PUBLICATION

Wnt-PLC-IP₃-Connexin-Ca²⁺ axis maintains ependymal motile cilia in zebrafish spinal cord

Authors

Zhang, J., Chandrasekaran, G., Li, W., Kim, D.Y., Jeong, I.Y., Lee, S.H., Liang, T., Bae, J.Y., Choi, I., Kang, H., Maeng, J.S., Kim, M.K., Lee, T., Park, S.W., Kim, M.J., Kim, H.S., Ro, H., Bae, Y.C., Park, H.C., Choi, E.Y., Choi, S.Y.

ID

ZDB-PUB-200422-170

Date

2020

Source

Nature communications 11: 1860 (Journal)

Registered Authors

Choi, Seok-Yong, Park, Hae-Chul

Keywords

none

MeSH Terms

Zebrafish/embryology*
Zebrafish Proteins/genetics
Zebrafish Proteins/metabolism
Gap Junctions
Cell Differentiation
Ependyma/metabolism*
Ependyma/pathology
Spinal Cord/metabolism*
Mice, Knockout
Animals
Cilia/genetics
Cilia/metabolism*
Humans
Male
Connexin 43/genetics
Connexin 43/metabolism*
Wnt Signaling Pathway/genetics
Mice
Signal Transduction/genetics
Gene Knockout Techniques
Gene Expression Regulation, Developmental
Connexins/metabolism*

(all 22)

PubMed

32312952 Full text @ Nat. Commun.

Abstract

Ependymal cells (ECs) are multiciliated neuroepithelial cells that line the ventricles of the brain and the central canal of the spinal cord (SC). How ependymal motile cilia are maintained remains largely unexplored. Here we show that zebrafish embryos deficient in Wnt signaling have defective motile cilia, yet harbor intact basal bodies. With respect to maintenance of ependymal motile cilia, plcδ3a is a target gene of Wnt signaling. Lack of Connexin43 (Cx43), especially its channel function, decreases motile cilia and intercellular Ca²⁺ wave (ICW) propagation. Genetic ablation of cx43 in zebrafish and mice diminished motile cilia. Finally, Cx43 is also expressed in ECs of the human SC. Taken together, our findings indicate that gap junction mediated ICWs play an important role in the maintenance of ependymal motile cilia, and suggest that the enhancement of functional gap junctions by pharmacological or genetic manipulations may be adopted to ameliorate motile ciliopathy.

Genes / Markers

Figures

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Expression

Gene	Fish	Conditions	Stage	Anatomy	Assay	Figure
EGFP	hsc16Tg	control	Long-pec	spinal cord ependymal cell	IFL	Fig. 3
EGFP	hsc16Tg + MO6-gja1b	standard conditions	Long-pec	spinal cord ependymal cell	IFL	Fig. 3
foxj1a	AB	control	Long-pec	spinal cord ependymal cell	ISH	Fig. 1
foxj1a	AB	control	Long-pec	whole organism	RTPCR	Fig. 1
foxj1a	AB + MO3-wnt4b + MO4-wnt11	standard conditions	Long-pec	spinal cord ependymal cell	ISH	Fig. 1
foxj1a	AB + MO3-wnt4b + MO4-wnt11	standard conditions	Long-pec	whole organism	RTPCR	Fig. 1
fzd7b	AB	standard conditions	Long-pec	spinal cord ependymal cell	ISH	Fig. 1
GCaMP	cms5Tg	standard conditions	Long-pec	spinal cord ependymal cell	IFL	Fig. 4
GCaMP	cms5Tg + MO6-gja1b	standard conditions	Long-pec	spinal cord ependymal cell	IFL	Fig. 4
GFP	hsc5Tg	control	Long-pec	ependymal cell motile cilium	IFL	Fig. 3

1 - 10 of 18

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
AB	chemical treatment by environment: 6-bromoindirubin-3'-oxime	Long-pec	spinal cord ependymal cell plcd3a expression increased amount, abnormal	Fig. 2
AB	chemical treatment by environment: Carbenoxolone sodium	Long-pec	ependymal cell motile cilium assembly decreased occurrence, abnormal	Fig. 4
AB	chemical treatment by environment: Carbenoxolone sodium	Long-pec	spinal cord has fewer parts of type ependymal cell motile cilium, abnormal	Fig. 4
AB	chemical treatment by environment: DAPT	Long-pec	ependymal cell motile cilium assembly normal occurrence, normal	Fig. 1
AB	chemical treatment by environment: DAPT	Long-pec	spinal cord has normal numbers of parts of type ependymal cell motile cilium, normal	Fig. 1
AB	chemical treatment by environment: IWR-1-endo	Long-pec	ependymal cell ciliary basal body organization normal occurrence, normal	Fig. 1
AB	chemical treatment by environment: IWR-1-endo	Long-pec	ependymal cell motile cilium assembly decreased occurrence, abnormal	Fig. 1
AB	chemical treatment by environment: IWR-1-endo	Long-pec	spinal cord has fewer parts of type ependymal cell motile cilium, abnormal	Fig. 1
AB	chemical treatment by environment: thapsigargin	Day 4	ependymal cell ciliary basal body organization normal occurrence, normal	Fig. 2
AB	chemical treatment by environment: thapsigargin	Day 4	ependymal cell motile cilium assembly decreased occurrence, abnormal	Fig. 2

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
cms5Tg	Tg(foxj1a:GCaMP6s)	Transgenic Insertion
cms6		Small Deletion	gja1b
hsc5Tg	Tg(actb2:Mmu.Arl13b-GFP)	Transgenic Insertion
hsc16Tg	Tg(foxj1a:EGFP)	Transgenic Insertion
t4		Deficiency	shha
w32Tg	Tg(hsp70l:dkk1b-GFP)	Transgenic Insertion

1 - 6 of 6

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Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
fzd7b	MO2-fzd7b	MRPHLNO
gja1b	CRISPR1-gja1b	CRISPR
gja1b	MO6-gja1b	MRPHLNO
plcd3a	MO1-plcd3a	MRPHLNO
wnt4b	MO3-wnt4b	MRPHLNO
wnt5b	MO1-wnt5b	MRPHLNO
wnt11	MO4-wnt11	MRPHLNO

1 - 7 of 7

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Fish

Fish
AB
AB + MO1-wnt5b
AB + MO2-fzd7b
AB + MO3-wnt4b
AB + MO3-wnt4b + MO4-wnt11
AB + MO4-wnt11
AB + MO6-gja1b
cms5Tg
cms5Tg + MO6-gja1b
Df(Chr07)t4/t4

Antibodies

Name	Type	Isotypes	Host Organism
Ab1-tuba	monoclonal	IgG2b	Mouse
Ab1-tubg1	monoclonal	IgG1	Mouse
Ab22-gfap	polyclonal	IgG	Rabbit

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
EGFP	EFG	EGFP
GCaMP	EFG	GCaMP
GFP	EFG	GFP

Mapping

Zhang et al., 2020 ZDB-PUB-200422-170 PMID:32312952

Wnt-PLC-IP3-Connexin-Ca2+ axis maintains ependymal motile cilia in zebrafish spinal cord

Zhang et al., 2020

ZDB-PUB-200422-170
PMID:32312952

Wnt-PLC-IP₃-Connexin-Ca²⁺ axis maintains ependymal motile cilia in zebrafish spinal cord