Figure 1. trr-1
Cloning and Expression. (A) The genetic map position of trr-1
) is shown above. The trr-1
gene structure as derived from cDNA and genomic sequences is indicated below. Shaded boxes indicate coding sequence and open boxes indicate 5' and 3' untranslated regions. Predicted translation initiation and termination codons and the poly(A) tail are shown. Positions of alternative splicing are indicated by asterisks. In all cases, the use of alternative splice acceptors would create small differences in the trr-1
coding sequence: alternative splicings of the fourth (ag/TTTCAGAC versus agtttcag/AC), fifth (ag/AATCTTCAGTC versus agaatcttcag/TC), eleventh (ag/AACTTTAAGAT versus agaactttaag/AT), and twelfth introns (ag/TTGCAGAA versus agttgcag/AA) would result in differences of two or three amino acids.(B) A schematic of the TRR-1 protein. All six trr-1
mutations are predicted to prematurely truncate the TRR-1 protein. The positions and natures of these nonsense mutations are indicated above. TRR-1 is similar to mammalian TRRAP and yeast Tra1p throughout the lengths of the proteins. The degree of similarity is especially high in the FAT (FRAP, ATM, TRRAP-like) and ATM/PI-3 kinase-like domains, which are indicated with solid and hatched boxes, respectively.(C-E) TRR-1 is expressed throughout development and is localized to nuclei. (C), a 4-cell stage embryo. (D), an L2 larva indicating TRR-1 expression in P4.p, P5.p, and P6.p (arrowheads). (E), a germline oocyte nucleus in an adult hermaphrodite. In (C) and (D), anterior is to the left and dorsal is up. To more clearly depict the weak expression of TRR-1 in somatic postembryonic cells in (D), we improved the signal-to-noise ratio of our initially captured image by adjusting pixel threshold values and applying a Gaussian filter using Adobe Photoshop 5.5. Scale bars, 5 um.(F-H) 4,6-Diamidino-2-phenylindole (DAPI) staining of the same samples shown in (C)-(E). In (E), P4.p, P5.p, and P6.p were identified by their positions and large, DAPI-negative nucleoli. In (F), the anterior-most cell, AB.a, and the dorsal-most cell, AB.p, have condensed mitotic chromosomes, which by comparison with (C) are not sites of concentrated TRR-1 localization. The discrete foci of DAPI staining in the oocyte nucleus shown in (H) indicate condensed meiotic chromosomes, and a comparison with (E) indicates localization of TRR-1 to these chromosomes. Scale bars, 5 um.