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A map of directional genetic interactions in a metazoan cell.

Fischer B, Sandmann T, Horn T, Billmann M, Chaudhary V, Huber W, Boutros M - Elife (2015)

Bottom Line: Gene-gene interactions shape complex phenotypes and modify the effects of mutations during development and disease.The effects of statistical gene-gene interactions on phenotypes have been used to assign genes to functional modules.Our study presents a powerful approach for reconstructing directional regulatory networks and provides a resource for the interpretation of functional consequences of genetic alterations.

View Article: PubMed Central - PubMed

Affiliation: Genome Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
Gene-gene interactions shape complex phenotypes and modify the effects of mutations during development and disease. The effects of statistical gene-gene interactions on phenotypes have been used to assign genes to functional modules. However, directional, epistatic interactions, which reflect regulatory relationships between genes, have been challenging to map at large-scale. Here, we used combinatorial RNA interference and automated single-cell phenotyping to generate a large genetic interaction map for 21 phenotypic features of Drosophila cells. We devised a method that combines genetic interactions on multiple phenotypes to reveal directional relationships. This network reconstructed the sequence of protein activities in mitosis. Moreover, it revealed that the Ras pathway interacts with the SWI/SNF chromatin-remodelling complex, an interaction that we show is conserved in human cancer cells. Our study presents a powerful approach for reconstructing directional regulatory networks and provides a resource for the interpretation of functional consequences of genetic alterations.

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Inference of directed genetic interaction of Cdc23 and sti.Testing for directed genetic interactions for the gene pair Cdc23 and sti. Fit of the 21-dimensional π-score vector as linear function of the single gene effects. Each dot shows the π-score and single gene effect in one phenotype. The π-scores are negatively correlated to the single gene effects of sti, indicating that the knockdown phenotypic effect of sti is strongly reduced by a depletion of Cdc23.DOI:http://dx.doi.org/10.7554/eLife.05464.013
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fig4s2: Inference of directed genetic interaction of Cdc23 and sti.Testing for directed genetic interactions for the gene pair Cdc23 and sti. Fit of the 21-dimensional π-score vector as linear function of the single gene effects. Each dot shows the π-score and single gene effect in one phenotype. The π-scores are negatively correlated to the single gene effects of sti, indicating that the knockdown phenotypic effect of sti is strongly reduced by a depletion of Cdc23.DOI:http://dx.doi.org/10.7554/eLife.05464.013

Mentions: We systematically determined whether such an epistatic relationship could be identified for each of the 93,096 gene pairs by testing if the 21-dimensional genetic interaction vectors were (anti-) parallel to either of the two single knockdown vectors (‘Materials and methods’, Figure 4—figure supplement 2). Using a conservative threshold, we found directional epistatic interactions between 1344 pairs of genes. Of these, 1040 were alleviating and 304 aggravating (Supplementary file 6). This directional genetic interaction network provided a rich snapshot of regulatory, causal and temporal relationships. For instance, we detected an alleviating interaction between the APC/C member Cdc23 and the cytokinesis regulator sti. After knockdown of Cdc23, we recorded reduced growth and slightly smaller nuclear area (Figure 4B–C, blue arrows for the two independent dsRNAs). Knockdown of sti also reduced growth, but in addition led to a strong increase in nuclear area (Figure 4B–C, orange arrows). The double knockdown of Cdc23 and sti, however, almost completely reproduced the phenotype of Cdc23 alone (Figure 4C, black arrows for 2 × 2 dsRNA design). This directional alleviating interaction between the genes reflects the known, consecutive functions of the encoded proteins. Without Cdc23, cells arrest in metaphase, leading to the observed reduction in cell number (Figure 4B, top). Without sti, a component of the contractile ring, which executes the abscission of the cytoplasm to form the two daughter cells (D'Avino et al., 2004), cells undergo karyokinesis (division of the nucleus) and re-replicate their genome without being able to divide their cytoplasm, resulting in giant cells with large nuclei and often multi-nucleated cells (Figure 4B, middle). However, when Cdc23 is depleted together with sti, the increase of nucleus size is avoided (Figure 4B, bottom) since depletion of Cdc23 arrests cells in metaphase thereby preventing re-replication (Zielke et al., 2008), explaining the detected directional interaction (Figure 4D).


A map of directional genetic interactions in a metazoan cell.

