PUBLICATION

Functional analysis of Sonic Hedgehog variants associated with holoprosencephaly in humans using a CRISPR/Cas9 zebrafish model

Authors

Hong, S., Hu, P., Jang, J.H., Carrington, B., Sood, R., Berger, S.I., Roessler, E., Muenke, M.

ID

ZDB-PUB-201002-85

Date

2020

Source

Human Mutation 41(12): 2155-2166 (Journal)

Registered Authors

Sood, Raman

Keywords

CRISPR/Cas9, hedgehog signaling, holoprosencephaly, variant reinterpretation, zebrafish

MeSH Terms

Alleles
Animals
CRISPR-Cas Systems/genetics*
Disease Models, Animal
Family
Genetic Association Studies
Genetic Variation*
Guidelines as Topic
Hedgehog Proteins/genetics*
Holoprosencephaly/genetics*
Humans
Loss of Function Mutation/genetics
Mutation/genetics
Phenotype
RNA, Messenger/genetics
RNA, Messenger/metabolism
Societies, Scientific
Zebrafish/genetics*
Zebrafish Proteins/genetics*

(all 19)

PubMed

32939873 Full text @ Hum. Mutat.

Abstract

Genetic variation in the highly conserved Sonic Hedgehog (SHH) gene is one of the most common genetic causes for the malformations of the brain and face in humans described as the holoprosencephaly clinical spectrum. However, only a minor fraction of known SHH variants have been experimentally proven to lead to abnormal function. Employing a phenotypic rescue assay with synthetic human messenger RNA variant constructs in shha^-/- knockout zebrafish, we evaluated 104 clinically reported in-frame and missense SHH variants. Our data helped us to classify them into loss of function variants (31), hypomorphic variants (33), and nonpathogenic variants (40). We discuss the strengths and weaknesses of currently accepted predictors of variant deleteriousness and the American College of Medical Genetics and Genomics guidelines for variant interpretation in the context of this functional model; furthermore, we demonstrate the robustness of model systems such as zebrafish as a rapid method to resolve variants of uncertain significance.

Genes / Markers

Marker	Marker Type	Name
ptch1	GENE	patched 1
ptch2	GENE	patched 2
shha	GENE	sonic hedgehog signaling molecule a
shhb	GENE	sonic hedgehog signaling molecule b

Figures

Show all Figures

Expression

Phenotype

Fish	Conditions	Stage	Phenotype	Figure
shha^hg126/hg126	standard conditions	Long-pec	caudal fin curled, abnormal	Fig. 1
shha^hg126/hg126	standard conditions	Long-pec	eye fused with eye, abnormal	Fig. 1
shha^hg126/hg126	standard conditions	Long-pec	somite U-shaped, abnormal	Fig. 1
shha^hg126/hg126; shhb^hg127/hg127	standard conditions	Long-pec	caudal fin curled, abnormal	Fig. 1
shha^hg126/hg126; shhb^hg127/hg127	standard conditions	Long-pec	eye fused with eye, abnormal	Fig. 1
shha^hg126/hg126; shhb^hg127/hg127	standard conditions	Long-pec	forebrain hypoplastic, abnormal	Fig. 1
shha^hg126/hg126; shhb^hg127/hg127	standard conditions	Long-pec	forebrain morphology, abnormal	Fig. 1
shha^hg126/hg126; shhb^hg127/hg127	standard conditions	Long-pec	somite U-shaped, abnormal	Fig. 1
shha^hg126/hg126; shhb^hg127/hg127	standard conditions	Long-pec	whole organism ptch2 expression absent, abnormal	Fig. 1

1 - 9 of 9

Show

Mutations / Transgenics

Allele	Type	Affected Genomic Region
hg125	Small Deletion	shha
hg126	Small Deletion	shha
hg127	Small Deletion	shhb
hg128	Small Deletion	shhb

Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
ptch1	MO1-ptch1	MRPHLNO
ptch2	MO1-ptch2	MRPHLNO
shha	CRISPR1-shha	CRISPR
shhb	CRISPR1-shhb	CRISPR

Fish

Fish
shha^hg126/hg126
shha^hg126/hg126; shhb^hg127/hg127
WT

Antibodies

Orthology

Engineered Foreign Genes

Mapping

Hong et al., 2020 ZDB-PUB-201002-85 PMID:32939873

Functional analysis of Sonic Hedgehog variants associated with holoprosencephaly in humans using a CRISPR/Cas9 zebrafish model

Hong et al., 2020

ZDB-PUB-201002-85
PMID:32939873