Suppression of aggregate and amyloid formation by a novel intrinsically disordered region in metazoan Hsp110 chaperones
Preprint posted on 13 January 2021 https://www.biorxiv.org/content/10.1101/2021.01.13.426581v1
Written by: Anna Enneking, Simon Hinfelaar, Inge Rothuis and Vincent Spit
Protein misfolding can cause devastating problems in the cell, as exemplified by the formation of amyloid fibrils in neurodegenerative diseases . Chaperones, including the ATP-dependent Hsp70, prevent protein aggregation . Hsp70 needs a nucleotide exchange factor (NEF) such as Hsp110 to promote cycling from ADP to ATP . Interestingly, Hsp110 is part of the Hsp70 superfamily, meaning they share the canonical Hsp70 architecture. They both have a nucleotide binding domain (NBD) that binds ATP, a flexible linker, and a substrate binding domain (SBD) composed of two parts: a β-sandwich-containing substrate binding site (SBD-β) and α-helical sequence (SBD-α).
Hsp110 also has an extended C-terminus, which is conserved in metazoan Hsp110 genes, e.g human Hsp105α and Apg-1, and has an unknown function . Although they share a canonical structure, Hsp110 has no refolding activity as Hsp70 does. However, Hsp110 overexpression can prevent aggregation of the huntingtin protein in a Drosophila model system . It is unknown whether this is a function of Hsp110 as a NEF or as a passive holdase. To understand this, the authors decide to disrupt the Hsp110 holdase function by targeting the SBD-β domain, because they suspect that this domain is responsible for the holdase activity. In their attempts to disrupt the SBD-β, they uncover a secondary binding site.
The authors introduce different deletions in the SBD-β domain of Drosophila Hsp110, leaving the regions that interact with Hsp70 (NBD, SBD-α) intact. They monitor protein aggregation by a light scattering aggregation assay using citrate synthase as substrate to test the holdase activity. Surprisingly, the Hsp110 protein in which SBD-β is deleted is more efficient in preventing aggregation than the full-length protein. So, if the SBD-β is not essential for the activity in this assay, then what else is?
The authors now suspect intrinsically disordered regions (IDRs), as such regions can act as sites for protein binding and are present in many chaperones . In silico analysis reveals one IDR in the C-terminus and another one in the SBD-α domain. It turns out that it is the C-terminal IDR that is responsible for the holdase activity, because when deleted the holdase activity is lost. Deletion of the SBD-α domain does not affect the holdase activity. The human homologs of Hsp110, Apg1a and Hsp105α, also have an IDR in the C-terminus. These C-termini also prevent protein aggregation when directly linked to the NBD (Fig. 4C), so the presence of an IDR in the C-terminus could indicate a general holdase function in the Hsp110 proteins.
The authors now study whether the IDRs can also prevent amyloid formation, which is associated with various diseases including Alzheimer ‘s . To this end, they examine the impact of variants of Drosophila Hsp110 (dHsp110) and its human homolog Hsp105α on fibril formation of amyloid-β 42 (Aβ42). Negative staining TEM shows that NBDdHsp110 alone does not decrease Aβ42 fibril formation, but the IDR fusions NBDdHsp110-IDRdHsp110 and NBDdHsp110-IDRHsp105α do (Fig. 5C). Thus, the IDR in Hsp110 homologs can also prevent amyloid formation.
What we like about the preprint
We were directly drawn to this article when we read the title. We found the fact that Hsp110 can prevent protein aggregation very interesting, mainly because we previously learned that Hsp110 works as NEF of Hsp70. It was therefore intriguing to read that Hsp70 and Hsp110 share a canonical structure, but that the SBD-β in Hsp110 is not solely responsible for its holdase activity. It was surprising to learn that an IDR contributes to this activity. To our excitement, this paper shows that the IDR in Hsp110 can actually prevent amyloid formation, which is associated with neurodegenerative diseases. This knowledge is important to better understand how chaperones prevent this pathogenic process in the cell.
Posted on: 14 April 2021 , updated on: 21 April 2021
doi: PendingRead preprint
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