PUBLICATION

Pacsin2 is required for endocytosis in the zebrafish pronephric tubule

Authors

Morgan, J., Yarwood, R., Starborg, T., Yan, G., Lowe, M.

ID

ZDB-PUB-220527-7

Date

2022

Source

Biology Open 11(6): (Journal)

Registered Authors

Lowe, Martin

Keywords

Endocytosis, Megalin, Proximal tubule, Zebrafish, pacsin2

MeSH Terms

Animals
Endocytosis/physiology
Mammals/metabolism
Biological Transport
Kidney Tubules, Proximal/metabolism
Zebrafish*/metabolism
Low Density Lipoprotein Receptor-Related Protein-2*/genetics
Low Density Lipoprotein Receptor-Related Protein-2*/metabolism

(all 8)

PubMed

35616009 Full text @ Biol. Open

Abstract

Endocytosis mediates the cellular uptake of numerous molecules from the extracellular space and is a fundamentally important process. In the renal proximal tubule, the scavenger receptor megalin and its co-receptor cubilin mediate endocytosis of low molecular weight proteins from the renal filtrate. However, the extent to which megalin endocytosis relies on different components of the trafficking machinery remains relatively poorly defined in vivo. In this study, we identify a functional requirement for the F-BAR protein pacsin2 in endocytosis in the renal proximal tubule of zebrafish larvae. Pacsin2 is expressed throughout development and in all zebrafish tissues, similar to the mammalian orthologue. Within renal tubular epithelial cells, pacsin2 is enriched at the apical pole where it is localised to endocytic structures. Loss of pacsin2 results in reduced endocytosis within the proximal tubule, which is accompanied by a reduction in the abundance of megalin and endocytic organelles. Our results indicate that pacsin2 is required for efficient endocytosis in the proximal tubule, where it likely cooperates with other trafficking machinery to maintain endocytic uptake and recycling of megalin.

Genes / Markers

Figures

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Expression

Fish	Conditions	Stage	Anatomy	Assay	Figure
AB	control	Protruding-mouth	pronephric proximal convoluted tubule early endosome	IHC	Fig. 4
AB + MO1-pacsin2	control	Protruding-mouth	pronephric proximal convoluted tubule early endosome	IHC	Fig. 4
mitfa^w2/w2; pacsin2^umc405/+; mpv17^a9/a9 (AB)	standard conditions	Day 4	pronephric proximal convoluted tubule early endosome	IHC	Fig. 4
mitfa^w2/w2; pacsin2^umc405/+; pacsin2^umc406/+; mpv17^a9/a9 (AB)	standard conditions	Day 4	pronephric proximal convoluted tubule early endosome	IHC	Fig. 4
mitfa^w2/w2; pacsin2^umc406/+; mpv17^a9/a9 (AB)	standard conditions	Day 4	pronephric proximal convoluted tubule early endosome	IHC	Fig. 4
AB	standard conditions	Adult	brain	RTPCR	Fig. 1
AB	standard conditions	Adult	eye	RTPCR	Fig. 1
AB	standard conditions	Adult	gonad	RTPCR	Fig. 1
AB	standard conditions	Adult	heart	RTPCR	Fig. 1
AB	standard conditions	Adult	kidney	RTPCR	Fig. 1

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
AB + MO1-pacsin2	control	Protruding-mouth	pronephric proximal convoluted tubule early endosome lrp2a expression decreased distribution, abnormal	Fig. 4
AB + MO1-pacsin2	control	Protruding-mouth	pronephric proximal convoluted tubule early endosome pacsin2 expression absent, abnormal	Fig. 4
AB + MO1-pacsin2	control	Protruding-mouth	pronephric proximal convoluted tubule early endosome ab1-eea1 labeling decreased distribution, abnormal	Fig. 4
AB + MO1-pacsin2	control	Protruding-mouth	pronephric proximal convoluted tubule early endosome Ab5-rab11 labeling decreased distribution, abnormal	Fig. 4
AB + MO1-pacsin2	control	Day 4	renal tubule endocytosis decreased process quality, abnormal	Fig. 3
AB + MO2-pacsin2	control	Day 4	renal tubule endocytosis decreased process quality, abnormal	Fig. 3
mitfa^w2/w2; pacsin2^umc405/+; mpv17^a9/a9 (AB)	control	Day 4	renal tubule endocytosis normal process quality, normal	Fig. 3
mitfa^w2/w2; pacsin2^umc405/+; mpv17^a9/a9 (AB)	standard conditions	Sphere	whole organism embryo development normal process quality, normal	Fig. 2
mitfa^w2/w2; pacsin2^umc405/+; mpv17^a9/a9 (AB)	standard conditions	Sphere	whole organism morphology, normal	Fig. 2
mitfa^w2/w2; pacsin2^umc405/+; mpv17^a9/a9 (AB)	standard conditions	1-4 somites	whole organism embryo development normal process quality, normal	Fig. 2

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Mutations / Transgenics

Allele	Type	Affected Genomic Region
a9	Complex	mpv17
umc405	Indel	pacsin2
umc406	Indel	pacsin2
w2	Point Mutation	mitfa

Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
pacsin2	CRISPR2-pacsin2	CRISPR
pacsin2	MO1-pacsin2	MRPHLNO
pacsin2	MO2-pacsin2	MRPHLNO

Fish

Fish
AB
AB + MO1-pacsin2
AB + MO2-pacsin2
mitfa^w2/w2; mpv17^a9/a9 (AB)
mitfa^w2/w2; pacsin2^umc405/+; mpv17^a9/a9 (AB)
mitfa^w2/w2; pacsin2^umc405/+; pacsin2^umc406/+; mpv17^a9/a9 (AB)
mitfa^w2/w2; pacsin2^umc406/+; mpv17^a9/a9 (AB)

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Antibodies

Name	Type	Antigen Genes	Isotypes	Host Organism
Ab1-eea1	polyclonal		IgG	Goat
Ab1-lrp2a	polyclonal	lrp2a		Rabbit
Ab1-pacsin2	polyclonal	pacsin2		Sheep
Ab-3G8	monoclonal			Mouse
Ab3-gapdh	polyclonal		IgG	Rabbit
Ab5-rab11	monoclonal			Mouse

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Orthology

Engineered Foreign Genes

Mapping

Morgan et al., 2022 ZDB-PUB-220527-7 PMID:35616009

Pacsin2 is required for endocytosis in the zebrafish pronephric tubule

Morgan et al., 2022

ZDB-PUB-220527-7
PMID:35616009