Diversity and structure of human T-cell receptor beta-chain variable region genes.

The nucleotide sequences of 27 T-cell receptor beta cDNA clones isolated from a human peripheral lymphocyte library were determined and compared to five additional published sequences. These cDNA clones represent 22 distinct T-cell receptor beta-chain variable region (V beta) gene segment sequences, which fall into 15 different V beta gene subfamilies, each containing six or fewer members. From this analysis, we estimate that the repertoire of expressed human V beta genes is less than 59, apparently much smaller than the immunoglobulin heavy chain and light chain variable region (VH and VL) repertoires. Variability plots comparing these human V beta regions with each other and with published mouse V beta regions provide evidence for only four hypervariable regions homologous to those seen in comparisons of immunoglobulin V regions. Somatic hypermutation appears to be used infrequently, if at all, in these V beta genes.

Characteristics of spontaneously produced and of virus-induced human LuKII cell interferons.

Several human lymphoblastoid cell lines produced significant levels of interferon (IFN) activity in the absence of any IFN induces. We have partially purified spontaneous IFN (SpIFN) from LuKII cells and compared it with the IFN activity produced by the same cells following Sendai virus induction. Virus-induced LuKII IFN reached maximum titers of 1000-6000 reference units/ml at 12 to 15 houts post-induction and produced a heterogeneous electrophoretic profile with major (18,500 dalton) and minor (23,500 dalton) species. Spontaneous LuKII IFN was produced very slowly over several days and reached maximum titers of 100-1000 reference units/ml. Crude SpIFN (25-100 ref. units/ml) was purified to 1-6 X 10(5) ref. units/ml with a 53-80 percent overall recovery, and it consistently migrated as a homogeneous 20,000 dalton band upon SDS gel electrophoresis. Although spontaneously produced and virus-induced lymphoblastoid IFNs differed in their ability to be neutralized by anti-mouse IFN antiserum (18), both types of IFN were only 1 percent cross-reactive on heterologous mouse L cells. We conclude that IFN definitely can be produced by many human lymphoblastoid cell lines in the absence of inducers, and that LuKII SpIFN has different characteristics than the IFNs produced by the same cell line following viral induction. Since it is now known that there are many IFN-alpha genes (3, 10), it seems likely that these cell lines may provide a system for studying the selective expression of one or more of these genes.

Heterogeneity of interferon mRNA species from Sendai virus-induced human lymphoblastoid (Namalva) cells and Newcastle disease virus-induced murine fibroblastoid (L) cells.

Cytoplasmic polyadenylated RNA preparations obtained from Sendai-induced human lymphoblastoid (Namalva) cells and from Newcastle disease virus (NDV)-induced murine (L) cells were denatured in 10-12.5 mM CH3HgOH and then electrophoresed in 2% agarose tube gels containing 10 mM CH3HgOH, the RNA eluted from gel slices and translationally active interferon mRNA species located using the Xenopus oocyte assay. The interferons synthesized were characterized as alpha or beta types based on neutralization tests using specific antisera against human or murine interferon-alpha and interferon-beta. At least two species of mRNA for human interferon-alpha and two for human interferon-beta were detected in RNA from Sendai-induced Namalva cells. These are (approximate mRNA length in parentheses) alpha (1.3 kb), alpha (1.9 kb), beta (1.1 kb) and beta (1.9 kb). Two populations of murine interferon mRNA of lengths approximately 1.4 kb and 3 kb were detected in mRNA preparations from NDV-induced L cells by electrophoresis. However, since the translation products of each of these two populations of mRNA consist of both murine interferon-alpha and murine interferon-beta it is likely that both the 1.4 kb and 3 kb populations contain at least one species each of murine interferon-alpha and murine interferon-beta mRNA.

Spontaneous production of human interferon.

Several established lines of human lymphoblastoid cells were evaluated for abilities to produce interferons. Some cell lines were able to produce interferon when induced with either Newcastle disease virus or Sendai virus, whereas others failed to produce detectable interferon when so induced. However, several cell lines were able to spontaneously produce interferon without induction. Spontaneously produced interferon was liberated by cells only during logarithmic growth phase, reaching levels ranging from about 10 reference units/ml of growth medium for some cell lines to 1000 reference units/ml for others. The interferons produced by induced lymphoblastoid cells and the spontaneously produced interferons were all characterized as type I human leukocyte interferon by high levels of cross-species antiviral activities on bovine cells and by neutralizations by antiserum to human leukocyte interferon but not by antiserum to human fibroblast interferon. However, analysis by electrophoresis in sodium dodecyl sulfate/polyacrylamide gels revealed that spontaneously produced interferon was less size heterogeneous than human leukocyte interferon, migrating as a single band of activity with a peak at 20,000 daltons, whereas human leukocyte interferon contained peaks of major activity at 23,000 and 18,000 daltons and virus-induced Namalva lymphoblastoid cell interferon migrated predominantly as the 18,000-dalton form. Also, although neither virus-induced primary leukocyte interferon nor any of the virus-induced lymphoblastoid cell interferons were neutralized by antiserum to mouse interferon, all of the spontaneously produced interferons were neutralized by antiserum to mouse interferons. These data suggest significant structural similarities between the active cores of certain interferons from phylogenetically diverse animal species.