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Published on: November 6, 2017
Patched-Related Is Required for Proper Development of Embryonic Drosophila Nervous System
Carmen Bolatto1,2, Sofía Nieves1, Agustina Reyes1
1Developmental Biology Laboratory, Histology and Embryology Department, Faculty of Medicine, Universidad de la República (UdelaR), Montevideo, Uruguay.
Insights
Patched-related (Ptr) protein is crucial for nervous system development. This study reveals Ptr acts as a negative regulator of the Hedgehog (Hh) signaling pathway, interacting directly with Hh.
Area of Science:
- Developmental biology
- Neuroscience
- Molecular genetics
Background:
- Patched-related (Ptr) protein shares similarities with Patched (Ptc), a key receptor in the Hedgehog (Hh) pathway.
- The precise role of Ptr in nervous system development is not fully understood.
Purpose of the Study:
- To investigate the physiological function of Ptr in the developing nervous system.
- To determine Ptr's role in the Hedgehog signaling pathway.
Main Methods:
- Generation and analysis of Ptr null mutant embryos using confocal microscopy.
- Immunolabeling to assess axonal tracts and glial cell distribution.
- Cell-based reporter assays and co-immunoprecipitation to study Hh pathway interactions.
- In vivo overexpression studies in imaginal wing discs.
Main Results:
- Ptr null mutant embryos exhibit severe defects in axonal tracts, glial cell populations, and peripheral nervous system morphology.
- Downregulation of Ptr in nerve cells recapitulates these nervous system phenotypes.
- Ptr acts as a negative regulator of Hh signaling, demonstrated by reporter assays.
- Direct interaction between Ptr and Hh was confirmed via co-immunoprecipitation.
- In vivo Ptr overexpression induces wing phenotypes at the anterior/posterior border.
Conclusions:
- Ptr plays a critical role in the development of the nervous system.
- Ptr functions as a negative regulator of the Hedgehog signaling pathway.
- Ptr directly interacts with Hh, influencing its signaling activity.
Abstract:
Patched-related (Ptr), classified primarily as a neuroectodermal gene, encodes a protein with predicted topology and domain organization closely related to those of Patched (Ptc), the canonical receptor of the Hedgehog (Hh) pathway. To investigate the physiological function of Ptr in the developing nervous system, Ptr null mutant embryos were immunolabeled and imaged under confocal microscopy. These embryos displayed severe alterations in the morphology of the primary axonal tracts, reduced number, and altered distribution of the Repo-positive glia as well as peripheral nervous system defects. Most of these alterations were recapitulated by downregulating Ptr expression, specifically in embryonic nerve cells. Because similar nervous system phenotypes have been observed in hh and ptc mutant embryos, we evaluated the Ptr participation in the Hh pathway by performing cell-based reporter assays. Clone-8 cells were transfected with Ptr-specific dsRNA or a Ptr DNA construct and assayed for changes in Hh-mediated induction of a luciferase reporter. The results obtained suggest that Ptr could act as a negative regulator of Hh signaling. Furthermore, co-immunoprecipitation assays from cell culture extracts premixed with a conditioned medium revealed a direct interaction between Ptr and Hh. Moreover, in vivo Ptr overexpression in the domain of the imaginal wing disc where Engrailed and Ptc coexist produced wing phenotypes at the A/P border. Thus, these results strongly suggest that Ptr plays a crucial role in nervous system development and appears to be a negative regulator of the Hh pathway.
