Journal
DEVELOPMENTAL BIOLOGY
Volume 372, Issue 1, Pages 17-27Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ydbio.2012.09.008
Keywords
Drosophila; Mesoderm migration; Rho GEF; Phosphoinositide; Pebble; Pleckstrin homology domain
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Funding
- RESTRACOMP scholarship from The Hospital for Sick Children
- The Cancer Research Society, Inc.
- ARC Special Research Centre for the Molecular Genetics of Development
- NHMRC project grant [316959]
- ARC Discovery Grant [DP0987338]
- NIH New Innovator Award [DP2OD004265, R01CA138676]
- American Cancer Society
- Australian Research Council [DP0987338] Funding Source: Australian Research Council
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The Drosophila RhoGEF Pebble (Pbl) is required for cytokinesis and migration of mesodermal cells. In a screen for genes that could suppress migration defects in pbl mutants we identified the phosphatidylinositol phosphate (PtdInsP) regulator pi5k59B. Genetic interaction tests with other PtdInsP regulators suggested that PtdIns(4,5)P2 levels are important for mesoderm migration when PIA is depleted. Consistent with this, the leading front of migrating mesodermal cells was enriched for PtdIns(4,5)P2. Given that Pbl contains a Pleckstrin Homology (PH) domain, a known PtdInsP-binding motif, we examined PtdInsP-binding of Pbl and the importance of the PH domain for PIA function. In vitro lipid blot assays showed that Pbl binds promiscuously to PtdInsPs, with binding strength associated with the degree of phosphorylation. PIA was also able to bind lipid vesicles containing PtdIns(4,5)P2 but binding was strongly reduced upon deletion of the PH domain. Similarly, in vivo, loss of the PH domain prevented localisation of Pbl to the cell cortex and severely affected several aspects of early mesoderm development, including flattening of the invaginated tube onto the ectoderm, extension of protrusions, and dorsal migration to form a monolayer. Pbl lacking the PH domain could still localise to the cytokinetic furrow, however, and cytokinesis failure was reduced in pbl(Delta PH) mutants. Taken together, our results support a model in which interaction of the PH-domain of Pbl with PtdIns(4,5)P2 helps localise it to the plasma membrane which is important for mesoderm migration. (c) 2012 Elsevier Inc. All rights reserved.
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