Fluorescence in-situ hybridization method reveals that carboxyl-terminal fragments of transactive response DNA-binding protein-43 truncated at the amino acid residue 218 reduce poly(A)+ RNA expression

Transactive response (TAR) DNA-binding protein 43 (TDP-43) has emerged as an important contributor to amyotrophic lateral sclerosis and frontotemporal lobar degeneration. To understand the association of TDP-43 with complex RNA processing in disease pathogenesis, we performed fluorescence in-situ hybridization using HeLa cells transfected with a series of deleted TDP-43 constructs and investigated the effect of truncation of TDP-43 on the expression of poly(A)(+) RNA. Endogenous and overexpressed full-length TDP-43 localized to the perichromatin region and interchromatin space adjacent to poly(A)(+) RNA. Deleted variants of TDP-43 containing RNA recognition motif 1 and truncating N-terminal region induced cytoplasmic inclusions in which poly(A)(+) RNA was recruited. Carboxyl-terminal TDP-43 truncated at residue 202 or 218 was distributed in the cytoplasm as punctate structures. Carboxyl-terminal TDP-43 truncated at residue 218, but not at 202, significantly decreased poly(A)(+) RNA expression by approximate to 24% compared with the level in control cells. Our results suggest that the disturbance of RNA metabolism induced by pathogenic fragments plays central roles in the pathogenesis of amyotrophic lateral sclerosis and frontotemporal lobar degeneration.