T cell expression cloning of a Mycobacterium tuberculosis gene encoding a protective antigen associated with the early control infection

Infection of C57BL/6 mice with Mycobacterium tuberculosis results in the de velopment of a progressive disease during the first 2 wk after challenge. T hereafter, the disease is controlled by the emergence of protective T cells . We have used this infection model in conjunction with direct T cell expre ssion cloning to identify Ags involved with the early control of the diseas e, A protective M. tuberculosis-specific CD4 T cell line derived from mice at 3 wk postchallenge was used to directly screen an ill, tuberculosis geno mic expression library. This screen resulted in the identification of a gen omic clone comprising two putative adjacent genes with predicted open readi ng frames of 10 and 41 kDa, MTB10 and MTB41, respectively (the products of Rv0916c and Rv0915c, respectively, in the TubercuList H37Rv database). MTB1 0 and MTB41 belong to the PE and PPE family of proteins recently identified to comprise 10% of the M. tuberculosis genome. Evaluation of the recombina nt proteins revealed that MTB41, but not MTB10, is the Ag recognized by the cell line and by M tuberculosis-sensitized human PBMC, Moreover, C57BL/6 m ice immunized with MTB41 DNA developed both CD4- (predominantly Th1) and CD 8-specific T cell responses to rMTB41 protein, More importantly, immunizati on of C57BL/6 mice with MTB41 DNA induced protection against infection with AL tubercrtlosis comparable to that induced by bacillus Calmette-Guerin. T hus, the use of a proven protective T cell line in conjunction with the T c ell expression cloning approach resulted in the identification of a candida te Ag for a subunit vaccine against tuberculosis.