PUBLICATION

The Rag-Ragulator Complex Regulates Lysosome Function and Phagocytic Flux in Microglia

Authors

Shen, K., Sidik, H., Talbot, W.S.

ID

ZDB-PUB-160118-4

Date

2016

Source

Cell Reports 14(3): 547-59 (Journal)

Registered Authors

Talbot, William S.

Keywords

none

MeSH Terms

Monomeric GTP-Binding Proteins/metabolism*
Zebrafish/growth & development
Zebrafish/metabolism
Phenotype
Mutagenesis
Lysosomes/metabolism*
Phagocytosis
In Situ Hybridization
Guanine Nucleotide Exchange Factors/metabolism*
Signal Transduction
TOR Serine-Threonine Kinases/genetics
TOR Serine-Threonine Kinases/metabolism
DNA Mutational Analysis
Microscopy, Confocal
Zebrafish Proteins/metabolism*
Animals
Embryo, Nonmammalian/metabolism
Microglia/metabolism*

(all 18)

PubMed

26774477 Full text @ Cell Rep.

Abstract

Microglia are resident macrophages of the CNS that are essential for phagocytosis of apoptotic neurons and weak synapses during development. We show that RagA and Lamtor4, two components of the Rag-Ragulator complex, are essential regulators of lysosomes in microglia. In zebrafish lacking RagA function, microglia exhibit an expanded lysosomal compartment, but they are unable to properly digest apoptotic neuronal debris. Previous biochemical studies have placed the Rag-Ragulator complex upstream of mTORC1 activation in response to cellular nutrient availability. Nonetheless, RagA and mTOR mutant zebrafish have distinct phenotypes, indicating that the Rag-Ragulator complex has functions independent of mTOR signaling. Our analysis reveals an essential role of the Rag-Ragulator complex in proper lysosome function and phagocytic flux in microglia.

Genes / Markers

Figures

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Expression

Gene	Fish	Conditions	Stage	Anatomy	Assay	Figure
apoeb	rraga^st77/st77	standard conditions	Protruding-mouth	brain microglial cell	ISH	Fig. 2
apoeb	WT	standard conditions	Protruding-mouth	brain microglial cell	ISH	Fig. 2
arhgdia	WT	chemical treatment by environment: torin 1	Day 5	whole organism	RTPCR	Fig. 6
asah1b	rraga^st77/st77	standard conditions	Day 5	whole organism	RTPCR	Fig. 6
asah1b	WT	chemical treatment by environment: torin 1	Day 5	whole organism	RTPCR	Fig. 6
bloc1s6	rraga^st77/st77	standard conditions	Day 5	whole organism	RTPCR	Fig. 6
ctsc	rraga^st77/st77	standard conditions	Day 5	whole organism	RTPCR	Fig. 6
ctsc	WT	chemical treatment by environment: torin 1	Day 5	whole organism	RTPCR	Fig. 6
DsRed	gl22Tg; zf148Tg	standard conditions	Day 6	brain neuron	IFL	Fig. 5
DsRed	gl22Tg; zf148Tg	standard conditions	Day 4	brain neuron	IFL	Fig. 5

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
WT	chemical treatment by environment: torin 1	Day 5	brain has normal numbers of parts of type microglial cell, normal	Fig. 6
WT	chemical treatment by environment: torin 1	Day 5	whole organism arhgdia expression decreased amount, abnormal	Fig. 6
WT	chemical treatment by environment: torin 1	Day 5	whole organism asah1b expression decreased amount, abnormal	Fig. 6
WT	chemical treatment by environment: torin 1	Day 5	whole organism ctsc expression decreased amount, abnormal	Fig. 6
WT	chemical treatment by environment: torin 1	Day 5	whole organism gabarapa expression decreased amount, abnormal	Fig. 6
WT	chemical treatment by environment: torin 1	Day 5	whole organism mapk3 expression decreased amount, abnormal	Fig. 6
WT	chemical treatment by environment: torin 1	Day 5	whole organism ab9-rps6 labeling decreased amount, abnormal	Fig. 1
irf8^st95/st95	standard conditions	Day 4	brain lacks all parts of type microglial cell, abnormal	Fig. 4
irf8^st95/st95	standard conditions	Day 4	optic tectum apoptotic cell clearance decreased occurrence, abnormal	Fig. 4
lamtor4^st99/st99	standard conditions	Protruding-mouth	brain has fewer parts of type microglial cell, abnormal	Fig. 1

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
gl22Tg	Tg(mpeg1:EGFP)	Transgenic Insertion
st74		Point Mutation	lamtor4
st77		Point Mutation	rraga
st95		Small Deletion	irf8
st99		Small Deletion	lamtor4
st110		Small Deletion	rraga
xu015Gt	Gt(GBT-P9)	Transgenic Insertion	mtor
zf148Tg	Tg(Xla.Tubb:DsRed)	Transgenic Insertion

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Human Disease / Model

Sequence Targeting Reagents

Fish

Fish
gl22Tg
gl22Tg; zf148Tg
irf8^st95/st95
lamtor4^st99/st99
mtor^{xu015Gt/xu015Gt}
rraga^st77/st77
rraga^st77/st77; gl22Tg
rraga^st77/st77; gl22Tg; zf148Tg
rraga^st110/st110
WT

Antibodies

Name	Type	Antigen Genes	Isotypes	Host Organism
Ab1-rraga	monoclonal	rraga	IgG	Rabbit
Ab2-map1lc3b	polyclonal			Rabbit
Ab6-sqstm1	polyclonal			Rabbit
Ab9-rps6	monoclonal		IgG	Rabbit

Orthology

Engineered Foreign Genes

Marker	Marker Type	Name
DsRed	EFG	DsRed
EGFP	EFG	EGFP
GFP	EFG	GFP

Mapping

Shen et al., 2016 ZDB-PUB-160118-4 PMID:26774477

The Rag-Ragulator Complex Regulates Lysosome Function and Phagocytic Flux in Microglia

Shen et al., 2016

ZDB-PUB-160118-4
PMID:26774477