dictyNews

Electronic Edition

Volume 49, number 15

June 2, 2023



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Abstracts

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Recruitment of both the ESCRT and autophagic machineries to 

ejecting Mycobacterium marinum





Lilli Gerstenmaier1, Ombretta Colasanti1, Hannah Behrens1, 

Margot Kolonko1, Christian Hammann2,3, Monica Hagedorn1,2,3



1Bernhard Nocht Institute for Tropical Medicine, 20359 Hamburg, 

Germany.

2Ribogenetics Biochemistry Lab, Department of Life Sciences and 

Chemistry, Jacobs University Bremen gGmbH, Campus Ring 1, 

28759 Bremen, Germany.

3Health and Medical University, Campus Potsdam, Olympischer 

Weg 1, 14471 Potsdam, Germany.





Current Protocols, Mol. Micrcrobiology 2023 May 25

doi: 10.1111/mmi.15075  



Cytosolic Mycobacterium marinum are ejected from host cells such 

as macrophages or the amoeba Dictyostelium discoideum in a 

non-lytic fashion. As described previously, the autophagic machinery 

is recruited to ejecting bacteria and supports host cell integrity during 

egress. Here, we show that the ESCRT machinery is also recruited to 

ejecting bacteria, partially dependent on an intact autophagic pathway. 

As such, the AAA-ATPase Vps4 shows a distinct localization at the 

ejectosome structure in comparison to fluorescently tagged Vps32, 

Tsg101 and Alix. Along the bacterium engaged in ejection, ESCRT 

and the autophagic component Atg8 show partial colocalization. We 

hypothesize that both, the ESCRT and autophagic machinery localize 

to the bacterium as part of a membrane damage response, as well as 

part of a “frustrated autophagosome” that is unable to engulf the 

ejecting bacterium.





Submitted by Monica Hagedorn 

[[log in to unmask]]

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Proteasomes of autophagy-deficient cells exhibit alterations in 

regulatory proteins and a marked reduction in activity



Qiuhong Xiong, Rong Feng, Sarah Fischer, Malte Karow, Maria 

Stumpf, Susanne Meßling, Leonie Nitz, Stefan Müller, Christoph S. 

Clemen , Ning Song, Ping Li, Changxin Wu, Ludwig Eichinger





Cells, accepted



Autophagy and the ubiquitin proteasome system are the two major 

processes for the clearance and recycling of proteins and organelles 

in eukaryotic cells. Evidence is accumulating that there is extensive 

cross-talk between the two pathways, but the underlying mechanisms 

are still unclear. We previously found, that autophagy (ATG9) and 16 

(ATG16) proteins are crucial for full proteasomal activity in the 

unicellular amoeba Dictyostelium discoideum. In comparison to AX2 

wild-type cells, ATG9¯ and ATG16¯ cells, displayed a 60% and 

ATG9¯/16¯ cells a 90% decrease in proteasomal activity. Mutant cells 

also showed a significant increase in poly-ubiquitinated proteins and 

contained large ubiquitin-positive protein aggregates. Here, we 

focused on possible reasons for these results. Re-analysis of 

published tandem mass tag-based quantitative proteomic results of 

AX2, ATG9¯, ATG16¯, and ATG9¯/16¯ cells revealed no change in the 

abundance of proteasomal subunits. To identify possible differences 

in proteasome-associated proteins, we generated AX2 wild-type and 

ATG16¯ cells expressing the 20S proteasomal subunit PSMA4 as 

GFP-tagged fusion protein, and performed co-immunoprecipitation 

experiments followed by mass spectrometric analysis. The results 

revealed no significant differences in the abundance of proteasomes 

between the two strains. However, we found enrichment as well as 

depletion of proteasomal regulators and differences in the ubiquitination 

of associated proteins for ATG16¯ as compared to AX2 cells. Recently, 

proteaphagy has been described as a mean to replace non-functional 

proteasomes. We propose that autophagy-deficient D. discoideum 

mutants suffer from inefficient proteaphagy, which results in the 

accumulation of modified less active and also of inactive proteasomes. 

As a consequence, these cells exhibit a dramatic decrease in 

proteasomal activity and deranged protein homeostasis.





Submitted by Ludwig Eichinger [[log in to unmask]]

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[End dictyNews, volume 49, number 15]