Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

New Toolset of Reporters Reveals That Glycogen Granules Are Neutral Substrates of Bulk Autophagy in Komagataella phaffii

Version 1 : Received: 26 September 2024 / Approved: 27 September 2024 / Online: 27 September 2024 (15:30:52 CEST)

How to cite: Wijewantha, N. V.; Battu, P.; Chen, K.; Kumar, R.; Nazarko, T. Y. New Toolset of Reporters Reveals That Glycogen Granules Are Neutral Substrates of Bulk Autophagy in Komagataella phaffii. Preprints 2024, 2024092233. https://doi.org/10.20944/preprints202409.2233.v1 Wijewantha, N. V.; Battu, P.; Chen, K.; Kumar, R.; Nazarko, T. Y. New Toolset of Reporters Reveals That Glycogen Granules Are Neutral Substrates of Bulk Autophagy in Komagataella phaffii. Preprints 2024, 2024092233. https://doi.org/10.20944/preprints202409.2233.v1

Abstract

Glycogen, a branched polysaccharide organized into glycogen granules (GGs), is delivered from the cytoplasm to the lysosomes of hepatocytes by the STBD1-driven selective autophagy (glycophagy). Recently, we developed Komagataella phaffii yeast as a simple model of GG autophagy and found that it proceeds non-selectively under nitrogen starvation conditions. However, another group, using Saccharomyces cerevisiae as a model, found that glycogen is a non-preferred cargo of the nitrogen-starvation induced bulk autophagy. To clarify cargo characteristics of K. phaffii GGs, we used the same glycogen synthase-based reporter (Gsy1-GFP) of GG autophagy in K. phaffii, as was used in S. cerevisiae. The K. phaffii Gsy1-GFP marked GGs and reported on their autophagic degradation during nitrogen starvation, as expected. However, unlike in S. cerevisiae, glycogen synthase-marked GGs were delivered to the vacuole and degraded there with the same efficiency as a cytosolic glycogen synthase in glycogen-deficient cells suggesting that glycogen is a neutral cargo of bulk autophagy in K. phaffii. We verified our findings with a new set of reporters based on the glycogen-binding CBM20 domain of human STBD1. The GFP-CBM20 and mCherry-CBM20 fusion proteins tagged GGs, reported about autophagy of GGs and confirmed that GGs in K. phaffii are neither preferred, nor non-preferred substrates of bulk autophagy. They are its neutral substrates.

Keywords

autophagy; bulk autophagy; CBM20; glycogen; glycogen granules; glycophagy; Komagataella phaffii; Pichia pastoris; selective autophagy; yeast

Subject

Biology and Life Sciences, Cell and Developmental Biology

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