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Cilia Dynamics During Embryonic Development

Reference : PhD Julien VERMOT

Offer publication : April 6, 2016

Our hypothesis is that mechanodetection is involved in setting left-right axis in zebrafish and that cilia act as the flow sensor dictating left-right axis in the KV.

To test this hypothesis, the project is divided in two main objectives as follows:

1. To determine the flow forces in the KV and their impact on asymmetric gene expression through an integrated approach combining biological and optical approaches.

2. To identify the gene target of the mechanodetection pathway in the KV involved in left-right axis specification.
The project is fragmented into five complementary tasks:
-Task 1 corresponds to the coordination/administration work.
-For Task 2, flow forces will first be analyzed using live, high-resolution imaging permitting to uncover the flow generated in the KV. This approach have been already used and validated. It allows both flow and cilia position to be monitored in 3D. Next we will ablate cilia function using chromophore assisted laser inactivation (CALI), in live embryo to test the impact of cilia motility on flow, calcium signaling and left-right patterning. In addition, we will analyze the relative impact of ectopic calcium fluxes in the KV using optogenetics during left-right specification. The technical development necessary to achieve this task will then be used in tasks 3 and 4.

-Task 3 will analyze the role of KV cilia in the mechanodetection process. First, we will test the properties of cilia to mechanically sense physical stimuli using optical tweezing and cilia ablation in vivo. We will then knock out PKD2, a non-selective calcium channel identified as the mechanodetector involved in left-right patterning, specifically within cilia using the deGradeFP approach. This will permit to show if the localization of PKD2 within cilia is necessary for the mechanodetection process.

-Task 4 will analyze the impact of PKD2 on asymmetric gene expression within the KV using a highly stringent gene profiling approach. We will identify the gene abnormally expressed in the absence of PKD2 function. This will uncover the ability of PKD2 to control gene expression and identify the gene
targets downstream of the mechanodection pathway.
These data should lead to a better understanding of the general processes controlling gene expression in response to mechanical forces, which is
essential during development and also in pathologies such as ciliopathies.

- WISHED SKILLS : Molecular biologie, cell biology


Your application

Application Deadline : Dec. 31, 2016

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