PUBLICATION

Investigation of Islet2a function in zebrafish embryos: Mutants and morphants differ in morphologic phenotypes and gene expression

Authors

Moreno, R.L., Williams, K., Jones, K.L., Ribera, A.B.

ID

ZDB-PUB-180622-43

Date

2018

Source

PLoS One 13: e0199233 (Journal)

Registered Authors

Ribera, Angie, Williams, Kris

Keywords

none

MeSH Terms

RNA, Messenger/genetics
RNA, Messenger/metabolism
Heterozygote
Phenotype
Larva/genetics
Animals
Zebrafish Proteins/genetics*
Zebrafish Proteins/metabolism
Morpholinos/pharmacology*
Transcription Activator-Like Effector Nucleases/metabolism
Transcription Factors/genetics*
Transcription Factors/metabolism
Motor Neurons/drug effects
Motor Neurons/metabolism
Base Sequence
Zebrafish/embryology*
Zebrafish/genetics*
Gene Knockdown Techniques
Mutagenesis/genetics
Mutation/genetics*
Gene Expression Regulation, Developmental*/drug effects
LIM-Homeodomain Proteins/genetics*
LIM-Homeodomain Proteins/metabolism
Gene Editing
Embryo, Nonmammalian/drug effects
Embryo, Nonmammalian/metabolism*

(all 26)

PubMed

29927984 Full text @ PLoS One

Abstract

Zebrafish primary motor neurons differ from each other with respect to morphology, muscle targets and electrophysiological properties. For example, CaP has 2-3-fold larger densities of both inward and outward currents than do other motor neurons. We tested whether the transcription factor Islet2a, uniquely expressed in CaP, but not other primary motor neurons, plays a role in specifying its stereotypic electrophysiological properties. We used both TALEN-based gene editing and antisense morpholino approaches to disrupt Islet2a function. Our electrophysiology results do not support a specific role for Islet2a in determining CaP's unique electrical properties. However, we also found that the morphological phenotypes of CaP and a later-born motor neuron differed between islet2a mutants and morphants. Using microarrays, we tested whether the gene expression profiles of whole embryo morphants, mutants and controls also differed. Morphants had 174 and 201 genes that were differentially expressed compared to mutants and controls, respectively. Further, islet2a was identified as a differentially expressed gene. To examine how mutation of islet2a affected islet gene expression specifically in CaPs, we performed RNA in situ hybridization. We detected no obvious differences in expression of islet1, islet2a, or islet2b in CaPs of mutant versus sibling control embryos. However, immunolabeling studies revealed that an Islet protein persisted in CaPs of mutants, albeit at a reduced level compared to controls. While we cannot exclude requirement for some Islet protein, we conclude that differentiation of the CaP's stereotypic large inward and outward currents does not have a specific requirement for Islet2a.

Genes / Markers

Figures

Show all Figures

Expression

Gene	Fish	Conditions	Stage	Anatomy	Assay	Figure
GFP	isl2a^{co7002/co7002}; ml2Tg	standard conditions	Prim-5	secondary motor neuron axon	IFL	Fig. 4
GFP	isl2a^{co7002/co7002}; ml2Tg	standard conditions	Prim-5	secondary motor neuron neuronal cell body	IFL	Fig. 4
GFP	isl2a^{co7002/co7002}; rw0Tg	standard conditions	Protruding-mouth	secondary motor neuron	IFL	Fig. 5
GFP	isl2a^co7002/+; ml2Tg	standard conditions	Prim-5	CaP motoneuron	IHC	Fig. 9
GFP	isl2a^co7002/+; ml2Tg	standard conditions	Prim-5	motor neuron	IHC	Fig. 8
GFP	isl2a^co7002/+; ml2Tg	standard conditions	Prim-5	secondary motor neuron axon	IFL	Fig. 4
GFP	isl2a^co7002/+; ml2Tg	standard conditions	Prim-5	secondary motor neuron neuronal cell body	IFL	Fig. 4
GFP	isl2a^co7002/+; rw0Tg	standard conditions	Protruding-mouth	secondary motor neuron	IFL	Fig. 5
GFP	ml2Tg	control	Prim-5	CaP motoneuron axon	IFL	Fig. 4
GFP	ml2Tg	control	Prim-5	CaP motoneuron neuronal cell body	IFL	Fig. 4

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
WT + MO1-isl2a	standard conditions	Long-pec	whole organism isl2a expression increased amount, abnormal	Fig. 7
isl2a^co7002/+; ml2Tg	standard conditions	Prim-5	CaP motoneuron ab2-isl labeling decreased amount, abnormal	Fig. 9
isl2a^co7002/+; ml2Tg	standard conditions	Prim-5	CaP motoneuron morphology, normal	Fig. 4
isl2a^co7002/+; rw0Tg	standard conditions	Protruding-mouth	secondary motor neuron amount, normal	Fig. 5
isl2a^co7002/+; rw0Tg	standard conditions	Protruding-mouth	secondary motor neuron morphology, normal	Fig. 5
isl2a^{co7002/co7002}	standard conditions	Prim-5	CaP motoneuron morphology, normal	Fig. 4
isl2a^{co7002/co7002}	standard conditions	Prim-5	proctodeum isl2a expression absent, abnormal	Fig. 1
isl2a^{co7002/co7002}	standard conditions	Prim-5	proctodeum ab2-isl labeling absent, abnormal	Fig. 1
isl2a^{co7002/co7002}; ml2Tg	standard conditions	Prim-5	CaP motoneuron morphology, normal	Fig. 4
isl2a^{co7002/co7002}; rw0Tg	standard conditions	Protruding-mouth	secondary motor neuron amount, normal	Fig. 5

1 - 10 of 16

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
co7002		Small Deletion	isl2a
co7003		Small Deletion	isl2a
ml2Tg	Tg(mnx1:GFP)	Transgenic Insertion
rw0Tg	Tg(isl1a:GFP)	Transgenic Insertion
zf35Tg	Tg(-7.3gata2a:GFP)	Transgenic Insertion

1 - 5 of 5

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Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
isl2a	MO1-isl2a	MRPHLNO
isl2a	MO6-isl2a	MRPHLNO
isl2a	TALEN3-isl2a	TALEN

Fish

Fish
isl2a^{co7002/co7002}
isl2a^{co7002/co7002}; ml2Tg
isl2a^{co7002/co7002}; rw0Tg
isl2a^co7002/+; ml2Tg
isl2a^co7002/+; rw0Tg
isl2a^{co7003/co7003}
ml2Tg
ml2Tg + MO1-isl2a
rw0Tg
rw0Tg + MO1-isl2a

Antibodies

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
GFP	EFG	GFP

Mapping

Moreno et al., 2018 ZDB-PUB-180622-43 PMID:29927984

Investigation of Islet2a function in zebrafish embryos: Mutants and morphants differ in morphologic phenotypes and gene expression

Moreno et al., 2018

ZDB-PUB-180622-43
PMID:29927984