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The RNA-binding motif 45 (RBM45) protein accumulates in inclusion bodies in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP) patients.

Collins M, Riascos D, Kovalik T, An J, Krupa K, Krupa K, Hood BL, Conrads TP, Renton AE, Traynor BJ, Bowser R - Acta Neuropathol. (2012)

Bottom Line: These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus.By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies.In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin.

View Article: PubMed Central - PubMed

Affiliation: Departments of Neurobiology and Pathology, University of Pittsburgh, PA, USA.

ABSTRACT
RNA-binding protein pathology now represents one of the best characterized pathologic features of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration patients with TDP-43 or FUS pathology (FTLD-TDP and FTLD-FUS). Using liquid chromatography tandem mass spectrometry, we identified altered levels of the RNA-binding motif 45 (RBM45) protein in the cerebrospinal fluid (CSF) of ALS patients. This protein contains sequence similarities to TAR DNA-binding protein 43 (TDP-43) and fused-in-sarcoma (FUS) that are contained in cytoplasmic inclusions of ALS and FTLD-TDP or FTLD-FUS patients. To further characterize RBM45, we first verified the presence of RBM45 in CSF and spinal cord tissue extracts of ALS patients by immunoblot. We next used immunohistochemistry to examine the subcellular distribution of RBM45 and observed in a punctate staining pattern within nuclei of neurons and glia in the brain and spinal cord. We also detected RBM45 cytoplasmic inclusions in 91 % of ALS, 100 % of FTLD-TDP and 75 % of Alzheimer's disease (AD) cases. The most extensive RBM45 pathology was observed in patients that harbor the C9ORF72 hexanucleotide repeat expansion. These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus. By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies. In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin. We identified RBM45 using a proteomic screen of CSF from ALS and control subjects for candidate biomarkers, and link this RNA-binding protein to inclusion pathology in ALS, FTLD-TDP and AD.

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Double-label immunofluorescence for RBM45 (green) and TDP-43 (red) in the spinal cord and hippocampus. a–c RBM45 co-localization with TDP-43 positive inclusions in ALS spinal cord motor neurons. DAPI visualizes nuclei (blue) in the merged images. RBM45 also remained in the nucleus of a motor neuron with cytoplasmic TDP-43 inclusions (c). d–f RBM45 co-localization with TDP-43 inclusions in the dentate gyrus of FTLD-TDP (d, e) and AD (f) cases. Arrows denote co-localization in intranuclear (d) and cytoplasmic (e, f) inclusions. Speckled RBM45 nuclear stain is observed in all panels and is devoid of TDP-43. Scale bars denote 20 μm in panelsa–c and 30 μm in panelsd–f. Each panel represents the following case numbers in Table 1: a 2, b 3, c 7, d 24, e 28, f 30
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Fig5: Double-label immunofluorescence for RBM45 (green) and TDP-43 (red) in the spinal cord and hippocampus. a–c RBM45 co-localization with TDP-43 positive inclusions in ALS spinal cord motor neurons. DAPI visualizes nuclei (blue) in the merged images. RBM45 also remained in the nucleus of a motor neuron with cytoplasmic TDP-43 inclusions (c). d–f RBM45 co-localization with TDP-43 inclusions in the dentate gyrus of FTLD-TDP (d, e) and AD (f) cases. Arrows denote co-localization in intranuclear (d) and cytoplasmic (e, f) inclusions. Speckled RBM45 nuclear stain is observed in all panels and is devoid of TDP-43. Scale bars denote 20 μm in panelsa–c and 30 μm in panelsd–f. Each panel represents the following case numbers in Table 1: a 2, b 3, c 7, d 24, e 28, f 30

Mentions: We next examined the co-localization of RBM45 and TDP-43 pathology in ALS, FTLD-TDP and AD by double-label confocal microscopy, with DAPI to identify nuclei. We detected co-localization of RBM45 and TDP-43 within cytoplasmic inclusions of spinal cord motor neurons in ALS (Fig. 5a–c). We also noted ALS motor neurons with both nuclear RBM45 and co-localization in cytoplasmic TDP-43 inclusions (Fig. 5c). Interestingly, Fig. 5a, b represents ALS cases with the C9ORF72 repeat expansion and exhibits reduced RBM45 nuclear staining when compared to ALS cases without the repeat expansion.Fig. 5


The RNA-binding motif 45 (RBM45) protein accumulates in inclusion bodies in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP) patients.

