PT  - JOURNAL ARTICLE
AU  - Tim Schulte
AU  - Marc D. Panas
AU  - Lucy Williams
AU  - Nancy Kedersha
AU  - Jonas Simon Fleck
AU  - Timothy J.C. Tan
AU  - Anders Olsson
AU  - Ainhoa Moliner Morro
AU  - Leo Hanke
AU  - Johan Nilvebrant
AU  - Kim Anh Giang
AU  - Per-Åke Nygren
AU  - Paul Anderson
AU  - Adnane Achour
AU  - Gerald M. McInerney
TI  - Caprin-1 binding to the critical stress granule protein G3BP1 is regulated by pH
AID  - 10.1101/2021.02.05.429362
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.02.05.429362
4099  - http://biorxiv.org/content/early/2021/02/05/2021.02.05.429362.short
4100  - http://biorxiv.org/content/early/2021/02/05/2021.02.05.429362.full
AB  - G3BP is the central hub within the protein-RNA interaction network of stress-induced bio-molecular condensates known as stress granules (SG). The SG-associated proteins Caprin-1 and USP10 exhibit mutually exclusive binding to the structured NTF2-domain of G3BP1, thereby regulating G3BP1-mediated condensation, but with opposite effects: Caprin-1 promotes but USP10 inhibits SG formation. Herein, we present the crystal structure of G3BP1-NTF2 in complex with a Caprin-1 derived short linear motif (SLiM), which provides a molecular understanding for the mutually exclusive binding of USP10 and Caprin-1 to G3BP1. Caprin-1 but not USP10 contacts two G3BP1-NTF2 histidine residues, which was confirmed using biochemical, biophysical and cellular biological binding assays. G3BP1/Caprin-1 interactions disrupted via point mutations resulted in fewer and smaller SG condensates. In addition, biochemical binding assays demonstrated reduced binding of Caprin-1 to G3BP1 at lower pH values. Finally, ratiometric pH sensitive measurements of SGs revealed a substantial drop in pH compared to the adjacent cytosol, suggesting that reduced pH can fine-tune and regulate the G3BP1-mediated interaction network via a NTF2-mediated pH-sensitive SLiM-selection mechanism.Competing Interest StatementThe authors have declared no competing interest.BLIBio-Layer InterferometryBMCsBio-molecular condensatesCaprin-1Cell cycle-associated protein 1co-IPCo-immunoprecipitationECDFEmpirical cumulative distribution functionFMR1/FMRPFragile X mental retardation proteinG3BP1Ras GTPase-activating protein-binding protein 1IPImmunoprecipitationITCIsothermal titration calorimetryLCR /IDRLow complexity or intrinsically disordered regionsnanoDSFNano differential scanning fluorimetryNTF2Nuclear transport factor 2 like domainRGGArginine-Glycine-Glycine motifSASodium arseniteSECSize exclusion chromatographySFVSemliki Forest virusSGStress granuleSLiMshort linear (peptide) motifUSP10Ubiquitin carboxyl-terminal hydrolase 10WTWild type