Fig. 3. milt mutants differentially express milt isoforms. (A) milt encodes two known transcripts, RA and RB. milt⁹² contains a 2 bp deletion affecting both isoforms. The three class II alleles, k06704, k14514 and EY01559, are all inserted within 100 bp of each other in the first intron of RA. Arrowheads indicate RT-PCR isoform-specific primer sites. (B) Two protein isoforms, Milt PA and PB, differ only at their N termini. Both contain the putative Kinesin-binding site, as uncovered for the mammalian homolog GRIF-1 using yeast two-hybrid analysis. However, some of their predicted HAP-1 domains differ. (C) RT-PCR shows that milt RA and RB are expressed equally in wild type. In milt^EY01559, RA is absent and RB is increased. In milt^{EY01559/k06704}, the ratio of RB to RA is still greatly elevated. (D) A Western blot shows that Milt is overexpressed in milt^EY01559 and milt^EY01559/CyO ovaries. The Milton antibody used recognizes both PA and PB. (E) Model of Milton-mediated bidirectional mitochondrial movement along microtubules. Milt-PA (magenta oval) and possibly Milt-PB (magenta square) link mitochondria to microtubules via the plus-end directed motor Kinesin. Additionally, Milt-PB positively influences the minus-end directed motor Dynein (large arrow).