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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.

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Related in: MedlinePlus

RNA-seq analysis of Spiroplasma-infected and uninfected embryos.(a) Epithelial cells of an infected female embryo at stage 11. Three DNA-stained z-sections are projected to show the surface of the epithelium. Boxed regions in a are magnified in a′ and a′′ to highlight Spiroplasma cells (arrows). Scale bars, 20 μm (a) and 5 μm (a′,a′′). (b) Expression of Sxl-Pe-EGFP in uninfected embryos at stage 10–11. Only female embryos express GFP. Bottom panel shows abbreviations for RNA-seq samples. (c) The number of differentially expressed genes identified in pairwise comparisons between RNA-seq samples. In UF (uninfected female) versus UM (uninfected male), for example, numbers of differently expressed genes up- or down-regulated in UM compared with UF are plotted (the same applies hereafter). (d) A heat map of selected 320 differentially expressed genes. On the top is a clustering dendrogram of RNA-seq samples based on similarity of gene expression patterns across the samples. On the left is a clustering dendrogram of differentially expressed genes based on similarity of gene expression patterns across the genes, wherein gene clusters #1–#6 are depicted by different colours. (e) GO categories enriched in gene clusters #1–#6. (f) Expression levels (fragments per kilobase per million, FPKM) of major genes related to apoptosis (left) and DNA damage response (right) categorized to the gene cluster #1, represented as mean±s.d. of three independent experiments.
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f1: RNA-seq analysis of Spiroplasma-infected and uninfected embryos.(a) Epithelial cells of an infected female embryo at stage 11. Three DNA-stained z-sections are projected to show the surface of the epithelium. Boxed regions in a are magnified in a′ and a′′ to highlight Spiroplasma cells (arrows). Scale bars, 20 μm (a) and 5 μm (a′,a′′). (b) Expression of Sxl-Pe-EGFP in uninfected embryos at stage 10–11. Only female embryos express GFP. Bottom panel shows abbreviations for RNA-seq samples. (c) The number of differentially expressed genes identified in pairwise comparisons between RNA-seq samples. In UF (uninfected female) versus UM (uninfected male), for example, numbers of differently expressed genes up- or down-regulated in UM compared with UF are plotted (the same applies hereafter). (d) A heat map of selected 320 differentially expressed genes. On the top is a clustering dendrogram of RNA-seq samples based on similarity of gene expression patterns across the samples. On the left is a clustering dendrogram of differentially expressed genes based on similarity of gene expression patterns across the genes, wherein gene clusters #1–#6 are depicted by different colours. (e) GO categories enriched in gene clusters #1–#6. (f) Expression levels (fragments per kilobase per million, FPKM) of major genes related to apoptosis (left) and DNA damage response (right) categorized to the gene cluster #1, represented as mean±s.d. of three independent experiments.

Mentions: We collected Spiroplasma-infected and uninfected Drosophila embryos of both sexes at stage 10–11 when infection-associated male-specific abnormal apoptosis starts26 (Fig. 1a). For embryonic sexing, we used a transgenic strain with green fluorescence protein (GFP) reporter of Sxl gene, Sxl-Pe-EGFP, which expresses GFP only in females (Fig. 1b and Supplementary Fig. 1a). Four groups of pooled embryos (uninfected females, uninfected males, infected females and infected males; three replicates for each group) were subjected to RNA-sequencing (RNA-seq) analysis. Of all the genes annotated in the Drosophila genome, 8,387 genes were substantially expressed in the embryos (Supplementary Methods), of which we identified 1,430 differentially expressed genes by all pairwise comparisons between the groups (false discovery rate<0.001). Notably, more differentially expressed genes were associated with infected male embryos than other groups (Fig. 1c and Supplementary Fig. 1b). In infected male embryos, up-regulated genes were concentrated on the second and third chromosomes, whereas down-regulated genes were preferentially found on the X chromosome (Supplementary Fig. 1c). At a glance, this pattern may look like reflecting dosage compensation defects in infected male embryos. However, comparison with uninfected male embryos revealed only a small number of down-regulated X-encoded genes in infected male embryos (51 of 1,447 analyzed genes encoded on the X chromosome) (Supplementary Fig. 1c), suggesting that dosage compensation is still functioning in infected male embryos.


