CHARACTERIZATION OF MONOCLONAL-ANTIBODIES DIRECTED AGAINST THE GP46 OF HUMAN T-CELL LEUKEMIA-VIRUS TYPE-I

Essential HTLV-I biological functions depend on the structural motives of the surface glycoprotein (gp46). Monoclonal antibodies (mAbs) have been generated in order to identify functional regions of gp46. We ob tained three monoclonal antibodies (3F3F10, 4F5F6 and 7G5D8) by immuni zing Balb/c mice with beta-propiolactone Inactivated HTLV-I producing cells and partially purified gp46. The mAbs are of the IgG 1 subclass. They have been characterized by western blot analysis, reactivity wit h HTLV-I and HTLV-II producing cells and ELISA binding assays using sy nthetic peptides. The immunoblot analysis performed with sheets prepar ed with the virus released by HUT 102 and 2060 cells (an HTLV-I virus producing cell line established in our laboratory) indicate that the t hree mAbs recognize a 46 kDa product as did the anti -gp46 mAb 0.5 alp ha (18). Reactivity of the three mAbs with various cell lines was exam ined by indirect immunofluorescence assay. The mAb 7G5D8 stained stron gly the membrane of all HTLV-I producing cells (MT2, C91/PL, HUT102 an d cells of seven lines established in our laboratory and by A. Gessain ); uninfected lymphoid cells (HSB-2, MOLT 4, CEM and PHA activated lym phocytes from normal donors) were negative. Interestingly cells of a H TLV-II producing line (344 MO) were positive. The mAbs 3F3F10 and 4F5F 6 reacted with the same cells as did 7G5D8 but the fluorescence intens ity was much lower than that observed with this later. A long syntheti c peptide corresponding to the immunodominant region of the gp46 defin ed by the amino acids 175-199 and 10-mer peptides overlaping this regi on were used in an approach to identify the recognized epitope(s). The long 175-199 peptide was recognized by the three mAbs. 3F3F10 and 4F5 F6 recognized none of the 10-mer peptides whereas 7G5D8 bound to 186-1 95 and 182-191 peptides. In addition 7G5D8 did not inhibit either sync ytia formation or virus infection. In view of the data concerning the previously described mAbs 0.5 alpha, LAT 27 (5) and KE36-11 (6), our r esults suggest that the epitope recognized by 7G5D8 is different from those recognized by the former ones. As the 183-191 sequence correspon ds to a region in which HTLV-I end HTLV-II harbour six common amino ac ids end two similar ones, this is consistent with the observation that 7G5D8 stained the HTLV-II producing cells 344 MO as well as all HTLV- I producing ones. Altogether our data support the hypothesis whereby t his epitope recognized by 7G5D8 is contained within a sequence defined by amino acids 183-191.