Endogenous retrovirus-like proteins recruit UBQLN2 to stress granules and shape their functional biology

Endogenous retrovirus-like proteins recruit UBQLN2 to stress granules and shape their functional biology Journal: Science Manuscript: adu6354 Milaganur Mohan, Harihar1; Fernandez, Martin2; Huang, Camellia1; Lin, Rita1; Ryou, Jaimie1; Seyfried, Donald1; Grotewold, Nikolas1; Basrur, Barmada, Sami1; Venkatesha1; Mosalaganti, Shyamal1; Whiteley, Alexandra3; Paulson, Henry1; Sharkey, Lisa1 Author affiliations 1 University of Michigan Medical School 2 University of Michigan 3 University of Colorado Boulder Abstract The human genome is replete with sequences derived from foreign elements including endogenous retrovirus-like proteins of unknown function. Here we show that UBQLN2, a ubiquitin-proteasome shuttle factor implicated in neurodegenerative diseases, is regulated by the linked actions of two retrovirus-like proteins, RTL8 and PEG10. RTL8 confers on UBQLN2 the ability to complex with and regulate PEG10. PEG10, a core component of stress granules, drives the recruitment of UBQLN2 to stress granules under various stress conditions, but can only do so when RTL8 is present. Changes in UBQLN2, RTL8 or PEG10 levels further remodel the kinetics of stress granule disassembly and translation recovery. PEG10 also alters overall stress granule composition by incorporating select extracellular vesicle proteins. Within stress granules, PEG10 forms virus-like particles, underscoring the structural heterogeneity of this class of biomolecular condensates. Together, these results reveal an unexpected link between pathways of cellular proteostasis and endogenous retrovirus-like proteins. Subjects Biological sciences, Molecular neuroscience, Retrotransposons, Retroviruses, Protein aggregation, Ubiquitin-proteasome system Funding Office of Environmental Management, P30AG072931 National Institute of Health, T32GM141840 National Institute of Health, T32GM007863 National Institute of Health, R01NS097542 National Institute of Health, R01NS113943 National Institute of Health, 1R56NS128110-01 National Institute of Health, DP2GM150019-01 National Institute of Health, R35NS122302 National Institute of Health, NIH S10OD030275 Description of the data and file structure The human genome is replete with sequences derived from foreign elements including endogenous retrovirus-like proteins of unknown function. Here we show that UBQLN2, a ubiquitin-proteasome shuttle factor implicated in neurodegenerative diseases, is regulated by the linked actions of two retrovirus-like proteins, RTL8 and PEG10. RTL8 confers on UBQLN2 the ability to complex with and regulate PEG10. PEG10, a core component of stress granules, drives the recruitment of UBQLN2 to stress granules under various stress conditions, but can only do so when RTL8 is present. Changes in UBQLN2, RTL8 or PEG10 levels further remodel the kinetics of stress granule disassembly and translation recovery. PEG10 also alters overall stress granule composition by incorporating select extracellular vesicle proteins. Within stress granules, PEG10 forms virus-like particles, underscoring the structural heterogeneity of this class of biomolecular condensates. Together, these results reveal an unexpected link between pathways of cellular proteostasis and endogenous retrovirus-like proteins. Files and variables File: Dryad_submission.zip Description: Data are grouped in appropriately named folders corresponding to each figure. Brief descriptions of the contents of each folder are as follows: Figure 1: Files for volcano plot generation (from the Excel file titled “PRF_F_2022_H_PAUL_836_54856_54870_1_12_RF1_RF2_Only_Unique_Pro”, columns C (gene name), N (abundance ratio), and Q (p-value) were used in the R code), MEGA11 files containing alignment of RTL proteins for phylogenetic tree construction, representative images in .jpg or .tif format and quantification of Western blots measuring PEG10 levels in HeLa, HEK293, and SK-N-SH cells treated with siRTL8 and siPEG10, representative images and quantification of Western blots measuring PEG10 levels in RTL8 KO HeLa cells rescued with HA-RTL8C, HA-RTL8C DN, Myc-LDOC1 or FLAG-UBQLN2, and representative images and quantification of Western blots measuring PEG10 levels in WT, non-targeting control, and RTL8 KO HEK293 cells. Figure 2: Representative images in .jpg or .tif format and quantification of Western blots showing UBQLN2-PEG10 co-IP after UBQLN2 IP with and without crosslinking, representative images and quantification of Western blots showing UBQLN2-PEG10 co-IP after UBQLN2 IP in RTL8 KO HeLa cells rescued with HA-RTL8C full-length or DN, or PEG10 KO HeLa cells rescued with V5-PEG10 gag-pol full-length or DPPR, and representative PLA images in lif and jpg format, contrast and brightness settings file for LAS X, CellProfiler and Prism analyses files for UBQLN2-PEG10 PLA experiments on non-targeting control, RTL8 KO, PEG10 KO, and UBQLN2 KO HeLa cells. Figure 3: Representative Western blot images in .jpg or .tif format of biotinylated isoxazole...