Aggregates of Small Nuclear Ribonucleic Acids (snRNAs) in Alzheimer's Disease

We recently discovered that protein components of the ribonucleic acid (RNA) spliceosome form cytoplasmic aggregates in Alzheimer's disease (AD) brain, resulting in widespread changes in RNA splicing. However, the involvement of small nuclear RNAs (snRNAs), also key components of the spliceosom...

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Published in	Brain pathology (Zurich, Switzerland) Vol. 24; no. 4; pp. 344 - 351
Main Authors	Hales, Chadwick M., Dammer, Eric B., Diner, Ian, Yi, Hong, Seyfried, Nicholas T., Gearing, Marla, Glass, Jonathan D., Montine, Thomas J., Levey, Allan I., Lah, James J.
Format	Journal Article
Language	English
Published	Switzerland Blackwell Publishing Ltd 01.07.2014 John Wiley & Sons, Inc John Wiley and Sons Inc
Subjects	2,2,7‐trimethylguanosine 7-trimethylguanosine Aged Aged, 80 and over Aggregates ALS Alzheimer Disease - metabolism Alzheimer's disease Brain - metabolism Cytoplasm - metabolism Female Fluorescent Antibody Technique Humans Male Microscopy, Electron, Transmission Middle Aged Neurofibrillary Tangles - metabolism quantitative PCR Real-Time Polymerase Chain Reaction RNA spliceosome RNA, Small Nuclear - metabolism snRNA Spinal Cord - metabolism tau Proteins - metabolism Tauopathies - metabolism 2,2,7-trimethylguanosine ALS quantitative PCR Alzheimer's disease RNA spliceosome snRNA
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ISSN	1015-6305 1750-3639 1750-3639
DOI	10.1111/bpa.12133

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Summary:	We recently discovered that protein components of the ribonucleic acid (RNA) spliceosome form cytoplasmic aggregates in Alzheimer's disease (AD) brain, resulting in widespread changes in RNA splicing. However, the involvement of small nuclear RNAs (snRNAs), also key components of the spliceosome complex, in the pathology of AD remains unknown. Using immunohistochemical staining of post‐mortem human brain and spinal cord, we identified cytoplasmic tangle‐shaped aggregates of snRNA in both sporadic and familial AD cases but not in aged controls or other neurodegenerative disorders. Immunofluorescence using antibodies reactive with the 2,2,7‐trimethylguanosine cap of snRNAs and transmission electron microscopy demonstrated snRNA localization with tau and paired helical filaments, the main component of neurofibrillary tangles. Quantitative real‐time polymerase chain reaction (PCR) showed U1 snRNA accumulation in the insoluble fraction of AD brains whereas other U snRNAs were not enriched. In combination with our previous results, these findings demonstrate that aggregates of U1 snRNA and U1 small nuclear ribonucleoproteins represent a new pathological hallmark of AD.
Bibliography:	Figure S1. snRNA cytoplasmic aggregates found throughout the AD brain. 2,2,7-TMG immunohistochemistry staining of fixed free-floating AD brain (Braak VI). Sections from hippocampus, temporal, frontal and occipital cortices are shown with black arrows pointing to cytoplasmic aggregates. Scale bar = 10 μm.Figure S2. snRNA aggregates colocalize with other snRNP aggregates in AD post-mortem brain. Immunfluorescence staining of fixed free-floating brain with (A,E) Hoechst, (B,F) 2,2,7-TMG, (C) U1-70k and (G) SmD. Overlay images are shown as well (D,H). White arrow point to overlapping snRNA and snRNPs. Scale bar = 10 μm.Figure S3. snRNA in ALS frontal cortex and spinal cord. Representative examples provided. Fifty micrometer free-floating cryopreserved section from (A) ALS human frontal cortex, (B) control spinal cord, (C) AD spinal cord and (D) ALS spinal cord were immunostained with 2,2,7-TMG cap antibody. Normal nuclear snRNA is present in all cells including motor neurons (B,C,D black arrows). Scale bar = 5 μm.Figure S4. RNA hybridization demonstrates U1-snRNA aggregates in neurons with neurofibrillary tangles. RNA hybridization was performed on an AD free-floating tissue section using biotinylated 2′-O-Me-RNA probe against a 17-nucleotide U1 snRNA-specific sequence. Counterstaining with thioflavin S (green) was applied before mounting slides. Black arrows point to U1 snRNA in a tangle-bearing neuron.Table S1. Demographic information of subjects in this study.Table S2. TaqMan Primer/Probe sets for quantitative PCR. ArticleID:BPA12133 Emory Alzheimer's Disease Research Center-NIA-AG025688 ADRC P50 AG05136 istex:EF30A60F43D02B44B914509A3747A68ABE4F7CBE Emory Neuroscience NINDS Core Facility-P30NS055077 NIA (AG038259) ark:/67375/WNG-GBR9SNTT-N NIAP01AG014449 Robert P. Apkarian Integrated Electron Microscopy Core of Emory University Alzheimer's Association New Investigator Research Award (NIRG-12-242297) ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 14 content type line 23 ObjectType-Article-2 ObjectType-Feature-1
ISSN:	1015-6305 1750-3639 1750-3639
DOI:	10.1111/bpa.12133