PABPN1
Polyadenylate-binding protein 2 (PABP-2) (Poly(A)-binding protein 2) (Nuclear poly(A)-binding protein 1) (Poly(A)-binding protein II) (PABII) (Polyadenylate-binding nuclear protein 1) [PAB2] [PABP2]
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Post-transcription mRNA processing in the 3'-untranslated region (UTR) of transcripts alters mRNA landscape. Alternative polyadenylation (APA) utilization in the 3'-UTR often leads to shorter 3'-UTR affecting mRNA stability, a process that is regulated by PABPN1. In skeletal muscles PABPN1 levels reduce with age and a greater decrease in found in Oculopharyngeal muscular dystrophy (OPMD). OPMD is a late onset autosomal dominant myopathy caused by expansion mutation in PABPN1. In OPMD models a shift from distal to proximal polyadenylation site utilization in the 3'-UTR, and PABPN1 was shown to play a prominent role in APA. Whether PABPN1-mediated APA transcripts are functional is not fully understood. We investigate nuclear export and translation efficiency of transcripts in OPMD models. We focused on autophagy-regulated genes (ATGs) with APA utilization in cell models with reduced functional PABPN1. We provide evidence that ATGs transcripts from distal PAS retain in the nucleus and thus have reduced translation efficiency in cells with reduced PABPN1. In contrast, transcripts from proximal PAS showed a higher cytoplasmic abundance but a reduced occupancy in the ribosome. We therefore suggest that in reduced PABPN1 levels ATG transcripts from APA may not effectively translate to proteins. In those conditions we found constitutive autophagosome fusion and reduced autophagy flux. Augmentation of PABPN1 restored autophagosome fusion, suggesting that PABPN1-mediated APA plays a role in autophagy in OPMD and in aging muscles.
Keywords
- Gerotarget
- PABPN1
- aging muscles
- alternative polyadenylation site
- autophagy
- mRNA processing
Poly(A) Binding Protein Nuclear 1 (PABPN1) is a multifunctional regulator of mRNA processing, and its expression levels specifically decline in aging muscles. An expansion mutation in PABPN1 is the genetic cause of oculopharyngeal muscle dystrophy (OPMD), a late onset and rare myopathy. Moreover, reduced PABPN1 expression correlates with symptom manifestation in OPMD. PABPN1 regulates alternative polyadenylation site (PAS) utilization. However, the impact of PAS utilization on cell and tissue function is poorly understood. We hypothesized that altered PABPN1 expression levels is an underlying cause of muscle wasting. To test this, we stably down-regulated PABPN1 in mouse tibialis anterior (TA) muscles by localized injection of adeno-associated viruses expressing shRNA to PABPN1 (shPab). We found that a mild reduction in PABPN1 levels causes muscle pathology including myofiber atrophy, thickening of extracellular matrix and myofiber-type transition. Moreover, reduced PABPN1 levels caused a consistent decline in distal PAS utilization in the 3'-UTR of a subset of OPMD-dysregulated genes. This alternative PAS utilization led to up-regulation of Atrogin-1, a key muscle atrophy regulator, but down regulation of proteasomal genes. Additionally reduced PABPN1 levels caused a reduction in proteasomal activity, and transition in MyHC isotope expression pattern in myofibers. We suggest that PABPN1-mediated alternative PAS utilization plays a central role in aging-associated muscle wasting.
MeSH Terms
- Aging
- Animals
- Dependovirus
- Gene Expression Regulation
- Humans
- Mice
- Muscle Proteins
- Muscle, Skeletal
- Muscular Dystrophy, Oculopharyngeal
- Poly(A)-Binding Protein I
- RNA, Messenger
- SKP Cullin F-Box Protein Ligases
Alanine expansion mutations in poly(A)-binding protein nuclear 1 (PABPN1) cause muscle weakness in the late-onset disorder oculopharyngeal muscular dystrophy. In affected muscles, expanded PABPN1 forms nuclear aggregates, depleting levels of soluble PABPN1 and inducing a genome-wide shift from distal to proximal polyadenylation site usage. PABPN1 protein accumulation is regulated by the ubiquitin proteasome system, which is highly dysregulated in oculopharyngeal muscular dystrophy. We show that ARIH2 E3-ligase regulates PABPN1 protein accumulation and aggregation. Levels of ARIH2 mRNA are regulated by PABPN1 via proximal polyadenylation site usage. We demonstrate that masking the proximal polyadenylation site in ARIH2 3' untranslated region by antisense oligonucleotides elevates the expression of ARIH2 and PABPN1 and restores myogenic defects that are induced by ARIH2 or PABPN1 down-regulation in cell culture. In vivo ARIH2 mRNA levels significantly decrease from midlife in vastus lateralis muscles and highly correlate with muscle degeneration. We suggest that the expression of both genes is maintained by a feed-forward loop between mRNA stability regulated by PABPN1 and protein turnover regulated by ARIH2.
MeSH Terms
- Aging
- Animals
- Blotting, Western
- Cell Line
- Gene Expression Regulation
- Humans
- Immunohistochemistry
- Immunoprecipitation
- Muscle, Skeletal
- Muscular Dystrophy, Oculopharyngeal
- Oligonucleotide Array Sequence Analysis
- Poly(A)-Binding Protein I
- Reverse Transcriptase Polymerase Chain Reaction
- Transfection
- Ubiquitin-Protein Ligases
Oculopharyngeal muscular dystrophy (OPMD) is caused by trinucleotide repeat expansion mutations in Poly(A) binding protein 1 (PABPN1). PABPN1 is a regulator of mRNA stability and is ubiquitously expressed. Here we investigated how symptoms in OPMD initiate only at midlife and why a subset of skeletal muscles is predominantly affected. Genome-wide RNA expression profiles from Vastus lateralis muscles human carriers of expanded-PABPN1 at pre-symptomatic and symptomatic stages were compared with healthy controls. Major expression changes were found to be associated with age rather than with expression of expanded-PABPN1, instead transcriptomes of OPMD and elderly muscles were significantly similar (P<0.05). Using k-means clustering we identified age-dependent trends in both OPMD and controls, but trends were often accelerated in OPMD. We report an age-regulated decline in PABPN1 levels in Vastus lateralis muscles from the fifth decade. In concurrence with severe muscle degeneration in OPMD, the decline in PABPN1 accelerated in OPMD and was specific to skeletal muscles. Reduced PABPN1 levels (30% to 60%) in muscle cells induced myogenic defects and morphological signatures of cellular aging in proportion to PABPN1 expression levels. We suggest that PABPN1 levels regulate muscle cell aging and OPMD represents an accelerated muscle aging disorder.
MeSH Terms
- Adolescent
- Adult
- Aged, 80 and over
- Aging
- Animals
- Case-Control Studies
- Cellular Senescence
- Gene Expression Regulation
- Humans
- Mice
- Middle Aged
- Muscle Weakness
- Muscle, Skeletal
- Muscular Dystrophy, Oculopharyngeal
- Poly(A)-Binding Protein I
- RNA, Messenger
- Transcriptome
- Young Adult