Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
FGF signalling during embryo development regulates cilia length in diverse epithelia.
Neugebauer JM
,
Amack JD
,
Peterson AG
,
Bisgrove BW
,
Yost HJ
.
???displayArticle.abstract???
Cilia are cell surface organelles found on most epithelia in vertebrates. Specialized groups of cilia have critical roles in embryonic development, including left-right axis formation. Recently, cilia have been implicated as recipients of cell-cell signalling. However, little is known about cell-cell signalling pathways that control the length of cilia. Here we provide several lines of evidence showing that fibroblast growth factor (FGF) signalling regulates cilia length and function in diverse epithelia during zebrafish and Xenopus development. Morpholino knockdown of FGF receptor 1 (Fgfr1) in zebrafish cell-autonomously reduces cilia length in Kupffer's vesicle and perturbs directional fluid flow required for left-right patterning of the embryo. Expression of a dominant-negative FGF receptor (DN-Fgfr1), treatment with SU5402 (a pharmacological inhibitor of FGF signalling) or genetic and morpholino reduction of redundant FGF ligands Fgf8 and Fgf24 reproduces this cilia length phenotype. Knockdown of Fgfr1 also results in shorter tethering cilia in the otic vesicle and shorter motile cilia in the pronephric ducts. In Xenopus, expression of a dn-fgfr1 results in shorter monocilia in the gastrocoel roof plate that control left-right patterning and in shorter multicilia in external mucociliary epithelium. Together, these results indicate a fundamental and highly conserved role for FGF signalling in the regulation of cilia length in multiple tissues. Abrogation of Fgfr1 signalling downregulates expression of two ciliogenic transcription factors, foxj1 and rfx2, and of the intraflagellar transport gene ift88 (also known as polaris), indicating that FGF signalling mediates cilia length through an Fgf8/Fgf24-Fgfr1-intraflagellar transport pathway. We propose that a subset of developmental defects and diseases ascribed to FGF signalling are due in part to loss of cilia function.
Figure 2. FGF signaling controls cilia length and directional fluid flow in Kupfferâs Vesicle(a-b) Confocal images of 10 SS embryos, KV labeled with antibodies against aPKC (red) and acetylated tubulin (green). Control and fgfr1 morphants had similar KV structure, but cilia were shorter in fgfr1 morphants (compare insets in a and b). (c) Cilia lengths were significantly different (p< 2.88e-06) in fgfr1 morphants (688 cilia; 18 embryos) versus Control morphants (437 cilia; 9 embryos). Cilia length was similar in WT uninjected (533 cilia; 10 embryos) and Control morphant (p< 0.93), cilia numbers per KV were similar in Control and fgfr1 morphants (p< 0.26). Cilia length defects in fgfr1 morphants were rescued by Xenopus FGFR1 (xFGFR1) mRNA (p< 4.70e-05; 807 cilia; 21 embryos). Injection of xFGFR1 mRNA alone had no affect on cilia length (p<0.73; 526 cilia, 14 embryos). (d) Embryos treated with SU5402 during shield stage (248 cilia; 12 embryos) had shorter cilia compared to DMSO control embryos (p<3.26e-06; 686 cilia; 15 embryos). (e) Cilia were shorter in transgenic DN-FGFR embryos that were heat shocked at 60% epiboly (656 cilia; 19 embryos) compared to heat shocked non-transgenic siblings (p<6.94e-03; 375 cilia; 10 embryos) and non-heat-shocked siblings (p<6.99e-03; 910 cilia; 16 embryos). (f) There was no difference in cilia length (p<0.28) in fgf24 morphants (455 cilia; 10 embryos) versus Control morphants (481 cilia; 10 embryos). However, cilia were shorter when both FGF8 and FGF24 ligands were diminished (fgf24 MO in ace mutants; 12 embryos; 244 cilia), compared to single ligand knockdown (FGF8/ace mutants: p< 1.39e-04; 10 embryos; 480 cilia; fgf24 MO in ace sibs: p< 3.44e-04; 15 embryos; 643 cilia) and WT ace siblings (p<3.63e-07; 13 embryos; 626 cilia). (g-h) DIC images of bead-injected KVs in Control and fgfr1 morphants injected with fluorescent beads. (i-j) Bead paths tracked by Metamorph software. Directional KV fluid flow was absent in fgfr1 morphants (j; p< 6.4e-15; 44 beads, 9 embryos) compared to counterclockwise flow in Control morphants (i; 39 beads, 8 embryos). Error bars are standard error of the mean (s.e.m).
