acid Phosphatase

Isolation of complementary DNA clones encoding the human lymphocyte glycoprotein T1/Leu-1.

The T1/Leu-1/CD5 molecule, a human T-cell surface glycoprotein of relative molecular mass (Mr) 67,000, has been implicated in the proliferative response of activated T cells and in T-cell helper function. A similar involvement in T-cell proliferation has been reported for Ly-1, the murine homologue of T1. Here we report the complete amino-acid sequence of the T1 precursor molecule deduced from complementary DNA clones. The protein contains a classical signal peptide; a 347-amino-acid extracellular segment; a transmembrane region; and a 93-amino-acid intracellular segment. The extracellular segment contains many cysteine residues and is composed of two related structural domains separated by a proline/threonine-rich region. The T1 molecule has structural features characteristic of other receptor molecules.

Amino Acid Sequence