PUBLICATION

Generation and characterization of Kctd15 mutations in zebrafish

Authors

Heffer, A., Marquart, G.D., Aquilina-Beck, A., Saleem, N., Burgess, H.A., Dawid, I.B.

ID

ZDB-PUB-171208-8

Date

2017

Source

PLoS One 12: e0189162 (Journal)

Registered Authors

Burgess, Harold, Dawid, Igor B., Marquart, Gregory

Keywords

none

MeSH Terms

Zebrafish Proteins/genetics*
Zebrafish/genetics*
Frameshift Mutation*
Animals
Potassium Channels, Voltage-Gated/genetics*

PubMed

29216270 Full text @ PLoS One

Abstract

Potassium channel tetramerization domain containing 15 (Kctd15) was previously found to have a role in early neural crest (NC) patterning, specifically delimiting the region where NC markers are expressed via repression of transcription factor AP-2a and inhibition of Wnt signaling. We used transcription activator-like effector nucleases (TALENs) to generate null mutations in zebrafish kctd15a and kctd15b paralogs to study the in vivo role of Kctd15. We found that while deletions producing frame-shift mutations in each paralog showed no apparent phenotype, kctd15a/b double mutant zebrafish are smaller in size and show several phenotypes including some affecting the NC, such as expansion of the early NC domain, increased pigmentation, and craniofacial defects. Both melanophore and xanthophore pigment cell numbers and early markers are up-regulated in the double mutants. While we find no embryonic craniofacial defects, adult mutants have a deformed maxillary segment and missing barbels. By confocal imaging of mutant larval brains we found that the torus lateralis (TLa), a region implicated in gustatory networks in other fish, is absent. Ablation of this brain tissue in wild type larvae mimics some aspects of the mutant growth phenotype. Thus kctd15 mutants show deficits in the development of both neural crest derivatives, and specific regions within the central nervous system, leading to a strong reduction in normal growth rates.

Genes / Markers

Figures

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Expression

Gene	Fish	Conditions	Stage	Anatomy	Assay	Figure
col10a1a	AB	control	Day 6	maxilla	ISH	Fig. 4
col10a1a	kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	Day 6	maxilla	ISH	Fig. 4
dlx3b	AB	control	5-9 somites	neural plate border	ISH	Fig. 1
dlx3b	kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural plate border	ISH	Fig. 1
EGFP	ba2Tg	control	Prim-5	migratory neural crest cell	IFL	Fig. 1
EGFP	kctd15a^y584/y584; kctd15b^y585/y585; ba2Tg	control	Prim-5	migratory neural crest cell	IFL	Fig. 1
foxd3	AB	control	5-9 somites	neural crest	ISH	Fig. 1
foxd3	AB	control	1-4 somites	neural crest	ISH	Fig. 1
foxd3	kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	1-4 somites	neural crest	ISH	Fig. 1
foxd3	kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural crest	ISH	Fig. 1

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	1-4 somites	neural crest foxd3 expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	1-4 somites	neural crest sox10 expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural crest foxd3 expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural crest snai1b expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural crest sox10 expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural crest sox9b expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural crest tfap2a expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	5-9 somites	neural plate border dlx3b expression increased amount, abnormal	Fig. 1
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	Prim-5	head pigment cell gch2 expression increased amount, abnormal	Fig. 3
kctd15a^y584/y584; kctd15b^y585/y585 (AB)	control	Prim-5	melanocyte tyrp1a expression increased amount, abnormal	Fig. 2

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
ba2Tg	Tg(-4.9sox10:EGFP)	Transgenic Insertion
y534Tg	Tg(-1.5kctd15a:GFP)	Transgenic Insertion
y584		Small Deletion	kctd15a
y585		Small Deletion	kctd15b

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Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
kctd15a	TALEN1-kctd15a	TALEN
kctd15b	TALEN1-kctd15b	TALEN

Fish

Fish
AB
ba2Tg
kctd15a^y584/y584 (AB)
kctd15a^y584/y584; kctd15b^y585/y585 (AB)
kctd15a^y584/y584; kctd15b^y585/y585; ba2Tg
kctd15b^y585/y585 (AB)
WT
y534Tg

Antibodies

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
EGFP	EFG	EGFP
GFP	EFG	GFP

Mapping

Heffer et al., 2017 ZDB-PUB-171208-8 PMID:29216270

Generation and characterization of Kctd15 mutations in zebrafish

Heffer et al., 2017

ZDB-PUB-171208-8
PMID:29216270