Fig. 2. mab-2 encodes RNT-1, a Runx transcription factor. (A) (i) him-5(e1490) male tail. (ii) mab-2(e1241); him-5(e1490) male tail. (iii) mab-2(e1241); him-5(e1490); ouEx15[B0414 + rol-6] male tail. Wild-type appearance is restored using this cosmid. (iv) mab-2(e1241); him-5(e1490); ouEx17[B0414.2 + rol-6] male tail. Wild-type appearance is restored using the single gene B0414.2, encoding the Runx transcription factor rnt-1. (v) male tail from a e1241/ok351 trans-heterozygote, showing that these two alleles fail to complement each other. (vi) Male tail from a e1241/os58 trans-heterozygote, again showing non-complementation between the two alleles. Scale bar: 10 µm. Posterior is towards the right in all panels. (B) Schematic representation showing the genomic structure of the rnt-1 gene, the nature and position of the mutations in e1241 and os58 alleles, and the deletion in ok351. The genomic region shown corresponds to the region cloned in the rescuing construct pAW258. The position of the GFP insertion in pAW260 is also indicated. (C) Alignment of Runx genes from various species showing the conserved DNA-binding domain. Identical amino acids are shown in black; similar amino acids are in grey. The Runt domain of rnt-1 (Accession Number O01834) is 51% similar to Drosophila Runt (Accession Number CAA39817), 50% similar to human RUNX1 (Accession Number NP_001745) and 50% similar to mouse Runx1 (Accession Number Q03347). Sequence alignments were performed using ClustalW and processed using BioEdit software. The nature of the mutations in e1241, os58 and ok351 animals are indicated with asterisks. The ok351 deletion removes 10 amino acids of the Runt domain plus 29 amino acids downstream in an in-frame deletion. (D) Bar chart showing a quantitative analysis of ray number in wild-type (n=33), rnt-1(os58) (n=37), rnt-1(e1241) (n=31), rnt-1(ok351) (n=35), rnt-1(RNAi) (n=28) and rnt-1(e1241); ouEx26[rnt-1::GFP+rol-6⁺] (n=35) males. Ray number in all three rnt-1 alleles, as well as in rnt-1(RNAi) males is significantly different from wild type (P<0.0001). Ray number in rnt-1(e1241); Ex[rnt-1::GFP] males is not significantly different from wild type (P>0.05), indicating rescue. Error bars represent the standard error of the mean. (E) Graph showing length growth curves of wild-type and rnt-1(ok351) animals. There is no significant reduction in the length of ok351 animals compared with wild-type animals until the worms have reached adulthood, 4 days after eggs were laid. A 5% decrease in ok351 body length is seen at this time point (n=33 for wild type, 30 for ok351, P<0.00001). Error bars represent the s.e.m.