PUBLICATION

foxc1 is required for embryonic head vascular smooth muscle differentiation in zebrafish

Authors

Whitesell, T.R., Chrystal, P.W., Ryu, J.R., Munsie, N., Grosse, A., French, C.R., Workentine, M.L., Li, R., Zhu, L.J., Waskiewicz, A., Lehmann, O.J., Lawson, N.D., Childs, S.J.

ID

ZDB-PUB-190615-13

Date

2019

Source

Developmental Biology 453(1): 34-47 (Journal)

Registered Authors

Childs, Sarah J., French, Curtis R., Lawson, Nathan, Lehmann, Ordan J., Waskiewicz, Andrew

Keywords

CRISPR, RNA-Seq transcriptome, Vascular smooth muscle cell, Zebrafish, acta2, foxc1b

Datasets

GEO:GSE119718

MeSH Terms

Gene Expression Regulation, Developmental
Embryo, Nonmammalian/cytology*
Embryo, Nonmammalian/metabolism
Zebrafish Proteins/metabolism*
Forkhead Transcription Factors/metabolism*
Pericytes/metabolism
Brain/embryology
Brain/metabolism
Transcriptome/genetics
Head/blood supply*
Head/embryology*
Mesoderm/metabolism
Up-Regulation
Animals
Myocytes, Smooth Muscle/metabolism
Cell Differentiation*/genetics
Endothelium/metabolism
Muscle, Smooth, Vascular/cytology*
Zebrafish/embryology*
Zebrafish/genetics

(all 20)

PubMed

31199900 Full text @ Dev. Biol.

Abstract

Vascular smooth muscle of the head derives from neural crest, but developmental mechanisms and early transcriptional drivers of the vSMC lineage are not well characterized. We find that in early development, the transcription factor foxc1b is expressed in mesenchymal cells that associate with the vascular endothelium. Using timelapse imaging, we observe that foxc1b expressing mesenchymal cells differentiate into acta2 expressing vascular mural cells. We show that in zebrafish, while foxc1b is co-expressed in acta2 positive smooth muscle cells that associate with large diameter vessels, it is not co-expressed in capillaries where pdgfrβ positive pericytes are located. In addition to being an early marker of the lineage, foxc1 is essential for vSMC differentiation; we find that foxc1 loss of function mutants have defective vSMC differentiation and that early genetic ablation of foxc1b or acta2 expressing populations blocks vSMC differentiation. Furthermore, foxc1 is expressed upstream of acta2 and is required for acta2 expression in vSMCs. Using RNA-Seq we determine an enriched intersectional gene expression profile using dual expression of foxc1b and acta2 to identify novel vSMC markers. Taken together, our data suggests that foxc1 is a marker of vSMCs and plays a critical functional role in promoting their differentiation.

Genes / Markers

Figures

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Expression

Gene	Fish	Conditions	Stage	Anatomy	Assay	Figure
acta2	c264Tg/+; ca62Tg/+	chemical treatment: metronidazole	Day 4	vascular associated smooth muscle cell	ISH	Fig. 9
acta2	c264Tg/+; ca72Tg/+	chemical treatment: metronidazole	Day 4	vascular associated smooth muscle cell	ISH	Fig. 9
acta2	foxc1a^{ua1017/ua1017}	standard conditions	Day 4	vascular associated smooth muscle cell	ISH	Fig. 8
acta2	foxc1a^{ua1017/ua1017}; foxc1b^{ua1018/ua1018}	standard conditions	Day 4	vascular associated smooth muscle cell	ISH	Fig. 8
acta2	foxc1b^{ua1018/ua1018}	standard conditions	Day 4	vascular associated smooth muscle cell	ISH	Fig. 8
acta2	WT	standard conditions	Day 4	vascular associated smooth muscle cell	ISH	Fig. 8
capn3b	ca7Tg	standard conditions	Day 4	aortic arch vascular associated smooth muscle cell	ISH	Fig. 7
capn3b	ca7Tg	standard conditions	Day 4	aortic arch	ISH	Fig. 7
capn3b	ca7Tg	standard conditions	Day 4	brain vascular associated smooth muscle cell	ISH	Fig. 7
capn3b	ca7Tg	standard conditions	Day 4	brain	ISH	Fig. 7

