PUBLICATION

Zebrafish snai2 mutants fail to phenocopy morphant phenotypes

Authors

Bickers, C., Española, S.D., Grainger, S., Pouget, C., Traver, D.

ID

ZDB-PUB-180913-28

Date

2018

Source

PLoS One 13: e0202747 (Journal)

Registered Authors

Grainger, Stephanie, Traver, David

Keywords

none

MeSH Terms

Amino Acid Sequence
Animals
Cell Differentiation
Embryo, Nonmammalian/metabolism
Forkhead Transcription Factors/metabolism
Gene Expression Regulation, Developmental
Genotype
Hematopoietic Stem Cells/cytology
Hematopoietic Stem Cells/metabolism
Morpholinos/metabolism
Mutagenesis, Site-Directed
PAX9 Transcription Factor/metabolism
Phenotype
Receptors, Notch/metabolism
Signal Transduction
Snail Family Transcription Factors/antagonists & inhibitors
Snail Family Transcription Factors/genetics*
Snail Family Transcription Factors/metabolism
Zebrafish/genetics
Zebrafish/growth & development
Zebrafish/metabolism*
Zebrafish Proteins/antagonists & inhibitors
Zebrafish Proteins/genetics*
Zebrafish Proteins/metabolism

(all 24)

PubMed

30208064 Full text @ PLoS One

Abstract

Snail2 is a zinc-finger transcription factor best known to repress expression of genes encoding cell adherence proteins to facilitate induction of the epithelial-to-mesenchymal transition. While this role has been best documented in the developmental migration of the neural crest and mesoderm, here we expand on previously reported preliminary findings that morpholino knock-down of snai2 impairs the generation of hematopoietic stem cells (HSCs) during zebrafish development. We demonstrate that snai2 morphants fail to initiate HSC specification and show defects in the somitic niche of migrating HSC precursors. These defects include a reduction in sclerotome markers as well as in the Notch ligands dlc and dld, which are known to be essential components of HSC specification. Accordingly, enforced expression of the Notch1-intracellular domain was capable of rescuing HSC specification in snai2 morphants. To parallel our approach, we obtained two mutant alleles of snai2. In contrast to the morphants, homozygous mutant embryos displayed no defects in HSC specification or in sclerotome development, and mutant fish survive into adulthood. However, when these homozygous mutants were injected with snai2 morpholino, HSCs were improperly specified. In summary, our morpholino data support a role for Snai2 in HSC development, whereas our mutant data suggest that Snai2 is dispensable for this process. Together, these findings further support the need for careful consideration of both morpholino and mutant phenotypes in studies of gene function.

Genes / Markers

Marker	Marker Type	Name
dlc	GENE	deltaC
dld	GENE	deltaD
eef1a1l1	GENE	eukaryotic translation elongation factor 1 alpha 1, like 1
etsrp	GENE	ETS1-related protein
fli1	GENE	Fli-1 proto-oncogene, ETS transcription factor
foxc1b	GENE	forkhead box C1b
jam2a	GENE	junctional adhesion molecule 2a
kdrl	GENE	kinase insert domain receptor like
meox1	GENE	mesenchyme homeobox 1
myb	GENE	v-myb avian myeloblastosis viral oncogene homolog

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Figures

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Expression

Phenotype

Fish	Conditions	Stage	Phenotype	Figure
AB + MO1-snai2	standard conditions	10-13 somites	somite dlc expression decreased amount, abnormal	Fig. 5
AB + MO1-snai2	standard conditions	10-13 somites	somite dld expression decreased amount, abnormal	Fig. 5
AB + MO1-snai2	standard conditions	14-19 somites	sclerotome pax9 expression decreased amount, abnormal	Fig. 4
AB + MO1-snai2	standard conditions	Prim-5	sclerotome foxc1b expression decreased amount, abnormal	Fig. 4
AB + MO1-snai2	standard conditions	Prim-5	ventral wall of dorsal aorta hematopoietic stem cell runx1 expression decreased amount, abnormal	Fig. 3
AB + MO1-snai2	standard conditions	Prim-5	ventral wall of dorsal aorta hematopoietic stem cell runx1 expression decreased amount, abnormal	Fig. 5
WT + MO1-snai2	standard conditions	Prim-5	ventral wall of dorsal aorta hematopoietic stem cell runx1 expression decreased amount, abnormal	Fig. 9
hzm3Tg; s896Tg; sd32Tg + MO1-snai2	standard conditions	Long-pec	ventral wall of dorsal aorta hematopoietic stem cell decreased amount, abnormal	Fig. 3
hzm3Tg; s896Tg; sd32Tg + MO1-snai2	standard conditions	Long-pec	ventral wall of dorsal aorta hematopoietic stem cell decreased amount, abnormal	Fig. 9
snai2^sd57/+ + MO1-snai2	standard conditions	Prim-5	ventral wall of dorsal aorta hematopoietic stem cell runx1 expression decreased amount, abnormal	Fig. 9

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
bw9Tg	Tg(kdrl:GAL4)	Transgenic Insertion
hzm3Tg	Tg(4xUAS:GFP)	Transgenic Insertion
kca3Tg	Tg(5xUAS-E1B:6xMYC-notch1a)	Transgenic Insertion
la2Tg	Tg(-6.0itga2b:EGFP)	Transgenic Insertion
mu271Tg	Tg(UAS:LIFEACT-GFP)	Transgenic Insertion
s896Tg	Tg(kdrl:Hsa.HRAS-mCherry)	Transgenic Insertion
sa24539		Point Mutation	snai2
sd32Tg	TgBAC(gata2b:KALTA4)	Transgenic Insertion
sd57		Small Deletion	snai2
um14Tg	Tg(EPV.Tp1-Mmu.Hbb:EGFP)	Transgenic Insertion

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Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
snai2	CRISPR1-snai2	CRISPR
snai2	CRISPR2-snai2	CRISPR
snai2	CRISPR3-snai2	CRISPR
snai2	MO1-snai2	MRPHLNO
snai2	MO2-snai2	MRPHLNO

Fish

Fish
AB
AB + MO1-snai2
hzm3Tg; s896Tg; sd32Tg
hzm3Tg; s896Tg; sd32Tg + MO1-snai2
la2Tg; s896Tg
snai2^{sa24539/sa24539}
snai2^sd57/sd57
snai2^sd57/sd57; hzm3Tg; s896Tg; sd32Tg
snai2^sd57/sd57; hzm3Tg; s896Tg; sd32Tg + MO1-snai2
snai2^sd57/sd57 + MO1-snai2

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Antibodies

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
EGFP	EFG	EGFP
GAL4	EFG	GAL4
GFP	EFG	GFP
KALTA4	EFG	KALTA4
mCherry	EFG	mCherry

Mapping

Bickers et al., 2018 ZDB-PUB-180913-28 PMID:30208064

Zebrafish snai2 mutants fail to phenocopy morphant phenotypes

Bickers et al., 2018

ZDB-PUB-180913-28
PMID:30208064