Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls.

Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Research Abstract Details 

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Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Abstract Text:

Isei Tanida,Toshiyuki Yamaji,Takashi Ueno,Shoichi Ishiura,Eiki Kominami,Kentaro Hanada,Isei Tanida,Toshiyuki Yamaji,Takashi Ueno,Shoichi Ishiura,Eiki Kominami,Kentaro Hanada,

A cytosolic form of LC3 is conjugated to phosphatidylethanolamine by Atg7, an E1-like enzyme, and Atg3, an E2-like enzyme, during autophagy. To monitor intracellular autophagosomes and autolysosomes, GFP-LC3 is a useful tool. However, GFP-LC3 can aggregate without being conjugated to phosphatidylethanolamine, especially when GFP-LC3 is transiently expressed (Kuma A, Matsui M, Mizushima N. Autophagy 2007; 3:323-8). Therefore, as a negative control, we investigated a mutant human LC3DeltaG protein in which the C-terminal Gly(120) essential for LC3-lipidation is deleted, and generated a set of expression plasmids for wild-type human LC3 and mutant LC3DeltaG fused to either CFP, GFP, YFP, or HcRed at the N terminus. We found that the mutant LC3DeltaG protein does not react with human Atg7 and Atg3, indicating that LC3-lipidation does not occur, and few puncta containing mutant LC3DeltaG form under starvation conditions. As observed with wildtype HcRed-LC3, mutant HcRed-LC3DeltaG also co-localizes with polyQ150-aggregates suggesting that the colocalization of HcRed- LC3 to polyQ150-aggregates is independent of LC3-lipidation. These mutant LC3DeltaG proteins will be useful negative controls in recognizing non-specific fluorescent protein-LC3 aggregates.

Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Publishing Authors By Initials

I Tanida,T Yamaji,T Ueno,S Ishiura,E Kominami,K Hanada,I Tanida,T Yamaji,T Ueno,S Ishiura,E Kominami,K Hanada,

For similar natural sciences: weights and measures: reference values research abstracts see: natural sciences: weights and measures: reference values research

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Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Autophagy

VOLUME: 4

Page Numbers: 131-4

Journal Abbreviation:

ISSN: 1554-8635

DAY: 1

MONTH: 11

YEAR: 2007

Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Information

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LANGUAGE: eng

NlmUniqueID: 101265188

Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Keywords Mesh Terms:

KEYWORDS: Reference Values

MESH TERMS: metabolism

Chemical & Substance for Abstract: Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls. Information

Substance Name: light chain 3, human

Registry Number: 0

Grant and Affiliation Information for Consideration about negative controls for LC3 and expression vectors for four colored fluorescent protein-LC3 negative controls.

AFFILIATION: Department of Biochemistry and Cell Biology, National Institute of Infectious Disease, Tokyo, Japan.

Country: United States

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MEDLINETA: Autophagy

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