Abstract
A quantitative polymerase chain reaction assay was developed that allowed us to monitor transcript levels corresponding to individual members of the cytotoxic cell proteinase (CCP) gene family during T cell activation. Selective expression was observed and shown to depend upon the mode of T cell antigen receptor stimulation. Mitogen or allogeneic stimulation of cells resulted in the appearance of transcripts corresponding to all the genes measured, whereas alpha CD3 antibody produced a response restricted to just two family members. This differential gene activation represents a heterogeneity in cytotoxic T lymphocytes that has not been recognized previously. It may indicate that the T cell branch of the immune system can distinguish between different forms of stimulation and respond by synthesizing a specific set of effector and ancillary molecules that is most appropriate for lysis of cells bearing that type of antigen. Only CCP1 transcripts correlated with cytotoxicity for all modes of stimulation. The patterns for the others are suggestive of distinct and ancillary, rather than direct effector, roles in the lytic mechanism.
Highlights
A quantitative polymerase chain reaction assay was developed that allowed us to monitor transcript levels corresponding to individual members of the cytotoxic cell proteinase (CCP) gene family during T cell activation
On the basis of similarities in amino acid and nucleotide sequences all but one belong to a closely related family that we have named cytotoxic cell proteinases (CCP; known as granzymes B-G) (Haddad et al, 1990).The genomic organization of CCPl to CCP5 genes has been determined and found to be almost identical amongst these family members (Lobe et al, 1988; Prendergast et al, 1991).In addition, the CCPl to CCP4 genes have been mapped close to the T cell antigen receptor a-chain locus at region 11of chromosome 14 (Crosby et al, 1990).The two human CCPs that have been mapped were found at this locus in human metaphase spreads (Lin et al, 1990)
Hanukah factor has a completely different genomic organization and is located on chromosome 5, and was not given the CCP designation (Gershenfeld et Cytotoxic T lymphocytes (CTL)' and natural killer cells play a critical role in the destruction of neoplastic and virally infected cells (Berke, 1987).The active components of these cytotoxic cells arecontained within the granules that are delivered by exocytosis to the surface of the target cell (reviewedby Joag et al (1989))
Summary
A quantitative polymerase chain reaction assay was developed that allowed us to monitor transcript levels corresponding to individual members of the cytotoxic cell proteinase (CCP) gene family during T cell activation. Mitogen or allogeneic stimulation of cells resulted in the appearance of transcripts corresponding to all the genes measured, whereas aCD3 antibody produced aresponse restricted to just two family members This differential gene activation represents a heterogeneity in cytotoxic T lymphocytes that has not been recognized previously. Cathepsin andat least one member of the latter group appear to map to the same chromosomal cluster as theCCP family (Hanson et al, 1990).these serine proteinases constitute a multigene family residing on chromosome 14, and most likely arising from gene duplication events These proteinases appear to be differentially expressed, i.e. CCPs in cytotoxic lymphocytes,cathepsin in neutrophils, and mast cell proteinase in mast cells. The abbreviations used are: CTL, cytotoxic T lymphocyte; CCP, cytotoxic cell proteinase; ConA, concanavalin A; IL2,interleukin 2; PCR, polymerase chain reaction; TCR, T cell receptor for antigen; Hepes, 4-(2-hydroxyethyl)-l-piperazineethanesulfoniaccid; kb, kilobase pair(s)
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