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Male-killing symbiont damages host's dosage-compensated sex chromosome to induce embryonic apoptosis

View Article: PubMed Central - PubMed

ABSTRACT

Some symbiotic bacteria are capable of interfering with host reproduction in selfish ways. How such bacteria can manipulate host's sex-related mechanisms is of fundamental interest encompassing cell, developmental and evolutionary biology. Here, we uncover the molecular and cellular mechanisms underlying Spiroplasma-induced embryonic male lethality in Drosophila melanogaster. Transcriptomic analysis reveals that many genes related to DNA damage and apoptosis are up-regulated specifically in infected male embryos. Detailed genetic and cytological analyses demonstrate that male-killing Spiroplasma causes DNA damage on the male X chromosome interacting with the male-specific lethal (MSL) complex. The damaged male X chromosome exhibits a chromatin bridge during mitosis, and bridge breakage triggers sex-specific abnormal apoptosis via p53-dependent pathways. Notably, the MSL complex is not only necessary but also sufficient for this cytotoxic process. These results highlight symbiont's sophisticated strategy to target host's sex chromosome and recruit host's molecular cascades toward massive apoptosis in a sex-specific manner.

No MeSH data available.


DNA damage and apoptosis in Spiroplasma-infected embryos mutant for stg.(a,b) Spiroplasma-infected control male embryo (genotype stgAR2/TM3 or stg7B/TM3) and stg mutant male embryo (genotype stgAR2/stg7B) at stage 12. Cell membranes and apoptotic cells are visualized by anti-Discs large (Dlg; green) and TUNEL (magenta) staining, respectively. Single-channel images of TUNEL staining and high magnification images of Dlg staining are shown in a′–b′ and a′′–b′′, respectively. (c) Quantification of TUNEL-positive areas in infected control and stg mutant embryos. Box plots are as in Fig. 2i,j. Different letters (a–d) indicate statistically significant differences (P<0.05 for stg females versus males, P<0.01 for the others; Kruskal–Wallis test followed by Mann–Whitney U-tests). Sample sizes are indicated at the bottom. (d,e) Infected stg mutant female (n=7) and male (n=11) embryos at stage 12, in which DNA damage (pH2Av; green), the X chromosome (MSL1; magenta), and cell membrane (DCAT-1; blue) are visualized. Single-channel images of pH2Av signals and MSL1 signals are shown in d′–e′ and d′′–e′′, respectively. Boxed regions in d and e are magnified in insets of d–d′′ and e–e′′. Scale bars, 100 μm (a–b′), 25 μm (a′′,b′′), 20 μm (d–e′′) and 5 μm (insets in d–e′′).
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f6: DNA damage and apoptosis in Spiroplasma-infected embryos mutant for stg.(a,b) Spiroplasma-infected control male embryo (genotype stgAR2/TM3 or stg7B/TM3) and stg mutant male embryo (genotype stgAR2/stg7B) at stage 12. Cell membranes and apoptotic cells are visualized by anti-Discs large (Dlg; green) and TUNEL (magenta) staining, respectively. Single-channel images of TUNEL staining and high magnification images of Dlg staining are shown in a′–b′ and a′′–b′′, respectively. (c) Quantification of TUNEL-positive areas in infected control and stg mutant embryos. Box plots are as in Fig. 2i,j. Different letters (a–d) indicate statistically significant differences (P<0.05 for stg females versus males, P<0.01 for the others; Kruskal–Wallis test followed by Mann–Whitney U-tests). Sample sizes are indicated at the bottom. (d,e) Infected stg mutant female (n=7) and male (n=11) embryos at stage 12, in which DNA damage (pH2Av; green), the X chromosome (MSL1; magenta), and cell membrane (DCAT-1; blue) are visualized. Single-channel images of pH2Av signals and MSL1 signals are shown in d′–e′ and d′′–e′′, respectively. Boxed regions in d and e are magnified in insets of d–d′′ and e–e′′. Scale bars, 100 μm (a–b′), 25 μm (a′′,b′′), 20 μm (d–e′′) and 5 μm (insets in d–e′′).

Mentions: In an attempt to gain further insight into the relationship between DNA damage, chromosomal breakage and abnormal apoptosis, we genetically blocked cell division during embryogenesis. String (Stg), a CDC25 homologue of Drosophila, is essential for the initiation of G2/M transition in the cell cycle52. In zygotic mutants of strong alleles of stg, embryonic cells initially undergo normal cleavage cycles by using maternal transcripts during mitoses 1–13, and after cellularization when zygotically regulated cell division starts (from mitosis 14 onward), cells are arrested at G2 phase during the rest of embryogenesis, thereby resulting in embryos with fewer and larger cells52 (Fig. 6a,b). Considering that the recruitment of the MSL complex to the male X chromosome is first detected in cellularized embryos at mitosis 14 (refs 53, 54), embryonic cells mutant for stg do not undergo cell division after the formation of the MSL complex, which is required for Spiroplasma-induced DNA damage. Therefore, using stg mutant embryos, we can genetically dissect whether bridge breakage in the male X chromosome has a causative role for the chromosome-specific DNA damage induced by Spiroplasma.


