Cilia at the Node of Mouse Embryos Sense Fluid Flow for Left-Right Determination via Pkd2 (original) (raw)
Related papers
Biophysical Journal, 2005
Nodal cilia dynamics is a key factor for left/right axis determination in mouse embryos through the induction of a leftward fluid flow. So far it has not been clearly established how such dynamics is able to induce the asymmetric leftward flow within the node. Herein we propose that an asymmetric two-phase nonplanar beating cilia dynamics that involves the bending of the ciliar axoneme is responsible for the leftward fluid flow. We support our proposal with a host of hydrodynamic arguments, in silico experiments and in vivo video microscopy data in wild-type embryos and inv mutants. Our phenomenological modeling approach underscores how the asymmetry and speed of the flow depends on different relevant parameters. In addition, we discuss how the combination of internal and external mechanisms might cause the two-phase beating cilia dynamics.
During mammalian development, left-right (L-R) asymmetry is established by a cilia-driven left-ward fluid flow within a midline embryonic cavity called the node. This 'nodal flow' is detected by peripherally-located crown cells that each assemble a primary cilium which contain the putative Ca 2+ channel PKD2. The interaction of flow and crown cell cilia promotes left side-specific expression of Nodal in the lateral plate mesoderm (LPM). Whilst the PKD2-interacting protein PKD1L1 has also been implicated in L-R patterning, the underlying mechanism by which flow is detected and the genetic relationship between Polycystin function and asymmet-ric gene expression remains unknown. Here, we characterize a Pkd1l1 mutant line in which Nodal is activated bilaterally, suggesting that PKD1L1 is not required for LPM Nodal pathway activation per se, but rather to restrict Nodal to the left side downstream of nodal flow. Epistasis analysis shows that Pkd1l1 acts as an upstream genetic repressor of Pkd2. This study therefore provides a genetic pathway for the early stages of L-R determination. Moreover, using a system in which cultured cells are supplied artificial flow, we demonstrate that PKD1L1 is sufficient to mediate a Ca 2+ signaling response after flow stimulation. Finally, we show that an extracellular PKD domain within PKD1L1 is crucial for PKD1L1 function; as such, destabilizing the domain causes L-R defects in the mouse. Our demonstration that PKD1L1 protein can mediate a response to flow coheres with a mechanosensation model of flow sensation in which the force of fluid flow drives asymmetric gene expression in the embryo.
Cilia-Driven Leftward Flow Determines Laterality in Xenopus
Current Biology, 2007
Determination of the vertebrate left-right body axis during embryogenesis results in asymmetric development and placement of most inner organs . Although the asymmetric Nodal cascade is conserved in all vertebrates, the mechanism of symmetry breakage has remained controversial . In mammalian and fish embryos, a cilia-driven leftward flow of extracellular fluid is required for initiation of the Nodal cascade. This flow is localized at the posterior notochord (''node'') and Kupffer's vesicle, respectively . In frog and chick embryos, however, molecular asymmetries are required earlier, from cleavage stages through gastrulation . The validity of a cilia-based mechanism for all vertebrates therefore has been questioned [3]. Here we show that a cilia-driven leftward flow precedes asymmetric nodal expression in the frog Xenopus. Motile monocilia emerged on the gastrocoel roof plate during neurulation and lengthened and polarized from an initially central position to the posterior pole of cells. Concomitantly, a robust leftward fluid flow developed from stage 15 onward, significantly before asymmetric nodal transcription started in the leftlateral-plate mesoderm at stage 19. Injection of 1.5% methylcellulose into the archenteron prevented leftward flow and resulted in laterality defects, demonstrating that the flow itself was required for asymmetric gene expression and organ placement.
Left-right determination: involvement of molecular motor KIF3, cilia, and nodal flow
Cold Spring Harbor perspectives in biology, 2009
Mammalian left-right determination is a good example for how multiple cell biological processes coordinate in the formation of a basic body plan. The leftward movement of fluid at the ventral node, called nodal flow, is the central process in symmetry breaking on the left-right axis. Nodal flow is autonomously generated by the rotation of posteriorly tilted cilia that are built by transport via KIF3 motor on cells of the ventral node. How nodal flow is interpreted to create left-right asymmetry has been a matter of debate. Recent evidence suggests that the leftward movement of sheathed lipidic particles, called nodal vesicular parcels (NVPs), may result in the activation of the noncanonical hedgehog signaling pathway, an asymmetric elevation in intracellular Ca(2+) and changes in gene expression.
