PUBLICATION

aura/mid1ip1L regulates the cytoskeleton at the zebrafish egg-to-embryo transition

Authors

Eno, C., Solanki, B., Pelegri, F.

ID

ZDB-PUB-160312-8

Date

2016

Source

Development (Cambridge, England) 143(9): 1585-99 (Journal)

Registered Authors

Pelegri, Francisco, Solanki, Bharti

Keywords

Mid1, Mid1ip1l, Cytoskeleton, Cortical granules, Membrane exocytosis, Cytokinesis, F-actin, Zebrafish, Opitz G/BBB syndrome

MeSH Terms

Cytoskeletal Proteins/genetics*
Cytoskeletal Proteins/metabolism
Animals
Cytoskeleton/metabolism*
Zebrafish Proteins/genetics*
Zebrafish Proteins/metabolism
Vesicle-Associated Membrane Protein 2/metabolism
Actins/metabolism
rab GTP-Binding Proteins/metabolism
Cytoplasmic Granules/metabolism
Cytokinesis/genetics*
Clathrin/metabolism
Dynamins/metabolism
Microtubules/metabolism*
Zebrafish/embryology*

(all 15)

PubMed

26965374 Full text @ Development

Abstract

Embryos from females homozygous for a recessive maternal-effect mutation in the gene aura exhibit defects including reduced cortical integrity, defective cortical granule (CG) release upon egg activation, failure to complete cytokinesis, and abnormal cell wound healing. Subcellular analysis shows that the cytokinesis defects observed in aura mutants are associated with aberrant cytoskeletal reorganization during furrow maturation, including abnormal F-actin enrichment and microtubule reorganization. Cortical F-actin prior to furrow formation fails to exhibit a normal transition into F-actin-rich arcs, and drug inhibition is consistent with aura function promoting F-actin polymerization and/or stabilization. In mutants, components of exocytic and endocytic vesicles, such as Vamp2, Clathrin and Dynamin, are sequestered in unreleased CGs, indicating a need for CG recycling in the normal redistribution of these factors. However, the exocytic targeting factor Rab11 is recruited to the furrow plane normally at the tip of bundling microtubules, suggesting an alternate anchoring mechanism independent of membrane recycling. A positional cloning approach indicates that the mutation in aura is associated with a truncation of Mid1 Interacting Protein 1L (Mid1ip1L), previously identified as an interactor of the X-linked Opitz G/BBB syndrome gene Mid1. A Cas9/CRISPR-induced mutant allele in mid1ip1L fails to complement the originally isolated aura maternal-effect mutation, confirming gene assignment. Mid1ip1L protein localizes to cortical F-actin aggregates, consistent with a direct role in cytoskeletal regulation. Our studies indicate that maternally provided aura/mid1ip1L acts during the reorganization of the cytoskeleton at the egg-to-embryo transition and highlight the importance of cytoskeletal dynamics and membrane recycling during this developmental period.

Genes / Markers

Figures

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Expression

Fish	Conditions	Stage	Anatomy	Assay	Figure
hm1Tg	standard conditions	1-cell	blastomere microtubule	IFL	Fig. S4
mid1ip1l^t9792/t9792; hm1Tg	standard conditions	1-cell	blastomere microtubule	IFL	Fig. S4
e114Tg	standard conditions	4-cell	blastomere contractile ring	IFL	Fig. 2
e114Tg	standard conditions	8-cell	blastomere contractile ring	IFL	Fig. 2
e114Tg	standard conditions	2-cell	blastomere contractile ring	IFL	Fig. 2
e114Tg	standard conditions	2-cell	blastomere filamentous actin	IFL	Fig. 2
e114Tg	standard conditions	4-cell	blastomere filamentous actin	IFL	Fig. 2
e114Tg	standard conditions	8-cell	blastomere filamentous actin	IFL	Fig. 2
mid1ip1l^t9792/t9792; e114Tg	standard conditions	2-cell	blastomere contractile ring	IFL	Fig. 2
mid1ip1l^t9792/t9792; e114Tg	standard conditions	4-cell	blastomere contractile ring	IFL	Fig. 2

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	8-cell	blastomere cleavage furrow ab2-ctnnb1 labeling decreased amount, abnormal	Fig. 9
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	2-cell	blastomere cleavage furrow morphology, abnormal	Fig. 9
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	8-cell	early embryonic cell cortical granule mislocalised, abnormal	Fig. 9
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	2-cell	yolk morphology, abnormal	Fig. 9
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	256-cell	blastomere cleavage furrow absent, abnormal	Fig. S1
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	256-cell	blastomere cleavage furrow incomplete structure, abnormal	Fig. S1
mid1ip1l^t9792/+; mid1ip1l^uw39/+	standard conditions	256-cell	blastomere cleavage furrow morphology, abnormal	Fig. S1
mid1ip1l^t9792/t9792	standard conditions	1-cell	blastodisc decreased height, abnormal	Fig. S2
mid1ip1l^t9792/t9792	standard conditions	1-cell	blastomere cleavage furrow mid1ip1l expression decreased amount, abnormal	Fig. 10
mid1ip1l^t9792/t9792	standard conditions	1-cell	blastomere cleavage furrow Ab2-cltc labeling decreased amount, abnormal	Fig. S5

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
e114Tg	Tg(actb2:LIFEACT-GFP)	Transgenic Insertion
hm1Tg	Tg(actb2:EMTB-3xEGFP)	Transgenic Insertion
t9792		Point Mutation	mid1ip1l
uw39		Insertion	mid1ip1l

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Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
mid1ip1l	CRISPR1-mid1ip1l	CRISPR

Fish

Fish
AB
e114Tg
hm1Tg
mid1ip1l^t9792/+; mid1ip1l^uw39/+
mid1ip1l^t9792/t9792
mid1ip1l^t9792/t9792; e114Tg
mid1ip1l^t9792/t9792; hm1Tg
mid1ip1l^uw39/uw39
WT

Antibodies

Name	Type	Antigen Genes	Isotypes	Host Organism
Ab1-dnm2	polyclonal		IgG	Rabbit
Ab1-rab11ba	polyclonal		IgG	Rabbit
Ab1-vamp2	polyclonal		IgG	Rabbit
Ab2-cltc	polyclonal		IgG	Rabbit
Ab2-ctnnb1	polyclonal	ctnnb1		Rabbit
Ab2-tuba	monoclonal		IgG1	Mouse

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Orthology

Gene	Orthology
mid1ip1a
mid1ip1b

Engineered Foreign Genes

Marker	Marker Type	Name
EGFP	EFG	EGFP
GFP	EFG	GFP

Mapping

Eno et al., 2016 ZDB-PUB-160312-8 PMID:26965374

aura/mid1ip1L regulates the cytoskeleton at the zebrafish egg-to-embryo transition

Eno et al., 2016

ZDB-PUB-160312-8
PMID:26965374