Abstract
Background
Ligand endocytosis plays a critical role in regulating the activity of the Notch pathway. TheDrosophila homolog of auxilin (dAux), a J-domain-containing protein best known for its role in the disassembly of clathrin coats from clathrin-coated vesicles, has recently been implicated in Notch signaling, although its exact mechanism remains poorly understood.
Results
To understand the role of auxilin in Notch ligand endocytosis, we have analyzed several point mutations affecting specific domains of dAux. In agreement with previous work, analysis using these stronger dAux alleles shows that dAux is required for several Notch-dependent processes, and its function during Notch signaling is required in the signaling cells. In support of the genetic evidences, the level of Delta appears elevated in dAux deficient cells, suggesting that the endocytosis of Notch ligand is disrupted. Deletion analysis shows that the clathrin-binding motif and the J-domain, when over-expressed, are sufficient for rescuing dAux phenotypes, implying that the recruitment of Hsc70 to clathrin is a critical role for dAux. However, surface labeling experiment shows that, in dAux mutant cells, Delta accumulates at the cell surface. In dAux mutant cells, clathrin appears to form large aggregates, although Delta is not enriched in these aberrant clathrin-positive structures.
Conclusion
Our data suggest that dAux mutations inhibit Notch ligand internalization at an early step during clathrin-mediated endocytosis, before the disassembly of clathrin-coated vesicles. Further, the inhibition of ligand endocytosis in dAux mutant cells possibly occurs due to depletion of cytosolic pools of clathrin via the formation of clathrin aggregates. Together, our observations argue that ligand endocytosis is critical for Notch signaling and auxilin participates in Notch signaling by facilitating ligand internalization.
Date of this Version
5-8-2008
DOI
10.1186/1471-213X-8-50
Recommended Citation
Kandachar, Vasundhara; Bai, Ting; and Chang, Henry C., "The Clathrin-Binding Motif and the J-Domain of Drosophila Auxilin are Essential for Facilitating Notch Ligand Endocytosis." (2008). Department of Biological Sciences Faculty Publications. Paper 68.
http://dx.doi.org/10.1186/1471-213X-8-50
Comments
This is the publisher pdf of Vasundhara Kandachar, Ting Bai, Henry C Chang. The Clathrin-Binding Motif and the J-Domain of Drosophila Auxilin are Essential for Facilitating Notch Ligand Endocytosis. BMC Developmental Biology 2008, 8:50 (8 May 2008) and is available at: 10.1186/1471-213X-8-50.