Cell, 1998
formed by KIF3, we generated KIF3B knockout mice. The kif3B Ϫ/Ϫ mice did not survive beyond midgestation, University of Tokyo 7-3-1 Hongo, Tokyo, 113-0033 and the embryos exhibited apparent morphological abnormalities including randomized left-right (L-R) asym-Japan metry, pericardial sac ballooning, growth retardation, neural tube closure defect, and neural tube staggering. Among these phenotypes, we focused on the random-Summary ization of L-R asymmetry. The L-R asymmetry first becomes anatomically ap-Microtubule-dependent motor, murine KIF3B, was disparent in the orientation of the heart tube looping (Kaufrupted by gene targeting. The null mutants did not man, 1992), but it is already detectable at the somitogensurvive beyond midgestation, exhibiting growth retaresis stage by asymmetric expression of several genes, dation, pericardial sac ballooning, and neural tube dissuch as lefty-1, lefty-2, nodal, and Pitx2, with expression organization. Prominently, the left-right asymmetry being observed in the left side in most cases (reviewed was randomized in the heart loop and the direction in Harvey, 1998). Ex utero culture experiments of rat of embryonic turning. lefty-2 expression was either embryos showed that the determination of L-R asymmebilateral or absent. Furthermore, the node lacked try occurs during the early neural plate stage (Brown et monocilia while the basal bodies were present. Immu-
Differentiation, 2005
The invariant asymmetric placement of thoracic and abdominal organs in the vertebrates is controlled by the left-asymmetric activity of the Nodal signaling cascade during embryogenesis. In the mouse embryo asymmetric induction of nodal is thought to be dependent on functional monocilia on the ventral node cells and on the Pkd2 gene, which encodes the calcium channel polycystin-2 (PC2). In humans mutations in PKD2 and PKD1 give rise to polycystic kidney disease. The PC1 and PC2 proteins are thought to function as part of a multifactorial complex. Localization of both proteins to the primary renal cilium suggested a function on cilia of the ventral node. Here we investigated Pkd1 knock-out embryos for laterality defects and found wild-type organ morphogenesis and normal expression of nodal and Pitx2. While PC2 localized to nodal cilia, no ciliary localization of PC1 was detected in mouse embryos. This finding was confirmed in an archetypical mammalian blastodisc, the rabbit embryo. Thus, absence of PC1 localization to cilia corresponded with a lack of laterality defects in Pkd1 knock-out embryos. Our results demonstrate a PC1-independent function of PC2 in left-right axis formation, and indirectly support a ciliary role of PC2 in this process.
Mechanism of Nodal Flow: A Conserved Symmetry Breaking Event in Left-Right Axis Determination
Cell, 2005
The leftward flow in extraembryonic fluid is critical for the initial determination of the left-right axis of mouse embryos. It is unclear if this is a conserved mechanism among other vertebrates and how the directionality of the flow arises from the motion of cilia. In this paper, we show that rabbit and medakafish embryos also exhibit a leftward fluid flow in their ventral nodes. In all cases, primary monocilia present a clockwise rotational-like motion. Observations of defective ciliary dynamics in mutant mouse embryos support the idea that the posterior tilt of the cilia during rotationallike beating can explain the leftward fluid flow. Moreover, we show that this leftward flow may produce asymmetric distribution of exogenously introduced proteins, suggesting morphogen gradients as a subsequent mechanism of left-right axis determination. Finally, we experimentally and theoretically characterize under which conditions a morphogen gradient can arise from the flow.
Pkd1l1 complexes with Pkd2 on motile cilia and functions to establish the left-right axis
Development, 2011
The internal organs of vertebrates show distinctive left-right asymmetry. Leftward extracellular fluid flow at the node (nodal flow), which is generated by the rotational movement of node cilia, is essential for left-right patterning in the mouse and other vertebrates. However, the identity of the pathways by which nodal flow is interpreted remains controversial as the molecular sensors of this process are unknown. In the current study, we show that the medaka left-right mutant abecobe (abc) is defective for left-right asymmetric expression of southpaw, lefty and charon, but not for nodal flow. We identify the abc gene as pkd1l1, the expression of which is confined to Kupffer's vesicle (KV, an organ equivalent to the node). Pkd1l1 can interact and interdependently colocalize with Pkd2 at the cilia in KV. We further demonstrate that all KV cilia contain Pkd1l1 and Pkd2 and left-right dynein, and that they are motile. These results suggest that Pkd1l1 and Pkd2 form a complex that ...