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 membr ane protein. The N-terminal cytoplasmic portion of this receptor contains f ull length and truncated BOX-1 motifs. These motifs are also found on cytok ine, erythropoietin and growth hormone receptors and provide ducking sites for JAK kinases, In the present study, we investigated a relationship betwe en NCCRP-1 and JAK2 kinase binding. A possible association with further dow nstream STAT activation was suggested. NCCR-1 was phosphorylated on C-termi nal domain serine residues. To examine the possibility that NCCRP-1 was ass ociated with JAK kinase(s), NCC were purified and lysates were probed by We sten 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-kD a secondary breakdown product. Both mol wt. forms were significantly smalle r than those reported for human JAK kinases. To determine if NCCRP-1 was ph ysically associated with JAK? kinase, chemical cross-linking experiments we re conducted. NCC membrane preparations were treated with the chemical cros s linker DSS, solubilised and immunoprecipitated with anti-NCCRP-1 (e.g., 3 2 kDa) mab 5C6. WE analysis using anti-JAK2 mab and mat 5C6 demonstrated th at the immunoprecipitate contained both the 32-kDa NCCRP-1 and. 85-90-kDa J AK? kinase. To examine further the possibility that STAT proteins may be as sociated with NCC/NCCRP-1 activation, NCC lysates were probed (WB) with var ious 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 in dicate that the N-terminus of NCCRP-1 may initiate cytokine gene transcript ion by the JAK-STAT signalling pathway. (C) 1999 Elsevier Science Inc.