Figure 3. Cilia length in pronephric ducts, otic vesicles, gastrocoel roof plate epithelia and mucociliary epithelia is controlled by FGF signaling(a-b) Pronephric duct cilia were shorter and disorganized in fgfr1 morphants (p<4.24Ã10-4; 528 cilia; 10 embryos,) compared to WT (517 cilia; 10 embryos) 26 SS embryos. (c-d) Otic vesicle tethering cilia (arrows, and inset) were shorter (p< 1.10e-07) in fgfr1 morphants (325 cilia; 10 embryos) compared to WT embryos (322 cilia; 8 embryos) at 24 hpf. (g-j, m). GRP cilia in Xenopus embryos were normal length in cells expressing GFP alone (green cells in g, outlined in h; 316 cilia 18 embryos, p<0.11), neighboring cells (outside boundaries in h; 653, 18 embryos), and cells neighboring DN-FGFR+GFP expression (outside boundaries in j; 652 cilia 15 embryos, p<0.99). In contrast, GRP cilia were shorter in cells expressing DN-FGFR+GFP (i, inside boundaries in j; 155 cilia, 15 embryos) compared to neighboring cells (p<6.1e-03) and cells expressing GFP alone (p<2.7e-03). (k, l) Z plane rendering of mucociliary epithelia (scale bar 20 um), showing shorter cilia in cells expressing DN-FGFR+GFP (13 cells, 7 embryos) compared to controls expressing GFP alone (14 cells, 4 embryos). (n) Multicilia area is reduced in cells expressing DN-FGFR+GFP (p<0.019). Error bars are s.e.m.
Figure 4. FGF signaling controls ciliogenic genes in DFC/KV cells(a, b) sox17 expression in DFC/KV (and endoderm cells in a different focal plane) in 90% epiboly embryos was normal in fgfr1 morphants and WT embryos. (c, d) Expression of dnah9 in 95 % epiboly embryos was normal in fgfr1 morphants and WT embryos. (e, f) In contrast, foxJ1 was down-regulated in fgfr1 morphants versus WT embryos at 90% epiboly. (g, h) Similarly, polaris was down-regulated in fgfr1 morphants versus WT embryos at tailbud stage. (i) Comparison of percentage of embryos with WT expression levels of each gene indicated. (j) Proposed mechanism by which FGF signaling controls length of motile cilia: FGF ligands bind to FGFR1 activating downstream transcription factors (TF) including foxj1 and rfx2, these TF activate IFT genes (e.g. polaris) to maintain motile cilia length on epithelial cells.
Albertson,
Roles for fgf8 signaling in left-right patterning of the visceral organs and craniofacial skeleton.
2005, Pubmed
Albertson,
Roles for fgf8 signaling in left-right patterning of the visceral organs and craniofacial skeleton.
2005,
Pubmed
Amack,
Two T-box genes play independent and cooperative roles to regulate morphogenesis of ciliated Kupffer's vesicle in zebrafish.
2007,
Pubmed
,
Xenbase
Amack,
The T box transcription factor no tail in ciliated cells controls zebrafish left-right asymmetry.
2004,
Pubmed
Amaya,
Expression of a dominant negative mutant of the FGF receptor disrupts mesoderm formation in Xenopus embryos.