Male-killing symbiont damages host's dosage-compensated sex chromosome to induce embryonic apoptosis
DNA damage and apoptosis in Spiroplasma-infected embryos mutant for stg.(a,b) Spiroplasma-infected control male embryo (genotype stgAR2/TM3 or stg7B/TM3) and stg mutant male embryo (genotype stgAR2/stg7B) at stage 12. Cell membranes and apoptotic cells are visualized by anti-Discs large (Dlg; green) and TUNEL (magenta) staining, respectively. Single-channel images of TUNEL staining and high magnification images of Dlg staining are shown in a′–b′ and a′′–b′′, respectively. (c) Quantification of TUNEL-positive areas in infected control and stg mutant embryos. Box plots are as in Fig. 2i,j. Different letters (a–d) indicate statistically significant differences (P<0.05 for stg females versus males, P<0.01 for the others; Kruskal–Wallis test followed by Mann–Whitney U-tests). Sample sizes are indicated at the bottom. (d,e) Infected stg mutant female (n=7) and male (n=11) embryos at stage 12, in which DNA damage (pH2Av; green), the X chromosome (MSL1; magenta), and cell membrane (DCAT-1; blue) are visualized. Single-channel images of pH2Av signals and MSL1 signals are shown in d′–e′ and d′′–e′′, respectively. Boxed regions in d and e are magnified in insets of d–d′′ and e–e′′. Scale bars, 100 μm (a–b′), 25 μm (a′′,b′′), 20 μm (d–e′′) and 5 μm (insets in d–e′′).
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f6: DNA damage and apoptosis in Spiroplasma-infected embryos mutant for stg.(a,b) Spiroplasma-infected control male embryo (genotype stgAR2/TM3 or stg7B/TM3) and stg mutant male embryo (genotype stgAR2/stg7B) at stage 12. Cell membranes and apoptotic cells are visualized by anti-Discs large (Dlg; green) and TUNEL (magenta) staining, respectively. Single-channel images of TUNEL staining and high magnification images of Dlg staining are shown in a′–b′ and a′′–b′′, respectively. (c) Quantification of TUNEL-positive areas in infected control and stg mutant embryos. Box plots are as in Fig. 2i,j. Different letters (a–d) indicate statistically significant differences (P<0.05 for stg females versus males, P<0.01 for the others; Kruskal–Wallis test followed by Mann–Whitney U-tests). Sample sizes are indicated at the bottom. (d,e) Infected stg mutant female (n=7) and male (n=11) embryos at stage 12, in which DNA damage (pH2Av; green), the X chromosome (MSL1; magenta), and cell membrane (DCAT-1; blue) are visualized. Single-channel images of pH2Av signals and MSL1 signals are shown in d′–e′ and d′′–e′′, respectively. Boxed regions in d and e are magnified in insets of d–d′′ and e–e′′. Scale bars, 100 μm (a–b′), 25 μm (a′′,b′′), 20 μm (d–e′′) and 5 μm (insets in d–e′′).
Mentions: In an attempt to gain further insight into the relationship between DNA damage, chromosomal breakage and abnormal apoptosis, we genetically blocked cell division during embryogenesis. String (Stg), a CDC25 homologue of Drosophila, is essential for the initiation of G2/M transition in the cell cycle52. In zygotic mutants of strong alleles of stg, embryonic cells initially undergo normal cleavage cycles by using maternal transcripts during mitoses 1–13, and after cellularization when zygotically regulated cell division starts (from mitosis 14 onward), cells are arrested at G2 phase during the rest of embryogenesis, thereby resulting in embryos with fewer and larger cells52 (Fig. 6a,b). Considering that the recruitment of the MSL complex to the male X chromosome is first detected in cellularized embryos at mitosis 14 (refs 53, 54), embryonic cells mutant for stg do not undergo cell division after the formation of the MSL complex, which is required for Spiroplasma-induced DNA damage. Therefore, using stg mutant embryos, we can genetically dissect whether bridge breakage in the male X chromosome has a causative role for the chromosome-specific DNA damage induced by Spiroplasma.

View Article: PubMed Central - PubMed

ABSTRACT

Some symbiotic bacteria are capable of interfering with host reproduction in selfish ways. How such bacteria can manipulate host's sex-related mechanisms is of fundamental interest encompassing cell, developmental and evolutionary biology. Here, we uncover the molecular and cellular mechanisms underlying Spiroplasma-induced embryonic male lethality in Drosophila melanogaster. Transcriptomic analysis reveals that many genes related to DNA damage and apoptosis are up-regulated specifically in infected male embryos. Detailed genetic and cytological analyses demonstrate that male-killing Spiroplasma causes DNA damage on the male X chromosome interacting with the male-specific lethal (MSL) complex. The damaged male X chromosome exhibits a chromatin bridge during mitosis, and bridge breakage triggers sex-specific abnormal apoptosis via p53-dependent pathways. Notably, the MSL complex is not only necessary but also sufficient for this cytotoxic process. These results highlight symbiont's sophisticated strategy to target host's sex chromosome and recruit host's molecular cascades toward massive apoptosis in a sex-specific manner.

No MeSH data available.