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Transvection-based gene regulation in Drosophila is a complex and plastic trait.

Bing X, Rzezniczak TZ, Bateman JR, Merritt TJ - G3 (Bethesda) (2014)

Bottom Line: We further show that the magnitude of transvection at the Men locus is modified by both genetic background and environment (temperature), demonstrating that transvection is a plastic phenotype.Our results suggest that transvection effects in D. melanogaster are shaped by a dynamic interplay between environment and genetic background.Interestingly, we find that cis-based regulation of the Men gene is more robust to genetic background and environment than trans-based.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, Laurentian University, Sudbury, ON, P3E 2C6, Canada.

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Effect of RNAi knockdown of transcription factors on Men in cis. (A) Relative gene expression in all three RNAi experiments showing significant knockdown of expression for all three genes analyzed (asterisk indicates significant reduction in gene expression, P < 0.05). (B) Men gene expression in Abd-B, mirr, and slbo knocked down lines, relative to control lines. (C) MEN enzyme activity in the same TF knocked down lines, relative to control lines. There are significant differences in MEN activity between columns with different letters (F3,24= 10.0164, P < 0.0001, Tukey’s honestly significant difference P < 0.05). In all panels, mean expression/activity of the control lines is indicated by the dotted line with the flanking faint lines indicating standard error.
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fig8: Effect of RNAi knockdown of transcription factors on Men in cis. (A) Relative gene expression in all three RNAi experiments showing significant knockdown of expression for all three genes analyzed (asterisk indicates significant reduction in gene expression, P < 0.05). (B) Men gene expression in Abd-B, mirr, and slbo knocked down lines, relative to control lines. (C) MEN enzyme activity in the same TF knocked down lines, relative to control lines. There are significant differences in MEN activity between columns with different letters (F3,24= 10.0164, P < 0.0001, Tukey’s honestly significant difference P < 0.05). In all panels, mean expression/activity of the control lines is indicated by the dotted line with the flanking faint lines indicating standard error.

Mentions: The predicted TFBSs described previously are present in multiple copies within highly conserved regions across the Men locus (Figure 5A; Lum and Merritt 2011) and we suspected that TFs bound to these sites could regulate Men expression in cis, in addition to any role in trans-regulation. Using RNAi, we reduced the expression of three TF genes with binding sites in this region, Abd-B, mirr, and slbo, and assayed for differences in Men expression and MEN activity (Figure 8A). Abd-B was selected because of its correlation with transvection at Men described previously, whereas mirr and slbo were selected as a contrast because no consistent correlations were found with these genes in trans. Following reports of larger RNAi effects with multiple heat shocks (Kristensen et al. 2003), vials were heat shocked five times to induce expression of hairpins: 1, 2, 4, 8, and 12 days after egg laying, resulting in reproducible reduction in TF expression (Figure 8). Note that all heat shock treatments were administered postembryogenesis to avoid the primary period during which Abd-B functions in embryonic patterning (Graveley et al. 2011, Akam 1987), and that no gross morphological changes were observed in any knockdown flies. Notably, Men expression and MEN activity were both significantly reduced only in flies that had Abd-B expression knocked down (Figure 8, B and C and Table S3). The reduction in malic enzyme with only a 50% reduction in Abd-B may be surprising at first, but previous studies have found malic enzyme to be sensitive to small differences in activity of other genes or the environment (e.g., Merritt et al. 2005; Geer et al. 1976). These results, and the correlation of Abd-B and Men expression seen in the previous experiment, support a model in which Abd-B plays a role in the regulation of Men expression both in cis and in trans.


Transvection-based gene regulation in Drosophila is a complex and plastic trait.

Bing X, Rzezniczak TZ, Bateman JR, Merritt TJ - G3 (Bethesda) (2014)

Effect of RNAi knockdown of transcription factors on Men in cis. (A) Relative gene expression in all three RNAi experiments showing significant knockdown of expression for all three genes analyzed (asterisk indicates significant reduction in gene expression, P < 0.05). (B) Men gene expression in Abd-B, mirr, and slbo knocked down lines, relative to control lines. (C) MEN enzyme activity in the same TF knocked down lines, relative to control lines. There are significant differences in MEN activity between columns with different letters (F3,24= 10.0164, P < 0.0001, Tukey’s honestly significant difference P < 0.05). In all panels, mean expression/activity of the control lines is indicated by the dotted line with the flanking faint lines indicating standard error.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4232543&req=5

fig8: Effect of RNAi knockdown of transcription factors on Men in cis. (A) Relative gene expression in all three RNAi experiments showing significant knockdown of expression for all three genes analyzed (asterisk indicates significant reduction in gene expression, P < 0.05). (B) Men gene expression in Abd-B, mirr, and slbo knocked down lines, relative to control lines. (C) MEN enzyme activity in the same TF knocked down lines, relative to control lines. There are significant differences in MEN activity between columns with different letters (F3,24= 10.0164, P < 0.0001, Tukey’s honestly significant difference P < 0.05). In all panels, mean expression/activity of the control lines is indicated by the dotted line with the flanking faint lines indicating standard error.
Mentions: The predicted TFBSs described previously are present in multiple copies within highly conserved regions across the Men locus (Figure 5A; Lum and Merritt 2011) and we suspected that TFs bound to these sites could regulate Men expression in cis, in addition to any role in trans-regulation. Using RNAi, we reduced the expression of three TF genes with binding sites in this region, Abd-B, mirr, and slbo, and assayed for differences in Men expression and MEN activity (Figure 8A). Abd-B was selected because of its correlation with transvection at Men described previously, whereas mirr and slbo were selected as a contrast because no consistent correlations were found with these genes in trans. Following reports of larger RNAi effects with multiple heat shocks (Kristensen et al. 2003), vials were heat shocked five times to induce expression of hairpins: 1, 2, 4, 8, and 12 days after egg laying, resulting in reproducible reduction in TF expression (Figure 8). Note that all heat shock treatments were administered postembryogenesis to avoid the primary period during which Abd-B functions in embryonic patterning (Graveley et al. 2011, Akam 1987), and that no gross morphological changes were observed in any knockdown flies. Notably, Men expression and MEN activity were both significantly reduced only in flies that had Abd-B expression knocked down (Figure 8, B and C and Table S3). The reduction in malic enzyme with only a 50% reduction in Abd-B may be surprising at first, but previous studies have found malic enzyme to be sensitive to small differences in activity of other genes or the environment (e.g., Merritt et al. 2005; Geer et al. 1976). These results, and the correlation of Abd-B and Men expression seen in the previous experiment, support a model in which Abd-B plays a role in the regulation of Men expression both in cis and in trans.

Bottom Line: We further show that the magnitude of transvection at the Men locus is modified by both genetic background and environment (temperature), demonstrating that transvection is a plastic phenotype.Our results suggest that transvection effects in D. melanogaster are shaped by a dynamic interplay between environment and genetic background.Interestingly, we find that cis-based regulation of the Men gene is more robust to genetic background and environment than trans-based.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, Laurentian University, Sudbury, ON, P3E 2C6, Canada.

Show MeSH
Related in: MedlinePlus