An antigen receptor (NCCRP-1) on nonspecific cytotoxic cells is a phosphoprotein associated with the JAK-STAT activation pathway

The antigen receptor (nonspecific cytotoxic cell receptor protein-1/NCCRP-1) on nonspecific cytotoxic cells (NCC) is a 32-kDa predicted Type III membrane protein. The N-terminal cytoplasmic portion of this receptor contains full length and truncated BOX-1 motifs. These motifs are also found on cytokine, erythropoietin and growth hormone receptors and provide ducking sites for JAK kinases, In the present study, we investigated a relationship between NCCRP-1 and JAK2 kinase binding. A possible association with further downstream STAT activation was suggested. NCCR-1 was phosphorylated on C-terminal domain serine residues. To examine the possibility that NCCRP-1 was associated with JAK kinase(s), NCC were purified and lysates were probed by Westen blotting (WB) for the presence of JAK2 kinase. Unlike their mammalian counterparts, NCC JAK? kinase existed as a 90-95-kDa primary and a 35-40-kDa secondary breakdown product. Both mol wt. forms were significantly smaller than those reported for human JAK kinases. To determine if NCCRP-1 was physically associated with JAK? kinase, chemical cross-linking experiments were conducted. NCC membrane preparations were treated with the chemical cross linker DSS, solubilised and immunoprecipitated with anti-NCCRP-1 (e.g., 32 kDa) mab 5C6. WE analysis using anti-JAK2 mab and mat 5C6 demonstrated that the immunoprecipitate contained both the 32-kDa NCCRP-1 and. 85-90-kDa JAK? kinase. To examine further the possibility that STAT proteins may be associated with NCC/NCCRP-1 activation, NCC lysates were probed (WB) with various anti STAT mabs. The strongest signal was produced by a 100-kDa STAT-6 protein. Lysates were negative fur STAT-1, STAT-3 and STAT-5. These data indicate that the N-terminus of NCCRP-1 may initiate cytokine gene transcription by the JAK-STAT signalling pathway. (C) 1999 Elsevier Science Inc.