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Evidence that the C-terminal domain (CtD) autoinhibits neural repression by Drosophila E(spl)M8
Bhaskar Kahali1, Jee-Eun Kim, Umesh Karandikar
1Department of Biology, West Virginia University, Morgantown, WV, USA.
Abstract:
Analysis of the retinal defects of a CK2 phosphomimetic variant of E(spl)M8 (M8S(159)D) and the truncated protein M8* encoded by the E(spl)D allele, suggest that the nonphosphorylated CtD "autoinhibits" repression. We have investigated this model by testing for inhibition (in "trans") by the CtD fragment in its nonphosphorylated (M8-CtD) and phosphomimetic (M8SD-CtD) states. In N(+) flies, ectopic M8-CtD compromises lateral inhibition, i.e., elicits supernumerary bristles as with loss of N signaling. This antimorphic activity of M8-CtD strongly rescues the reduced eye and/or bristle loss phenotypes that are elicited by ectopic M8SD or wild type M8. Additionally, the severely reduced eye of N(spl)/Y; E(spl)D/+ flies is also rescued by M8-CtD. Rescue is specific to the time and place, the morphogenetic furrow, where "founding" R8 photoreceptors are specified. In contrast, the phosphomimetic M8SD-CtD that is predicted to be deficient for autoinhibition, exhibits significantly attenuated or negligible activity. These studies provide evidence that autoinhibition by the CtD regulates M8 activity in a phosphorylation-dependent manner.
Insights
The C-terminal domain (CtD) of E(spl)M8 protein autoinhibits its function. Phosphorylation disrupts this autoinhibition, impacting eye development and bristle formation in flies.
Area of Science:
- Developmental Biology
- Genetics
- Molecular Biology
Background:
- The E(spl)M8 protein plays a crucial role in regulating cell fate during development.
- Previous studies suggested that the C-terminal domain (CtD) of E(spl)M8 might regulate its activity through autoinhibition.
- Phosphorylation of the CtD is hypothesized to affect this autoinhibitory mechanism.
Purpose of the Study:
- To investigate the role of the nonphosphorylated and phosphomimetic CtD fragments of E(spl)M8 in regulating its function.
- To test the hypothesis that the nonphosphorylated CtD autoinhibits repression by E(spl)M8.
- To determine the phosphorylation-dependent regulation of E(spl)M8 activity.
Main Methods:
- Analysis of retinal defects in Drosophila melanogaster (flies) expressing specific E(spl)M8 variants.
- Testing the 'in trans' inhibitory activity of nonphosphorylated (M8-CtD) and phosphomimetic (M8SD-CtD) CtD fragments.
- Evaluating the effects of these fragments on lateral inhibition and photoreceptor specification.
Main Results:
- Ectopic expression of nonphosphorylated M8-CtD in N(+) flies compromised lateral inhibition, leading to supernumerary bristles.
- M8-CtD exhibited antimorphic activity, rescuing eye and bristle loss phenotypes caused by ectopic M8SD or wild-type M8.
- The phosphomimetic M8SD-CtD showed significantly attenuated or negligible activity, suggesting a loss of autoinhibition.
Conclusions:
- The C-terminal domain (CtD) of E(spl)M8 functions as an autoinhibitory domain.
- Phosphorylation of the CtD disrupts this autoinhibition, altering E(spl)M8's regulatory function.
- Autoinhibition by the CtD regulates E(spl)M8 activity in a phosphorylation-dependent manner, crucial for proper development.
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