DRUSE

In the journal DNA Repair (doi:10.1016/j.dnarep.2008.10.007), Minoru Nakayama and colleagues recently reported that loss of RecQ5 leads to spontaneous mitotic defects and chromosomal aberrations in Drosophila melanogaster. Below is the abstract:

“RecQ5 belongs to the RecQ DNA helicase family that includes genes causative of Bloom, Werner, and Rothmund-Thomson syndromes. Although no human disease has been genetically linked to a mutation in RecQ5, Drosophila melanogaster RecQ5 is highly expressed in early embryos, suggesting an important role for it in the DNA metabolism of the early embryo. In this present study, we generated RecQ5 mutants in D. melanogaster. Embryos lacking maternally derived RecQ5 contained irregular nuclei in early embryogenesis. These irregular nuclei emerged in nuclear cycle 11–13, lost cell-cycle markers, and were located below the surface monolayer of nuclei. By time-lapse microscopy, these irregular nuclei were observed not to divide, whereas all neighboring nuclei proceeded through normal mitotic division with synchrony. These data suggest that the irregular nuclei exited from the nuclear division cycle. This phenotype is reminiscent of the effect of X-ray irradiation on wild-type embryos and was rescued by expression of RecQ5. Thus, the maternal supply of RecQ5 is important for the nuclear cycles in syncytical embryos. Furthermore, the frequencies of spontaneous and induced chromosomal aberrations were increased in RecQ5 mutant neuroblasts. These data imply that DNA damage accumulates spontaneously in RecQ5 mutants. Therefore, endogenous genomic damage may be produced in Drosophila development, and RecQ5 would be involved in the maintenance of genomic stability by suppressing the accumulation of DNA damage.”

Fig. 2. Abnormal nuclei during mitosis in syncytial embryos of RecQ5 mutants. Virgin females of wild-type or recq5^D1 mutant flies were crossed with wild-type males. Embryos (0–2 h old) were collected, fixed, and stained for DNA (red) and PH3 (green). Co-localization of red and green signals results in yellow fluorescence. Representative images of embryos (prophase/metaphase) derived from females homozygous for wild-type (A–D) and recq5^D1 mutation (E–H) are shown at cycles 10 (A and E), 11(B and F), 12 (C and G), and 13 (D and H). Each panel shows a field that covers 80% of the surface of a formaldehyde-fixed, PI-stained, anti-PH3-stained embryo. Arrows indicate abnormal nuclei. Scale bar, 10 μm. (I) Frequency of abnormal nuclei at nuclear cycle stages of wild-type (open bars) and recq5^D1 (filled bars) embryos. The number of abnormal nuclei was obtained by scoring the number of PH3-negative nuclei, and the percentage of abnormal nuclei was determined by scoring the number of abnormal nuclei in the total population of nuclei in the cortex of embryos at prophase/metaphase. At least 2000 nuclei were scored in each experiment. Error bars indicate standard deviation from 4 independent experiments. Frequencies of abnormal nuclei between wild-type and recq5^D1 embryos were not significantly different at cycle 10, but these frequencies were significantly different at cycles 11–13 (P < 0.01 in all cases, Student’s t-test) (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

Fig. 3. Mitotic defects in nuclei of RecQ5 mutant embryos. Embryos derived from females [wild-type (A), recq5^D1 (B), recq5^D1/+ (C), recq5^D2 (D), and arm-GAL4/UAS-Recq5qe; recq5^D1 (E)] were fixed and stained for DNA (red) and PH3 (green). (F) X-ray-irradiated wild-type embryos (3 Gy). Arrowheads indicate abnormal nuclei. A and B, anaphase; C–F, prophase/metaphase (inset: magnified picture). Scale bar, 10 μm. (G) Frequency of abnormal nuclei at cycles 11–13 of arm-GAL4/UAS-Recq5qe; recq5^D1 is shown as in Fig. 2I. Frequencies of abnormal nuclei between arm-GAL4/UAS-Recq5qe; recq5^D1 and wild-type embryos were not significantly different, but the frequency for arm-GAL4/UAS-Recq5qe; recq5^D1 was significantly different from that for recq5^D1 (P < 0.01, Student’s t-test) (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

Fig. 4. The abnormal nuclei dropped down into the interior in recq5 mutants. Embryos (prophase) derived from wild-type females (A–C) and those from females homozygous for the recq5^D1 mutation (D–F) were fixed and stained for DNA (red) and PH3 (green). Surface views (A and D) and the corresponding optical cross sections at every 3 μm toward the interior of the embryo (B and C, E and F) are shown. Arrowheads indicate abnormal nuclei. Scale bar, 10 μm (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

Fig. 5. Interphase of wild-type and mutants. Embryos (interphase) derived from wild-type females (A) and those from females homozygous for the recq5^D1 mutation (B) were fixed and stained for DNA (red) and PCNA (green). Arrowheads indicate abnormal nuclei (inset: magnified picture). Scale bar, 10 μm (For interpretation of the references to color in this figure legend, the reader is referred to the web version of the article).

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