CD4-independent signal transduction through the T-cell receptor (TCR/CD3).

The membrane-bound CD4 glycoprotein has been proposed to act like a co-receptor along with the T-cell antigen receptor (TCR/CD3) during ligand recognition and cell activation. Due to its association with the protein tyrosine kinase (PTK) p56lck, CD4 is believed to transduce a signal and support CD3 activation of T cells. In this study we have shown that CD3 ligation on murine T-cell hybridomas induces tyrosine phosphorylation of proteins, including phospholipase C-gamma 1 (PLC gamma 1), both in the presence as well as in the absence of CD4-linked p56lck. Furthermore, using HPB clones deficient in CD3/PTK association, it has been found that the presence of CD4/p56lck does not overcome the defect in signalling. Not even co-aggregation of CD4 with CD3 triggers tyrosine phosphorylation of proteins in these cells. Together, the present results indicate that CD3-linked PTK(s) plays a primary role in the induction of signalling through TCR/CD3, and the presence of CD4/p56lck is neither necessary nor sufficient to elicit these events. In the light of these results a possible role for CD4 in antigen presentation has been proposed.

Advances in HLA genetics.

The major histocompatibility complex (MHC) is a genetic system of over 70 known genes that occupies the midportion of the short arm of the sixth chromosome (C6p) and spans about 4 million base pairs of DNA. The high-resolution typing of class I and class II MHC genes and the identification of genes between and near them has increased the definition of the genetic basis of immune responses and diseases of unknown etiology such as autoimmune diseases in man. Although there are many more genetic systems that participate in the rejection of tissues and in the immune response, the MHC plays a central role in tissue compatibility and immune response against cancer and infectious diseases. In this paper, the authors review evidence about the role of HLA polymorphism in the pathogenesis and development of cancer, infectious diseases, autoimmune diseases and transplantation.

MHC class I antigens regulate CD3-induced tyrosine phosphorylation of proteins in T cells.

The MHC class I antigens enhance the T cell response to various mitogenic stimuli. Class I antigens co-cap and associate with the CD3 structure on these cells. The present work shows that co-aggregation of these MHC antigens with CD3 induces a sustained elevated Ca2+ response in T cells. The duration required for a 50% decline in peak response is five to 10 times longer when class I antigens are cross-linked with CD3 as compared to that when cells are stimulated through CD3 alone. Further analysis reveals that class I antigens prolong the duration of CD3-induced tyrosine phosphorylation of several proteins. No protein tyrosine kinase activity is found associated with these MHC antigens which could explain their influence on CD3 activation. Similarly, class I antigens do not depend on the membrane-bound protein tyrosine phosphatase CD45, since they elevate the degree of CD3-induced Ca2+ response in a CD45-deficient Jurkat cell line. On the contrary, in a CD45- HPB cell line defective in CD3 signaling, co-aggregation of class I antigens with CD3 does not induce Ca2+ flux. Therefore, the effect of class I antigens on CD3 function depends on the ability of CD3 to transduce a signal. Furthermore, cytofluorometric studies show that cross-linking of class I antigens with CD3 inhibits the internalization of the latter. Thus, class I antigens seem to prolong the duration of signal transduction through CD3 by retarding its down-regulation.

New HLA-DR haplotypes containing the DRB6 pseudogene.

An unexpected probe reaction pattern was observed in two samples during HLA-DR typing by PCR-Sequence Specific Oligonucleotide Probes. In order to confirm the unusual typings, samples were analyzed by PCR-Sequence Specific Primers, cloning, and nucleotide sequencing of the second exon of the HLA-DRB-genes. The confirmed DR, DQ phenotype for one sample was DRB1*0701, DRB4*01, DRB5*0101, DRB6*0201, DQB*0602, DQB1*0202. The phenotype of other sample was DRB1*1602, DRB1*1302, DRB3*0301, DRB6*0101, DQB1*0501, DQB1*0502. The first sample has the novel combination of DRB1*0701 with DRB5*0101 and DRB6*0201. The second sample has either DRB6*0101 together with DRB1*1602 in absence of any DRB5 allele or DRB6*0101 together with DRB1*1302, DRB3*0301. We postulate that the most likely haplotype in sample #1 is DRB1*0701; DRB5*0101, DRB1*0602 which could have arisen from gene conversion. The most likely haplotype in sample #2, DRB1*1602, DRB6*0101, DQB1*0502 would have arisen from an homologous recombination event.

Comparison of HLA-A antigen typing by serology with two polymerase chain reaction based DNA typing methods: implications for proficiency testing.

Serology has been routinely used for class I HLA typing for the selection of donors for allotransplantation. However, serology is not adequate for the assignment of all class I specificities especially when testing non-Caucasians subjects and it is necessary to adopt new strategies for routine testing. At the present time the extent of incorrect serologic HLA-A assignments in clinical testing is not known. The polymerase chain reaction (PCR) based techniques have become useful standard clinical typing methods of HLA class II alleles but most laboratories still use serology for class I typing. In this report we have compared two PCR based techniques, PCR amplification with sequence-specific primers (PCR-SSP) and PCR amplification and subsequent hybridization with sequence-specific oligonucleotide probes (PCR-SSOP), for the assignment of HLA-A specificities in 56 blood samples from patients and families serologically typed for HLA-A. This side-by-side comparison of PCR methods showed 100% correlation between them. However, serology showed 7.1% misassignments and, in an additional panel of 19 cells where serology produced equivocal results, the PCR-SSP and SSOP methods identified the correct HLA-A specificity. Our results emphasize the need to complement routine serologic testing of HLA specificities with a small number of primers designed to test HLA-A34, A36, A43, A66, A74 and A80, that are not detected with high precision by serology. We concluded that the PCR-SSP and -SSOP methods can be used in routine HLA-A typing of patients and donors for transplantation with a greater precision than serology.