The influence of pathological mutations and proline substitutions in TDP-43 glycine-rich peptides on its amyloid properties and cellular toxicity

• Published in: PloS one 2014-08, Vol.9 (8), p.e103644
• Main Authors: Sun, Chia-Sui, Wang, Cindy Yu-Hsiang, Chen, Bryan Po-Wen, He, Ruei-Yu, Liu, Gerard Chun-Hao, Wang, Chih-Hsien, Chen, Wenlung, Chern, Yijuang, Huang, Joseph Jen-Tse
• Format: Article
• Language: English
• Subjects: Alzheimer's disease; Alzheimers disease; Amino Acid Sequence; Amino Acid Substitution; Amyloid - metabolism; Amyloid - ultrastructure; Amyloidogenesis; Amyloidosis; Amyotrophic lateral sclerosis; Animals; Asparagine; Assaying; Biochemistry; Biology and Life Sciences; C-Terminus; Calcein; Cell Membrane - drug effects; Cell Membrane - metabolism; Cell survival; Circular dichroism; Cytotoxicity; Degeneration; Deoxyribonucleic acid; Dichroism; DNA; DNA-binding protein; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; DNA-Binding Proteins - toxicity; Electron microscopy; Fibers; Fourier transforms; Frontotemporal dementia; Genetic aspects; Glutamine; Glycine; Glycine - metabolism; Inclusion bodies; Lasers; Leakage; Medicine and Health Sciences; Mice; Molecular Sequence Data; Mutant Proteins - metabolism; Mutant Proteins - toxicity; Mutation; Mutation - genetics; Neurotoxicity; Pathogenesis; Peptides; Peptides - chemistry; Peptides - metabolism; Peptides - toxicity; Proline; Proline - genetics; Protein Aggregates - drug effects; Protein binding; Protein Multimerization - drug effects; Protein Structure, Secondary; Proteins; Raman spectroscopy; Spectroscopy; Spectrum Analysis, Raman; Thiazoles - metabolism; Time Factors; Toxicity; Transmission electron microscopy; Ubiquitin; β-Amyloid
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0103644

TAR DNA-binding protein (TDP-43) was identified as the major ubiquitinated component deposited in the inclusion bodies in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U) in 2006. Later on, numerous ALS-related mutations were found in either the glycine or glutamine/asparagine-rich region on the TDP-43 C-terminus, which hinted on the importance of mutations on the disease pathogenesis. However, how the structural conversion was influenced by the mutations and the biological significance of these peptides remains unclear. In this work, various peptides bearing pathogenic or de novo designed mutations were synthesized and displayed their ability to form twisted amyloid fibers, cause liposome leakage, and mediate cellular toxicity as confirmed by transmission electron microscopy (TEM), circular dichroism (CD), Thioflavin T (ThT) assay, Raman spectroscopy, calcein leakage assay, and cell viability assay. We have also shown that replacing glycines with prolines, known to obstruct β-sheet formation, at the different positions in these peptides may influence the amyloidogenesis process and neurotoxicity. In these cases, GGG308PPP mutant was not able to form beta-amyloid, cause liposome leakage, nor jeopardized cell survival, which hinted on the importance of the glycines (308-310) during amyloidogenesis.