Nuclear inclusions in oculopharyngeal muscular dystrophy consist of poly(A) binding protein 2 aggregates which sequester poly(A) RNA

Abstract

Oculopharyngeal muscular dystrophy (OPMD) is an adult-onset disease characterized by progressive eyelid drooping, swallowing difficulties and proximal limb weakness. The autosomal dominant form of the disease is caused by short (GCG)(8-13) expansions in the PABP2 gene. This gene encodes the poly(A) binding protein 2 (PABP2), an abundant nuclear protein that binds with high affinity to nascent poly(A) tails, stimulating their extension and controlling their length. In this work we report that PABP2 is detected in filamentous nuclear inclusions, which are the pathological hallmark of OPMD. Using both immunoelectron microscopy and fluorescence confocal microscopy, the OPMD-specific nuclear inclusions appeared decorated by anti-PABP2 antibodies. In addition, the inclusions were labeled with antibodies directed against ubiquitin and the subunits of the proteasome and contained a form of PABP2 that was more resistant to salt extraction than the protein dispersed in the nucleoplasm. This suggests that the polyalanine expansions in PABP2 induce a misfolding and aggregation of the protein into insoluble inclusions, similarly to events in neurodegenerative diseases caused by CAG/polyglutamine expansions. No significant differences were observed in the steady-state poly(A) tail length in OPMD and normal myoblasts. However, the nuclear inclusions were shown to sequester poly(A) RNA. This raises the possibility that in OPMD the polyalanine expansions in the PABP2 protein may interfere with the cellular traffic of poly(A) RNA.

Regulation of the Nuclear Poly(A)-binding Protein by Arginine Methylation in Fission Yeast

“…Interestingly, the extent of PAPB2 oligomerization is a key factor associated with the formation of nuclear aggregates found in cells of OPMD-afflicted individuals (20,51). Having established that Pab2 oligomerization was modulated by arginine methylation (Fig.…”

“…Pab2 Overexpression Causes Toxicity That Is Exacerbated in rmt1-null Cells-Mutations in the human pabp2 gene are linked to oculopharyngeal muscular dystrophy (OMPD) (19), a disease characterized by fibrous inclusions in the nuclei of skeletal muscle fibers (20). Interestingly, the extent of PAPB2 oligomerization is a key factor associated with the formation of nuclear aggregates found in cells of OPMD-afflicted individuals (20,51).…”

“…An understanding of the in vivo mechanism by which PABP2 regulates polyadenylation is significant, because the human genetic disorder oculopharyngeal muscular dystrophy (OPMD) is linked to mutations in the pabp2 gene (19). A physiological hallmark of this disorder is the accumulation of fibrous inclusions in the nuclei of skeletal muscle fibers that consist of PABP2 aggregates (20). Yet, the molecular defects in PABP2 function that lead to the establishment of this disease remain unclear.…”

Modeling Oculopharyngeal Muscular Dystrophy in Myotube Cultures Reveals Reduced Accumulation of Soluble Mutant PABPN1 Protein

“…36 Fluorescence in situ hybridization was performed using biotinylated 2=-O-alkyl polyuridine, as previously described. 10 Images were recorded with a Zeiss Axiovert, Oberkochen, Germany (model 135TV) fluorescence microscope equipped with a 100-W mercury arc lamp and a ϫ40/1.3 NA plan Apo objective. Single confocal sections were obtained with a Leica TCS/SP2 confocal microscope system equipped with a ϫ100/1.4 NA plan Apo objective.…”

“…Taken together, these observations suggest that the expression of PABPN1 in myotube cultures of the WTA or D7E clones led to protein accumulation in insoluble nuclear structures that resembled the INIs that are formed in affected patient muscles. 10 …”