dictyNews
Electronic Edition
Volume 41, number 26
December 12, 2015

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Abstracts
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The two Dictyostelium autophagy eight proteins, ATG8a and ATG8b, 
associate with the autophagosome in succession

Jan Matthias, Susanne Meßling and Ludwig Eichinger


European Journal of Cell Biology, in press.

Autophagy is an ancient cellular pathway that is conserved from 
yeast to man. It contributes to many physiological and pathological 
processes and plays a major role in the degradation of proteins 
and/or organelles in response to starvation and stress. In the 
autophagic process cytosolic material is captured into double 
membrane-bound vesicles, the autophagosomes. After fusion with 
lysosomes, the cargo is degraded in the generated autolysosomes and 
then recycled for further use.Autophagy 8 (ATG8, in mammals LC3), a 
well-established marker of autophagy, is covalently linked to 
phosphatidylethanolamine on the autophagic membrane during 
autophagosome formation. Bioinformatic analysis of the Dictyostelium 
genome revealed two atg8 genes which encode the ATG8a and ATG8b 
paralogs, which are with around 14 kDa similar in size, 54 % identical 
to one another and more closely related to the corresponding proteins 
in fungi and plants than in animals. For ATG8a we found a strong up-
regulation throughout the 24 h developmental time course while ATG8b 
expression was highest in vegetative cells followed by a moderate 
reduction during early development. Confocal microscopy of 
fluorescently tagged ATG8a and ATG8b in vegetative AX2 wild-type and 
in ATG9  minus cells showed that both proteins mainly co-localised on 
vesicular structures with a diameter above 500 nm while those smaller 
than 500 nm were predominantly positive for ATG8b. In ATG9 minus cells 
we found a strong increase in the relative abundance of ATG8a-positive 
large vesicular structures and of total ATG8b-positive structures per 
cell indicating autophagic flux problems in this mutant. We also found 
that vesicular structures positive for ATG8a and/or ATG8b were also 
positive for ubiquitin. Live cell imaging of AX2 and ATG9 minus cells 
co-expressing combinations of red and green tagged ATG8a, ATG8b or 
ATG9 revealed transient co-localisations of these proteins. Our results 
suggest that ATG8b associates with nascent autophagosomes before 
ATG8a. We further find that the process of autophagosome formation in 
Dictyostelium is highly dynamic. We infer from our data that 
Dictyostelium ATG8a and ATG8b have distinct functions in autophagosome 
formation and that ATG8b is the functional ortholog of the mammalian 
LC3 subfamily and ATG8a of the GABARAP subfamily.


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