4 research outputs found
Detection of TAR DNA-binding protein 43 (TDP-43) oligomers as initial intermediate species during aggregate formation
Aggregates of the RNA-binding protein TDP-43 (TAR DNAbinding protein) are a hallmark of the overlapping neurodegenerative disorders amyotrophic lateral sclerosis (ALS) and
frontotemporal dementia. The process of TDP-43 aggregation
remains poorly understood, and whether it includes formation
of intermediate complexes is unknown. Here, we analyzed
aggregates derived from purified TDP-43 under semidenaturing conditions, identifying distinct oligomeric complexes at the
initial time points before the formation of large aggregates. We
found that this early oligomerization stage is primarily driven by
TDP-43’s RNA-binding region. Specific binding to GU-rich
RNA strongly inhibited both TDP-43 oligomerization and
aggregation, suggesting that RNA interactions are critical for
maintaining TDP-43 solubility. Moreover, we analyzed TDP-43
liquid–liquid phase separation and detected similar detergentresistant oligomers upon maturation of liquid droplets into solid-like fibrils. These results strongly suggest that the oligomers
form during the early steps of TDP-43 misfolding. Importantly,
the ALS-linked TDP-43 mutations A315T and M337V significantly accelerate aggregation, rapidly decreasing the monomeric
population and shortening the oligomeric phase. We also show
that aggregates generated from purified TDP-43 seed intracellular
aggregation detected by established TDP-43 pathology markers.
Remarkably, cytoplasmic aggregate seeding was detected earlier
for the A315T and M337V variants and was 50% more widespread
than forWTTDP-43 aggregates.We provide evidence for aninitial
step of TDP-43 self-assembly into intermediate oligomeric complexes, whereby these complexes may provide a scaffold for aggregation. This process is altered by ALS-linked mutations, underscoring the role of perturbationsin TDP-43 homeostasisin protein
aggregation and ALS-FTD pathogenesis
Genome-Wide Mapping and Interrogation of the Nmp4 Antianabolic Bone Axis
PTH is an osteoanabolic for treating osteoporosis but its potency wanes. Disabling the transcription factor nuclear matrix protein 4 (Nmp4) in healthy, ovary-intact mice enhances bone response to PTH and bone morphogenetic protein 2 and protects from unloading-induced osteopenia. These Nmp4(-/-) mice exhibit expanded bone marrow populations of osteoprogenitors and supporting CD8(+) T cells. To determine whether the Nmp4(-/-) phenotype persists in an osteoporosis model we compared PTH response in ovariectomized (ovx) wild-type (WT) and Nmp4(-/-) mice. To identify potential Nmp4 target genes, we performed bioinformatic/pathway profiling on Nmp4 chromatin immunoprecipitation sequencing (ChIP-seq) data. Mice (12 w) were ovx or sham operated 4 weeks before the initiation of PTH therapy. Skeletal phenotype analysis included microcomputed tomography, histomorphometry, serum profiles, fluorescence-activated cell sorting and the growth/mineralization of cultured WT and Nmp4(-/-) bone marrow mesenchymal stem progenitor cells (MSPCs). ChIP-seq data were derived using MC3T3-E1 preosteoblasts, murine embryonic stem cells, and 2 blood cell lines. Ovx Nmp4(-/-) mice exhibited an improved response to PTH coupled with elevated numbers of osteoprogenitors and CD8(+) T cells, but were not protected from ovx-induced bone loss. Cultured Nmp4(-/-) MSPCs displayed enhanced proliferation and accelerated mineralization. ChIP-seq/gene ontology analyses identified target genes likely under Nmp4 control as enriched for negative regulators of biosynthetic processes. Interrogation of mRNA transcripts in nondifferentiating and osteogenic differentiating WT and Nmp4(-/-) MSPCs was performed on 90 Nmp4 target genes and differentiation markers. These data suggest that Nmp4 suppresses bone anabolism, in part, by regulating IGF-binding protein expression. Changes in Nmp4 status may lead to improvements in osteoprogenitor response to therapeutic cues