Fischer B, Sandmann T, Horn T, Billmann M, Chaudhary V, Huber W, Boutros M - Elife (2015)

Inference of directed genetic interaction of Cdc23 and sti.Testing for directed genetic interactions for the gene pair Cdc23 and sti. Fit of the 21-dimensional π-score vector as linear function of the single gene effects. Each dot shows the π-score and single gene effect in one phenotype. The π-scores are negatively correlated to the single gene effects of sti, indicating that the knockdown phenotypic effect of sti is strongly reduced by a depletion of Cdc23.DOI:http://dx.doi.org/10.7554/eLife.05464.013
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4384530&req=5

fig4s2: Inference of directed genetic interaction of Cdc23 and sti.Testing for directed genetic interactions for the gene pair Cdc23 and sti. Fit of the 21-dimensional π-score vector as linear function of the single gene effects. Each dot shows the π-score and single gene effect in one phenotype. The π-scores are negatively correlated to the single gene effects of sti, indicating that the knockdown phenotypic effect of sti is strongly reduced by a depletion of Cdc23.DOI:http://dx.doi.org/10.7554/eLife.05464.013
Mentions: We systematically determined whether such an epistatic relationship could be identified for each of the 93,096 gene pairs by testing if the 21-dimensional genetic interaction vectors were (anti-) parallel to either of the two single knockdown vectors (‘Materials and methods’, Figure 4—figure supplement 2). Using a conservative threshold, we found directional epistatic interactions between 1344 pairs of genes. Of these, 1040 were alleviating and 304 aggravating (Supplementary file 6). This directional genetic interaction network provided a rich snapshot of regulatory, causal and temporal relationships. For instance, we detected an alleviating interaction between the APC/C member Cdc23 and the cytokinesis regulator sti. After knockdown of Cdc23, we recorded reduced growth and slightly smaller nuclear area (Figure 4B–C, blue arrows for the two independent dsRNAs). Knockdown of sti also reduced growth, but in addition led to a strong increase in nuclear area (Figure 4B–C, orange arrows). The double knockdown of Cdc23 and sti, however, almost completely reproduced the phenotype of Cdc23 alone (Figure 4C, black arrows for 2 × 2 dsRNA design). This directional alleviating interaction between the genes reflects the known, consecutive functions of the encoded proteins. Without Cdc23, cells arrest in metaphase, leading to the observed reduction in cell number (Figure 4B, top). Without sti, a component of the contractile ring, which executes the abscission of the cytoplasm to form the two daughter cells (D'Avino et al., 2004), cells undergo karyokinesis (division of the nucleus) and re-replicate their genome without being able to divide their cytoplasm, resulting in giant cells with large nuclei and often multi-nucleated cells (Figure 4B, middle). However, when Cdc23 is depleted together with sti, the increase of nucleus size is avoided (Figure 4B, bottom) since depletion of Cdc23 arrests cells in metaphase thereby preventing re-replication (Zielke et al., 2008), explaining the detected directional interaction (Figure 4D).

Bottom Line: Gene-gene interactions shape complex phenotypes and modify the effects of mutations during development and disease.The effects of statistical gene-gene interactions on phenotypes have been used to assign genes to functional modules.Our study presents a powerful approach for reconstructing directional regulatory networks and provides a resource for the interpretation of functional consequences of genetic alterations.

View Article: PubMed Central - PubMed

Affiliation: Genome Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
Gene-gene interactions shape complex phenotypes and modify the effects of mutations during development and disease. The effects of statistical gene-gene interactions on phenotypes have been used to assign genes to functional modules. However, directional, epistatic interactions, which reflect regulatory relationships between genes, have been challenging to map at large-scale. Here, we used combinatorial RNA interference and automated single-cell phenotyping to generate a large genetic interaction map for 21 phenotypic features of Drosophila cells. We devised a method that combines genetic interactions on multiple phenotypes to reveal directional relationships. This network reconstructed the sequence of protein activities in mitosis. Moreover, it revealed that the Ras pathway interacts with the SWI/SNF chromatin-remodelling complex, an interaction that we show is conserved in human cancer cells. Our study presents a powerful approach for reconstructing directional regulatory networks and provides a resource for the interpretation of functional consequences of genetic alterations.

Show MeSH
Related in: MedlinePlus