Male-killing symbiont damages host's dosage-compensated sex chromosome to induce embryonic apoptosis
RNA-seq analysis of Spiroplasma-infected and uninfected embryos.(a) Epithelial cells of an infected female embryo at stage 11. Three DNA-stained z-sections are projected to show the surface of the epithelium. Boxed regions in a are magnified in a′ and a′′ to highlight Spiroplasma cells (arrows). Scale bars, 20 μm (a) and 5 μm (a′,a′′). (b) Expression of Sxl-Pe-EGFP in uninfected embryos at stage 10–11. Only female embryos express GFP. Bottom panel shows abbreviations for RNA-seq samples. (c) The number of differentially expressed genes identified in pairwise comparisons between RNA-seq samples. In UF (uninfected female) versus UM (uninfected male), for example, numbers of differently expressed genes up- or down-regulated in UM compared with UF are plotted (the same applies hereafter). (d) A heat map of selected 320 differentially expressed genes. On the top is a clustering dendrogram of RNA-seq samples based on similarity of gene expression patterns across the samples. On the left is a clustering dendrogram of differentially expressed genes based on similarity of gene expression patterns across the genes, wherein gene clusters #1–#6 are depicted by different colours. (e) GO categories enriched in gene clusters #1–#6. (f) Expression levels (fragments per kilobase per million, FPKM) of major genes related to apoptosis (left) and DNA damage response (right) categorized to the gene cluster #1, represented as mean±s.d. of three independent experiments.
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Related In: Results  -  Collection

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f1: RNA-seq analysis of Spiroplasma-infected and uninfected embryos.(a) Epithelial cells of an infected female embryo at stage 11. Three DNA-stained z-sections are projected to show the surface of the epithelium. Boxed regions in a are magnified in a′ and a′′ to highlight Spiroplasma cells (arrows). Scale bars, 20 μm (a) and 5 μm (a′,a′′). (b) Expression of Sxl-Pe-EGFP in uninfected embryos at stage 10–11. Only female embryos express GFP. Bottom panel shows abbreviations for RNA-seq samples. (c) The number of differentially expressed genes identified in pairwise comparisons between RNA-seq samples. In UF (uninfected female) versus UM (uninfected male), for example, numbers of differently expressed genes up- or down-regulated in UM compared with UF are plotted (the same applies hereafter). (d) A heat map of selected 320 differentially expressed genes. On the top is a clustering dendrogram of RNA-seq samples based on similarity of gene expression patterns across the samples. On the left is a clustering dendrogram of differentially expressed genes based on similarity of gene expression patterns across the genes, wherein gene clusters #1–#6 are depicted by different colours. (e) GO categories enriched in gene clusters #1–#6. (f) Expression levels (fragments per kilobase per million, FPKM) of major genes related to apoptosis (left) and DNA damage response (right) categorized to the gene cluster #1, represented as mean±s.d. of three independent experiments.
Mentions: We collected Spiroplasma-infected and uninfected Drosophila embryos of both sexes at stage 10–11 when infection-associated male-specific abnormal apoptosis starts26 (Fig. 1a). For embryonic sexing, we used a transgenic strain with green fluorescence protein (GFP) reporter of Sxl gene, Sxl-Pe-EGFP, which expresses GFP only in females (Fig. 1b and Supplementary Fig. 1a). Four groups of pooled embryos (uninfected females, uninfected males, infected females and infected males; three replicates for each group) were subjected to RNA-sequencing (RNA-seq) analysis. Of all the genes annotated in the Drosophila genome, 8,387 genes were substantially expressed in the embryos (Supplementary Methods), of which we identified 1,430 differentially expressed genes by all pairwise comparisons between the groups (false discovery rate<0.001). Notably, more differentially expressed genes were associated with infected male embryos than other groups (Fig. 1c and Supplementary Fig. 1b). In infected male embryos, up-regulated genes were concentrated on the second and third chromosomes, whereas down-regulated genes were preferentially found on the X chromosome (Supplementary Fig. 1c). At a glance, this pattern may look like reflecting dosage compensation defects in infected male embryos. However, comparison with uninfected male embryos revealed only a small number of down-regulated X-encoded genes in infected male embryos (51 of 1,447 analyzed genes encoded on the X chromosome) (Supplementary Fig. 1c), suggesting that dosage compensation is still functioning in infected male embryos.

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.


Related in: MedlinePlus