Pkd2 affects the architecture of zebrafish left-right organizer

Background

Dorsal anterior clustering (DAC) of motile cilia in the left-right organizer (LRO) is crucial for normal fluid flow dynamics and correct laterality in zebrafish [1]. We directly demonstrated that charon/dand5 transcription is negatively regulated by strong flow in zebrafish LRO [1] which suggests that LRO cells have the ability to sense fluid flow and influence gene expression patterns, but how? Pkd2 ion channel is a good candidate because it participates in a mechanosensory complex that senses fluid flow and induces a calcium inward flux in kidney cells [2] and in nodal cells [3]. In agreement, mouse and zebrafish mutants for Pkd2 have LR defects [4, 5]. However, Pkd2 is also involved in cell polarity during migration [6] and in extracellular matrix deposition [7] implying a role for Pkd2 in cell morphogenesis.

Objective

Determine whether Pkd2 knockdown affects DAC of ciliated LRO cells.

Methods

dnah7 morpholino [1] was injected to generate static cilia without affecting DAC [1]. Each embryo was screened for static cilia by high-speed videomicroscopy. In parallel, Pkd2 knockdowns were imaged for flow dynamics followed by quantification of anterior / posterior cilia number through two-photon microscopy.

Results

Pkd2 knockdown, contrary to Dnah7 knockdown, caused a remodelling in the LRO architecture by disrupting DAC. Moreover, Pkd2 knockdown resulted in abnormal fluid flow as a consequence of defective DAC.

Conclusions

Comparing Dnah7 and Pkd2 knockdowns we concluded that Pkd2 mediated pathway affects LRO morphogenesis by a mechanism that seems to be independent of its role in fluid flow mechanosensation. Meaning that Pkd2 triggers additional responses from those caused by LRO flow.

Supported by FCT-ANR/BEX-BID/0153/2012 grant.