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
c264Tg/+; ca62Tg/+	chemical treatment: metronidazole	Day 4	vascular associated smooth muscle cell acta2 expression decreased amount, abnormal	Fig. 9
c264Tg/+; ca72Tg/+	chemical treatment: metronidazole	Day 4	pericyte pdgfrb expression increased amount, abnormal	Fig. 9
c264Tg/+; ca72Tg/+	chemical treatment: metronidazole	Day 4	vascular associated smooth muscle cell acta2 expression decreased amount, abnormal	Fig. 9
c264Tg/+; ca72Tg/+	chemical treatment: metronidazole	Day 4	vascular associated smooth muscle cell foxc1b expression decreased amount, abnormal	Fig. 9
foxc1a^ua1017/+; ca7Tg/+; ci5Tg/+	standard conditions	Protruding-mouth	ventral aorta length, normal	Fig. 8
foxc1a^ua1017/+; ca7Tg/+; ci5Tg/+	standard conditions	Protruding-mouth	ventral aorta vascular associated smooth muscle cell amount, normal	Fig. 8
foxc1a^ua1017/+; ca7Tg/+; ci5Tg/+	standard conditions	Day 4	ventral aorta length, normal	Fig. 8
foxc1a^ua1017/+; ca7Tg/+; ci5Tg/+	standard conditions	Day 4	ventral aorta vascular associated smooth muscle cell amount, normal	Fig. 8
foxc1a^ua1017/+; foxc1b^ua1018/+; ca7Tg/+; ci5Tg/+	standard conditions	Protruding-mouth	ventral aorta length, normal	Fig. 8
foxc1a^ua1017/+; foxc1b^ua1018/+; ca7Tg/+; ci5Tg/+	standard conditions	Protruding-mouth	ventral aorta vascular associated smooth muscle cell amount, normal	Fig. 8

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Mutations / Transgenics

Allele	Construct	Type
c264Tg	Tg(UAS-E1B:NTR-mCherry)	Transgenic Insertion
ca7Tg	Tg(acta2:EGFP)	Transgenic Insertion
ca8Tg	Tg(acta2:mCherry)	Transgenic Insertion
ca41Tg	TgBAC(pdgfrb:EGFP)	Transgenic Insertion
ca42Tg	TgBAC(pdgfrb:GAL4FF)	Transgenic Insertion
ca62Tg	Tg(acta2:GAL4FF,myl7:EGFP)	Transgenic Insertion
ca72Tg	Tg(foxc1b:GAL4FF,myl7:EGFP)	Transgenic Insertion
ci5Tg	Tg(-6.5kdrl:mCherry)	Transgenic Insertion
la116Tg	Tg(kdrl:GFP)	Transgenic Insertion
mw44Tg	Tg(foxc1b:EGFP)	Transgenic Insertion

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Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
foxc1a	CRISPR12-foxc1a	CRISPR
foxc1a	CRISPR13-foxc1a	CRISPR
foxc1b	CRISPR2-foxc1b	CRISPR
foxc1b	CRISPR4-foxc1b	CRISPR

Fish

Fish
c264Tg; ca42Tg; mw44Tg
c264Tg/+; ca62Tg/+
c264Tg/+; ca72Tg/+
ca7Tg
ca7Tg/+; ci5Tg/+
ca7Tg; ci5Tg
ca8Tg; mw44Tg
ci5Tg; mw44Tg
foxc1a^ua1017/+; ca7Tg/+; ci5Tg/+
foxc1a^ua1017/+; foxc1b^ua1018/+; ca7Tg/+; ci5Tg/+

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Antibodies

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
EGFP	EFG	EGFP
GAL4FF	EFG	GAL4FF
GFP	EFG	GFP
mCherry	EFG	mCherry
NTR	EFG	NTR

Mapping

Whitesell et al., 2019 ZDB-PUB-190615-13 PMID:31199900

foxc1 is required for embryonic head vascular smooth muscle differentiation in zebrafish

Whitesell et al., 2019

ZDB-PUB-190615-13
PMID:31199900