Collins M, Riascos D, Kovalik T, An J, Krupa K, Krupa K, Hood BL, Conrads TP, Renton AE, Traynor BJ, Bowser R - Acta Neuropathol. (2012)

Double-label immunofluorescence for RBM45 (green) and TDP-43 (red) in the spinal cord and hippocampus. a–c RBM45 co-localization with TDP-43 positive inclusions in ALS spinal cord motor neurons. DAPI visualizes nuclei (blue) in the merged images. RBM45 also remained in the nucleus of a motor neuron with cytoplasmic TDP-43 inclusions (c). d–f RBM45 co-localization with TDP-43 inclusions in the dentate gyrus of FTLD-TDP (d, e) and AD (f) cases. Arrows denote co-localization in intranuclear (d) and cytoplasmic (e, f) inclusions. Speckled RBM45 nuclear stain is observed in all panels and is devoid of TDP-43. Scale bars denote 20 μm in panelsa–c and 30 μm in panelsd–f. Each panel represents the following case numbers in Table 1: a 2, b 3, c 7, d 24, e 28, f 30
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Related In: Results  -  Collection

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Fig5: Double-label immunofluorescence for RBM45 (green) and TDP-43 (red) in the spinal cord and hippocampus. a–c RBM45 co-localization with TDP-43 positive inclusions in ALS spinal cord motor neurons. DAPI visualizes nuclei (blue) in the merged images. RBM45 also remained in the nucleus of a motor neuron with cytoplasmic TDP-43 inclusions (c). d–f RBM45 co-localization with TDP-43 inclusions in the dentate gyrus of FTLD-TDP (d, e) and AD (f) cases. Arrows denote co-localization in intranuclear (d) and cytoplasmic (e, f) inclusions. Speckled RBM45 nuclear stain is observed in all panels and is devoid of TDP-43. Scale bars denote 20 μm in panelsa–c and 30 μm in panelsd–f. Each panel represents the following case numbers in Table 1: a 2, b 3, c 7, d 24, e 28, f 30
Mentions: We next examined the co-localization of RBM45 and TDP-43 pathology in ALS, FTLD-TDP and AD by double-label confocal microscopy, with DAPI to identify nuclei. We detected co-localization of RBM45 and TDP-43 within cytoplasmic inclusions of spinal cord motor neurons in ALS (Fig. 5a–c). We also noted ALS motor neurons with both nuclear RBM45 and co-localization in cytoplasmic TDP-43 inclusions (Fig. 5c). Interestingly, Fig. 5a, b represents ALS cases with the C9ORF72 repeat expansion and exhibits reduced RBM45 nuclear staining when compared to ALS cases without the repeat expansion.Fig. 5

Bottom Line: These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus.By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies.In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin.

View Article: PubMed Central - PubMed

Affiliation: Departments of Neurobiology and Pathology, University of Pittsburgh, PA, USA.

ABSTRACT
RNA-binding protein pathology now represents one of the best characterized pathologic features of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration patients with TDP-43 or FUS pathology (FTLD-TDP and FTLD-FUS). Using liquid chromatography tandem mass spectrometry, we identified altered levels of the RNA-binding motif 45 (RBM45) protein in the cerebrospinal fluid (CSF) of ALS patients. This protein contains sequence similarities to TAR DNA-binding protein 43 (TDP-43) and fused-in-sarcoma (FUS) that are contained in cytoplasmic inclusions of ALS and FTLD-TDP or FTLD-FUS patients. To further characterize RBM45, we first verified the presence of RBM45 in CSF and spinal cord tissue extracts of ALS patients by immunoblot. We next used immunohistochemistry to examine the subcellular distribution of RBM45 and observed in a punctate staining pattern within nuclei of neurons and glia in the brain and spinal cord. We also detected RBM45 cytoplasmic inclusions in 91 % of ALS, 100 % of FTLD-TDP and 75 % of Alzheimer's disease (AD) cases. The most extensive RBM45 pathology was observed in patients that harbor the C9ORF72 hexanucleotide repeat expansion. These RBM45 inclusions were observed in spinal cord motor neurons, glia and neurons of the dentate gyrus. By confocal microscopy, RBM45 co-localizes with ubiquitin and TDP-43 in inclusion bodies. In neurons containing RBM45 cytoplasmic inclusions we often detected the protein in a punctate pattern within the nucleus that lacked either TDP-43 or ubiquitin. We identified RBM45 using a proteomic screen of CSF from ALS and control subjects for candidate biomarkers, and link this RNA-binding protein to inclusion pathology in ALS, FTLD-TDP and AD.

Show MeSH
Related in: MedlinePlus