Figure 7. Actin gene expression in oocytes, early embryos, and adult muscle; motility defects in
act-2(
or295) adults. (A) Amplification of actin gene transcripts from isolated oocytes. Nested RT-PCR was performed using gene-specific 3'UTR primer sequences (see Materials and Methods). (B) Immunofluorescence photomicrographs of fixed wild-type and mutant embryos stained with fluorescently labeled antibodies that recognize actin (red) or tubulin (green); DNA (blue) was labeled with TOTO3. Confocal laser strength and antibody concentration were identical in all cases (see Materials and Methods). Little or no actin was detected in
act-1(3'UTR RNAi);
act-3(3'UTR RNAi);
act-2(
ok1229) embryos (i, k, and l). Extra chromosomes in panel k presumably are due to a failure in polar body extrusion during meiosis. (C) Cytoplasmic GFP::ACT-2 expression in live embryos was detected throughout the cytoplasm and at the cortex of all early embryonic cells (a), beginning at about the 20-cell stage (unpublished data), and in epidermal cells during embryonic elongation (b). GFP::ACT-2 also was detected in contractile filaments of adult striated muscle cells (c). Scale bar, 25 µm. (D) Motility defects in
act-2(
or295) adult animals at the restrictive temperature. Animals were placed on individual plates for 6 h before recording images, and track marks were recorded at 400x and 125x magnification, respectively. Also see Supplementary Movies 14 and 15, in which adult worm movements were recorded at 1 frame/s. Motility defects are apparent for
act-2(
or295), whereas
act-2(
or621) mutants resemble wild-type adults in their motility.