MEMBRANE-BOUND OR SOLUBLE TRUNCATED RT1.A(A) RAT CLASS-I MAJOR HISTOCOMPATIBILITY ANTIGENS INDUCE SPECIFIC ALLOIMMUNITY

Transfectants that express membrane-bound (IMB) or secrete soluble tru ncated (TR) rat class I RT1.A(a) major histocompatibility (MHC) antige ns induce alloimmunity in vivo. The MB-RT1.A(a) was produced by transf ecting the full-length RT1.A(a) cDNA, including the alpha(1), alpha(2) , and alpha(3), transmembrane and intracellular domains. The TR-RT1.A( a) cDNA insert included only the extracellular (alpha(1), alpha(2), an d alpha(3) domains; a stop codon was placed in front of the transmembr ane domain. Following full-length sequencing, MB-RT1.A(a) and TR-RT1.A (a) cDNAs were translated in vitro into glycosylated MB-RT1.A(a) (45 k Da) and TR-RT1.A(a) (36 kDa) proteins, respectively. Each cDNA constru ct was individually subcloned into the pSG5 vector before transfection into Buffalo (BUF; RT1(b)) hepatoma cells. FACscan analysis with anti -RT1.A(a)-specific R2/15S monoclonal antibody (MAb) confirmed surface expression of RT1.A(a) molecules on the MB-RT1.A(a), but not on the TR -RT1.A(a), transfectants. In contrast, enzyme-linked immunoadsorbent a ssays documented the presence of soluble RT1.A(a) molecules in superna tes from cells transfected with the TR-RT1.A(a), but not from cells tr ansfected with the TR-RT1.A(a), cDNA. Subcutaneous injection of MB-RT1 .A(a) or TR-RT1.A(a) transfectants to BUF or Wistar Furth (WF; RT1(u)) rats induced accelerated rejection of ACI (RT1(a)) but not third-part y Brown Norway (RT1(n)) heart allografts. Furthermore, supernates of T R-RT1.A(a), but not of MB-RT1.A(a), transfectants immunized WF hosts t oward ACI hearts. Thus, both intact MB-RT1.A(a) and soluble TR-RT1.A(a ) class I alloantigens induce potent sensitization against alloantigen s.