Isolation and characterization of a cDNA encoding a putative cytokine which is induced by stimulation via the CD2 structure on human T lymphocytes

To define activation-specific sequences in human T lymphocytes, a cDNA library was constructed by subtractive hybridization using resting and stimulated peripheral blood lymphocyte pairs. Stimulation of peripheral blood lymphocytes was achieved by triggering with mitogenic anti-CD2 (T11) monoclonal antibodies. Differential library screening with cDNA probes derived from stimulated vs. resting peripheral blood lymphocytes led to identification of a novel sequence, termed HC21. The predicted primary and secondary structure of HC21 deduced from the translated nucleotide sequence suggest that the gene encodes a secreted protein of 92 amino acids including a 23-residue leader sequence. Northern blot analysis demonstrates that HC21 mRNA is induced in peripheral blood T lymphocytes and interleukin 2-dependent T cell clones within 10 min following stimulation via CD2, while steady-state RNA expression is maximal by 2 h. Although the kinetics of HC21 expression are similar after stimulation via the antigen/major histocompatibility complex receptor (CD3-Ti), CD3-Ti triggering leads to less accumulation of HC21 RNA than CD2 triggering. In contrast, recombinant interleukin 2 does not induce detectable HC21 RNA expression in T cell clones. Transient expression of the HC21 cDNA in COS cells results in a readily detectable protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of radiolabeled cell supernatants consistent with HC21 encoding a secreted product. Protein sequence analysis reveals a striking homology (up to 76% identity at amino acid level) with other members of a recently described lymphokine family whose functions are yet to be defined.