Figure 1.

Three-hybrid formation with the A.aeolicus β subunit and tRNA^Leu. (A) Schematic presentation of the yeast 3HS adapted to a tRNA bait. The two hybrid proteins are the LexA-MS2 coat protein and the LeuRS β subunit in fusion with the GAL4 activation domain. The hybrid RNA consists of the MS2 RNA and the A.aeolicus tRNA^Leu. Three-hybrid formation leads to expression of the two reporter genes HIS3 and lacZ. (B) Assay of the expression level of HIS3 by drop test. The L40_coat yeast cells containing pACT-LRSB and pIIIA--MS2 were grown on 3-AT-containing medium. The highest permissive concentration of 3-AT (50 mM) was measured for the β subunit with its cognate tRNA (lane 3). The positive control is IRP with IRE (25 mM, lane 6). HIS3 activation was not detectable when the GAL4 activation domain alone was expressed with the A.aeolicus tRNA^Leu bait (lane 1), or when the β subunit was expressed with the MS2 RNA alone (lane 2), or when the β subunit was expressed with the E.coli tRNA^Arg (lane 4), or when the β subunit was proposed to the E.coli tRNA^Leu (lane 5).