TWENTY-SECOND ANNUAL EASTERN FISH HEALTH WORKSHOP ROYAL PAVILLION RESORT, ATLANTIC BEACH, NC MARCH 18 - 20, 1997 Molecular Properties of a Nonspecific Cytotoxic Cell Receptor Protein Liliana Jaso-Friedmann and Donald L. Evans Department of Medical Microbiology, University of Georgia, Athens, GA. USA Nonspecific cytotoxic cells (NCC) may be the teleost equivalent of lymphokine activated NK cells. We have previously identified a protein on NCC membranes (NCC receptor protein/NCCRP-1) which may be involved in the recognition of (conventional) tumor cell and protozoan antigens. Binding to NCCRP-1 by monoclonal antibody 5C6, by target cell antigen or by cognate synthetic peptide initiates a signalling response leading to increased cytotoxicity. In the present study, three 20-mer microsequences were obtained from tryptic digests of purified NCCRP-1. Degenerate primers were synthesized (based on each peptide sequence) and were used for RT-PCR with mRNA purified from homogeneous NCC populations. An NCCRP-1 specific cDNA sequence was used to synthesize nondegenerate primers. These primers were used in a 5'/3' RACE PCR to obtain the entire NCCRP-1 specific cDNA. A deduced amino acid (aa) sequence consisted of 235 aa. The identity of the protein was confirmed by finding the previously microsequenced peptides in the derived sequence. Homology searches revealed that NCCRP-1 is a novel protein. Northern blot analysis of mRNA content from teleost NCC, B-cells and T-cells revealed only one band in NCC preparations. Functional studies demonstrated a decrease in membrane NCCRP-1 expression and inhibition of NCC cytotoxicity following treatment with NCCRP-1 anti-sense oligonucleotides. Treatment of NCC with sense oligonucleotides had no inhibitory effects on cytotoxicity. An algorithm predicting the membrane conformation of NCCRP-1 suggests one extracellular proline rich (16.8%) domain, a transmembrane portion of 15-18 aa and a cytoplasmic tail composed of a high frequency (14.8%) of serine and tyrosine residues. Current studies suggest that NCC and NCCRP-1 may participate in innate resistance functions in teleost fish. Return to 22nd Annual Eastern Fish Health Workshop Return to Leetown Science Center Home Page