Domain-specific regulation of PABPN1 uncouples aggregation from function and enables therapeutic rescue

PABPN1 is a multifunctional protein whose expression is tightly regulated to maintain cellular homeostasis. PABPN1 dysregulation contributes to both common acquired diseases, such as bladder cancer, and rare inherited disorders, including oculopharyngeal muscular dystrophy (OPMD). In OPMD, PABPN1 forms insoluble aggregates that reduce its functional levels, leading to genome-wide shifts in alternative polyadenylation (APA) at 3'-UTRs and disruption of mRNA metabolism, including nuclear export and translation. OPMD is caused by a short alanine expansion at the N-terminus of PABPN1 within an intrinsically disordered region (IDR) followed by a coiled-coil domain (CCD). How these domains contribute to PABPN1 function and aggregation remains unclear. Here, we show that the N-terminal IDR promotes aggregation and modulates protein-protein interactions with longer IDRs suppressing interaction between PABPN1 and its binding partners. We further demonstrate that the CCD has dual functions: its N-terminal domain dictates PABPN1 stability, whereas its C-terminal domain enhances the PABPN1 interactome. We identified a naturally occurring variant lacking exon 1 encoding the IDR and the CCD N-terminal domain (named trPAB). That variant forms a stable, non-aggregating protein isoform. Interactome analysis and structural modeling indicate improved molecular function by restoring PABPN1 activity, APA profiles, and cellular phenotypes in OPMD and bladder cancer models. Importantly, delivery of trPAB via Adeno-associated viral vector in an OPMD mouse model improves muscle histopathology, supporting its potential as a therapeutic strategy. A mechanistic basis of a novel gene therapy approach for OPMD O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/731084v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@121f4b7org.highwire.dtl.DTLVardef@4942f7org.highwire.dtl.DTLVardef@1cb8613org.highwire.dtl.DTLVardef@cf6810_HPS_FORMAT_FIGEXP M_FIG C_FIG Top panel: A schematic presentation of three PABPN1 variant functional states. Left: the normal full-length protein interacts with key RNA binding protein (RBP) partners supporting normal cellular function. Middle: A pathogenic full-length alanine-expanded PABPN1 leads to protein aggregation, resulting in limited interactors, dysfunctional complexes and impaired cellular processes. Right: a truncated natural variant (trPAB), not associated with OPMD pathology, shows enhanced stability and reduced aggregation, suggesting a potential protective or "super functional" profile. The bottom panel translates these mechanistic insights into therapeutic strategy. In cell models, expression of trPAB is associated with reduced protein aggregation, restoration of normal cellular function, and improved myogenesis. These effects are recapitulated in a relevant animal model of OPMD, where trPAB treatment leads to decreased aggregation, functional recovery, and reversal of muscle atrophy. Our study supports the rationale for leveraging a stabilized PABPN1 variant as a novel gene therapy approach to counteract aggregation-driven pathology in OPMD. HighlightsO_LIThe central coiled-coil domain (CCD) of PABPN1 has two opposing functions: its N-terminal region stabilizes the protein, while its C-terminal region promotes stability but hampers aggregation. C_LIO_LIAt the N-terminus, an intrinsically disordered region (IDR) acts as a gatekeeper for protein interactions. The alanine tract, including its pathogenic expansion, restricts this interactome, limiting PABPN1 binding capacity. C_LIO_LIWe identified a naturally occurring PABPN1 isoform, trPAB, which lacks exon 1, including the IDR and the N-terminal portion of the CCD. trPAB forms a stable and functional protein and exerts beneficial effects in muscle cells. C_LIO_LIFunctionally, trPAB rescues key cellular phenotypes in bladder cancer cells and reverses nuclear aggregation and muscle atrophy in a mouse model of OPMD. C_LI