Invited Speaker Oral Presentation Inaugural Australian Ubiquitin Summit 2025

The accumulation and precipitation of unbranched glycogen in human tissues as toxic polyglucosan bodies (PB) underlies a number of human diseases that cause cardiomyopathy and heart failure, liver failure and/or fatal neurological disorders. They result from mutations in the genes encoding HOIL-1 (also called RBCK1), malin, laforin, glycogen branching enzyme (GBE1) or glycogenin-1 (GYG1). HOIL-1 and malin are both E3 ubiquitin ligases. We have reported that HOIL-1, a component of the linear ubiquitin assembly complex (LUBAC) ubiquitylates hydroxyl groups in proteins forming ester bonds (1-3) and the 6-hydroxyl group of glucose in small unbranched glucosaccharides (4). The following lines of evidence let us to propose that unbranched (or sparsely branched) glycogen molecules, formed by errors of metabolism, undergo ubiquitylation, marking them for uptake into lysosomes and destruction by the lysosomal acid alpha-1:4 glucosidase (GAA). The failure of this process leads to the accumulation of unbranched glycogen and its precipitation as PB in cells. This hypothesis was based on the following findings:-

1. PB are formed in the heart and brain of mice in which HOIL-1 is replaced by an E3 ligase-inactive mutant, establishing that the E3 ligase function of HOIL-1 is essential to prevent PB formation in vivo (4)
2. HOIL-1 only ubiquitylates the C6 hydroxyl groups of glucose molecules in unbranched glucosaccharides in vitro, the same hydroxyl group at which the branch points in glycogen are formed (4).
3. The HOIP component of LUBAC binds specifically to unbranched glycogen and generates Met1-linked ubiquitin oligomers that accelerate the HOIL-1-catalysed ubiquitylation of unbranched glycogen >100-fold, by an allosteric mechanism (4).

In the talk, I will present more recent results, which demonstrate that unbranched glycogen is polyubiquitylated in human cells, but normally branched glycogen is not. I will also present the identification of the inter-ubiquitin linkage types within the polyubiquitin chains attached to unbranched glycogen, which indicate that a minimum of three E3 ubiquitin ligases participate in the polyubiquitylation of unbranched glycogen in human cells. These results support the hypothesis that the polyubiquitylation of unbranched glycogen tinitiatesa quality control process that marks these molecules for destruction by glycophagy, a type of autophagy.

References

1. Kelsall, I.R., Zhang, J., Knebel, A., Arthur, J.S.C. and Cohen, P. (2019) ‘The E3 ligase HOIL-1 catalyses ester bond formation between ubiquitin and components of the myddosome in mammalian cells.” Proc. Nat. Acad. Sci. USA 116, 13293-13298
2. Petrova, T., Zhang, J., Nanda, S., Figueras-Vadillo, C. and Cohen, P. (2021)

“HOIL-1-catalysed, ester-linked ubiquitylation restricts IL-18 signaling in cytotoxic T cells but promotes TLR signalling in macrophages.” FEBS J. 288, 5909-5924

1. McCrory, E., Akimov^, V., Cohen, P.^* and Blagoev, B.* (2022) “ Identification of ester-linked ubiquitylation sites during TLR7 signalling increases the number of inter-ubiquitin linkages from 8 to 12” Biochem. J. 479, 2419-2431
2. Kelsall, I.R., McCrory, E.H., Xu, Y., Scudamore^, C.L., Nanda S.K., Mancebo-Gamella, P., Wood^, N.T., Knebel, A., Matthews, S.J. and Cohen, P. (2022) “HOIL-1 ubiquitin ligase activity targets unbranched glucosaccharides and is required to prevent polyglucosan accumulation” EMBO Journal 41:e109700