1991,
Pubmed
,
Xenbase
Bisgrove,
Polaris and Polycystin-2 in dorsal forerunner cells and Kupffer's vesicle are required for specification of the zebrafish left-right axis.
2005,
Pubmed
,
Xenbase
Boettger,
FGF8 functions in the specification of the right body side of the chick.
1999,
Pubmed
Bonnafe,
The transcription factor RFX3 directs nodal cilium development and left-right asymmetry specification.
2004,
Pubmed
Brody,
Ciliogenesis and left-right axis defects in forkhead factor HFH-4-null mice.
2000,
Pubmed
Draper,
Zebrafish fgf24 functions with fgf8 to promote posterior mesodermal development.
2003,
Pubmed
Draper,
Inhibition of zebrafish fgf8 pre-mRNA splicing with morpholino oligos: a quantifiable method for gene knockdown.
2001,
Pubmed
,
Xenbase
Eggenschwiler,
Cilia and developmental signaling.
2007,
Pubmed
Essner,
Kupffer's vesicle is a ciliated organ of asymmetry in the zebrafish embryo that initiates left-right development of the brain, heart and gut.
2005,
Pubmed
Fischer,
FGF8 acts as a right determinant during establishment of the left-right axis in the rabbit.
2002,
Pubmed
Fischer,
The zebrafish fgf24 mutant identifies an additional level of Fgf signaling involved in vertebrate forelimb initiation.
2003,
Pubmed
Gerdes,
Disruption of the basal body compromises proteasomal function and perturbs intracellular Wnt response.
2007,
Pubmed
Kramer-Zucker,
Cilia-driven fluid flow in the zebrafish pronephros, brain and Kupffer's vesicle is required for normal organogenesis.
2005,
Pubmed
Lee,
Fgf signaling instructs position-dependent growth rate during zebrafish fin regeneration.
2005,
Pubmed
Long,
The zebrafish nodal-related gene southpaw is required for visceral and diencephalic left-right asymmetry.
2003,
Pubmed
Meyers,
Differences in left-right axis pathways in mouse and chick: functions of FGF8 and SHH.
1999,
Pubmed
Millimaki,
Zebrafish atoh1 genes: classic proneural activity in the inner ear and regulation by Fgf and Notch.
2007,
Pubmed
Nonaka,
Randomization of left-right asymmetry due to loss of nodal cilia generating leftward flow of extraembryonic fluid in mice lacking KIF3B motor protein.
1998,
Pubmed
Park,
Dishevelled controls apical docking and planar polarization of basal bodies in ciliated epithelial cells.
2008,
Pubmed
,
Xenbase
Pownall,
An inducible system for the study of FGF signalling in early amphibian development.
2003,
Pubmed
,
Xenbase
Riley,
A critical period of ear development controlled by distinct populations of ciliated cells in the zebrafish.
1997,
Pubmed
Roehl,
Zebrafish pea3 and erm are general targets of FGF8 signaling.
2001,
Pubmed
Scholpp,
Zebrafish fgfr1 is a member of the fgf8 synexpression group and is required for fgf8 signalling at the midbrain-hindbrain boundary.
2004,
Pubmed
Schweickert,
Cilia-driven leftward flow determines laterality in Xenopus.
2007,
Pubmed
,
Xenbase
Tanaka,
FGF-induced vesicular release of Sonic hedgehog and retinoic acid in leftward nodal flow is critical for left-right determination.
2005,
Pubmed
Thummel,
Inhibition of zebrafish fin regeneration using in vivo electroporation of morpholinos against fgfr1 and msxb.
2006,
Pubmed
Urban,
FGF is essential for both condensation and mesenchymal-epithelial transition stages of pronephric kidney tubule development.
2006,
Pubmed
,
Xenbase
Zhang,
Receptor specificity of the fibroblast growth factor family. The complete mammalian FGF family.
2006,
Pubmed
