dictyNews

Electronic Edition

Volume 43, number 9

May 5, 2017



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Abstracts

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The two Dictyostelium discoideum autophagy 8 proteins have distinct 

autophagic functions



Susanne Meßling, Jan Matthias, Qiuhong Xiong, Sarah Fischer and 

Ludwig Eichinger



Center for Biochemistry, Institute of Biochemistry I, Medical Faculty, 

University of Cologne, Joseph-Stelzmann-Str. 52, 50931 Cologne, Germany





European Journal of Cell Biology, in press.



Autophagy is a highly conserved cellular degradation pathway which is

crucial for various cellular processes. The autophagic process is 

subdivided in the initiation, autophagosome maturation and lysosomal 

degradation phases and involves more than forty core and accessory 

autophagy-related (ATG) proteins. Autophagy 8 (ATG8, in mammals LC3) 

is a well-established marker of autophagy and is linked to the autophagic 

membrane from initiation until fusion with the lysosome.



We generated single and double knock-out mutants of the two Dictyostelium 

paralogues, ATG8a and ATG8b, as well as strains that expressed 

RFP-ATG8a and/or GFP-ATG8b, RFP-ATG8b, RFP-GFP-ATG8a or 

RFP-GFP-ATG8b in different knock-out mutants. The ATG8b¯ mutant 

displayed only subtle phenotypic changes in comparison to AX2 wild-type 

cells. In contrast, deletion of ATG8a resulted in a complex phenotype with 

delayed development, reduced growth, phagocytosis and cell viability, an 

increase in ubiquitinylated proteins and a concomitant decrease in 

proteasomal activity. The phenotype of the ATG8a¯/b¯ strain was, except for

cell viability, in all aforementioned aspects more severe, showing that both 

proteins function in parallel during most analysed cellular processes. 

Immunofluorescence analysis of knock-out strains expressing either 

RFP-GFP-ATG8a or RFP-GFP-ATG8b suggests a crucial function for 

ATG8b in autophagosome-lysosome fusion. Quantitative analysis of strains 

expressing RFP-ATG8a, RFP-ATG8b, or RFP-ATG8a and GFP-ATG8b 

revealed that ATG8b generally localised to small and large vesicles, 

whereas ATG8a preferentially co-localised with ATG8b on large vesicles, 

indicating that ATG8b associated with nascent autophagosomes before 

ATG8a, which is supported by previous results (Matthias et al., 2016). 

Deconvoluted confocal fluorescence images showed that ATG8b localised 

around ATG8a and was presumably mainly present on the outer membrane 

of the autophagosome while ATG8a appears to be mainly associated with

the inner membrane.



In summary, our data show that ATG8a and ATG8b have distinct functions 

and are involved in canonical as well as non-canonical  autophagy. The 

data further suggest that ATG8b predominantly acts as adapter for the 

autophagy machinery at the outer and ATG8a as cargo receptor at the inner 

membrane of the autophagosome.





submitted by: Ludwig Eichinger [[log in to unmask]]

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[End dictyNews, volume 43, number 9]