bims-unfpre 2025-08-17 papers

Issue of 2025–08–17
five papers selected by
Susan Logue, University of Manitoba

bioRxiv. 2025 Aug 07. pii: 2025.08.05.666879. [Epub ahead of print]

Secreted Protein Production is Improved by Controlling Endoplasmic Reticulum Stress Associated Protein Degradation.

Ryan Chauncey Splichal, Christina Chan, S Patrick Walton.

Therapeutic proteins are produced frequently by mammalian cells in large-scale bioreactors. As a result, producer cells are exposed to a chemically (nutrients, gas exchange, target protein overexpression) and physically (shear due to mixing) stressful environment, which can lead to loss of proteostasis and endoplasmic reticulum (ER) stress. In response, cells activate the unfolded protein response (UPR). The UPR includes activation of autophagy and proteasomes, both of which target unfolded/misfolded proteins for degradation. To investigate the impacts of autophagy and proteasome activity on secreted protein production in ER-stressed cells, we used HeLa and MDA-MB-231 cells transfected to express Gaussia luciferase (as a model for therapeutic protein production) and exposed to tunicamycin (TM) (to activate ER stress). As expected, TM exposure decreased protein production and secretion. Inhibiting autophagy improved secretion in stressed cells as expected. However, counterintuitively, increasing proteasomal degradation improved secretion while inhibiting proteasomal activity decreased secretion, that is proteasomal activity was directly correlated to secretion. Taken together, our results demonstrate that protein secretion can be improved through control of autophagy and proteasomal activity, providing insight into strategies for improving yield from protein production bioprocesses.

DOI: https://doi.org/10.1101/2025.08.05.666879

bioRxiv. 2025 Jul 14. pii: 2025.07.11.664427. [Epub ahead of print]

SCOPE: Revealing Hidden Mechanisms in Phenotypic Screens Through Target and Pathway Enrichment.

Abhijeet Kapoor, Keith Kelleher, Suzanne Underhill, Sankalp Jain, Brandon K Harvey, Mark J Henderson.

Phenotypic screening enables discovery of small molecules without requiring predefined targets, but mechanistic interpretation remains challenging due to polypharmacology and pathway complexity. We developed SCOPE (Screening Compound Ontology for Pathway Enrichment), a KNIME-based computational framework that resolves the molecular drivers of phenotypic activity by linking compound-level screening data to annotated targets and pathways. SCOPE integrates multi-source target annotations and performs statistical enrichment to identify shared mechanisms of action. Applied to a high-throughput screen for modulators of ER-stress induced secretion of endoplasmic reticulum (ER) resident proteins, a process known as exodosis, SCOPE identified calcium signaling as the most enriched KEGG pathway without prior biological context. Target enrichment revealed G protein-coupled receptors (GPCRs) involved in inositol 1,4,5-trisphosphate receptors (IP3Rs)-mediated signaling, with widespread antagonism among hit compounds implicating this pathway in the regulation of exodosis. Notably, SCOPE uncovered a novel role for the histamine receptor HRH1, which was validated by RNAi knockdown and pharmacological inhibition, implicating HRH1 as a potential therapeutic target in ER stress-related disorders. These results highlight SCOPE's potential to deconvolute phenotypic screens and uncover actionable mechanisms in complex cellular systems.

DOI: https://doi.org/10.1101/2025.07.11.664427

Mol Biol Cell. 2025 Aug 13. mbcE25010024

Endosome maturation during ER stress relies on the ubiquitin binding domain of Histone Deacetylase 6.

Katherine M Piscopo, Brooke Larson, Anna M Christiansen, Jason M Perry, Julie Hollien.

Histone deacetylase 6 (HDAC6) helps cells manage misfolded proteins by transporting ubiquitin-associated structures toward the microtubule organizing center, where they can be sequestered and degraded by lysosomes. Here we show that when cells are subjected to acute protein folding stress in the endoplasmic reticulum (ER), HDAC6 depletion results in the appearance of enlarged endosomes that are highly decorated with ubiquitin and colocalize with both early and late endosome markers. The C-terminal ubiquitin-binding domain and adjacent disordered regions of HDAC6 are necessary and sufficient to rescue this endosomal phenotype in cells lacking endogenous HDAC6. HDAC6 deficiency does not appear to prevent the recruitment of endosomal sorting complexes required for transport (ESCRTs), which coordinate endosome maturation. However, overexpression of HDAC6 can reverse endosome phenotypes associated with the depletion of the early ESCRT factor HRS. We speculate that HDAC6 facilitates the packaging and processing of endosomal cargo when the endomembrane system is under stress.

DOI: https://doi.org/10.1091/mbc.E25-01-0024

Nat Cell Biol. 2025 Aug 11.

A multichaperone condensate enhances protein folding in the endoplasmic reticulum.

Anna Leder, Guillaume Mas, Viktória Szentgyörgyi, Roman P Jakob, Timm Maier, Anne Spang, Sebastian Hiller.

Protein folding in the endoplasmic reticulum (ER) relies on a network of molecular chaperones that facilitates the folding and maturation of client proteins. How the ER chaperones organize in a supramolecular manner to exert their cooperativity has, however, remained unclear. Here we report the discovery of a multichaperone condensate in the ER lumen, which is formed around the chaperone PDIA6 during protein folding homeostasis. The condensates form in a Ca2+-dependent manner and we resolve the underlying mechanism at the atomic and cellular levels. The PDIA6 condensates recruit further chaperones-Hsp70 BiP, J-domain protein ERdj3, disulfide isomerase PDIA1 and Hsp90 Grp94-which constitute some of the essential components of the early folding machinery. The chaperone condensates enhance folding of proteins, such as proinsulin, and prevent protein misfolding in the ER lumen. The PDIA6-scaffolded chaperone condensates hence provide the functional basis for spatial and temporal coordination of the dynamic ER chaperone network.

DOI: https://doi.org/10.1038/s41556-025-01730-w

Nat Commun. 2025 Aug 13. 16(1): 7523

Fbxo42 promotes the degradation of Ataxin-2 granules to trigger terminal Xbp1 signaling.

Cristiana C Santos, Nadine Schweizer, Fátima Cairrão, Juanma Ramirez, Nerea Osinalde, Ming Yang, Catarina J Gaspar, Vanya I Rasheva, Miguel L Trigo, Zach Hensel, Colin Adrain, Tiago N Cordeiro, Franka Voigt, Paulo A Gameiro, Ugo Mayor, Pedro M Domingos.

The Unfolded Protein Response (UPR) is activated by the accumulation of misfolded proteins in the Endoplasmic Reticulum (ER), a condition known as ER stress. Prolonged ER stress and UPR activation cause cell death, by mechanisms that remain poorly understood. Here, we report that regulation of Ataxin-2 by Fbxo42 is a crucial step during UPR-induced cell death. From a genetic screen in Drosophila, we identify loss of function mutations in Fbxo42 that suppress cell death and retinal degeneration induced by the overexpression of Xbp1spliced, an important mediator of the UPR. We identify the RNA binding protein Ataxin-2 as a substrate of Fbxo42, which, as part of a Skp-A/Cullin-1 complex, promotes the ubiquitylation and degradation of Ataxin-2. Upon ER-stress, the mRNA of Xbp1 is sequestered and stabilized in Ataxin-2 granules, where it remains untranslated. Fbxo42 recruitment to these granules promotes the degradation of Ataxin-2, allowing for the translation of Xbp1 mRNA and triggering cell death during the terminal stages of UPR activation.

DOI: https://doi.org/10.1038/s41467-025-62417-2