title: Alanine repeats influence protein localization in splicing speckles and paraspeckles.
abstract: Mammalian splicing regulatory protein RNA-binding motif protein 4 (RBM4) has an alanine repeat-containing C-terminal domain (CAD) that confers both nuclear- and splicing speckle-targeting activities. Alanine-repeat expansion has pathological potential. Here we show that the alanine-repeat tracts influence the subnuclear targeting properties of the RBM4 CAD in cultured human cells. Notably, truncation of the alanine tracts redistributed a portion of RBM4 to paraspeckles. The alanine-deficient CAD was sufficient for paraspeckle targeting. On the other hand, alanine-repeat expansion reduced the mobility of RBM4 and impaired its splicing activity. We further took advantage of the putative coactivator activator (CoAA)-RBM4 conjoined splicing factor, CoAZ, to investigate the function of the CAD in subnuclear targeting. Transiently expressed CoAZ formed discrete nuclear foci that emerged and subsequently separated-fully or partially-from paraspeckles. Alanine-repeat expansion appeared to prevent CoAZ separation from paraspeckles, resulting in their complete colocalization. CoAZ foci were dynamic but, unlike paraspeckles, were resistant to RNase treatment. Our results indicate that the alanine-rich CAD, in conjunction with its conjoined RNA-binding domain(s), differentially influences the subnuclear localization and biogenesis of RBM4 and CoAZ.
authors.affiliation: Program in Molecular Medicine, National Yang-Ming University and Academia Sinica, Taipei, Taiwan Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei, Taiwan.
authors.forename: Shuo-Hsiu
authors.initials: SH
authors.lastname: Chang
authors.affiliation: Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
authors.forename: Wei-Lun
authors.initials: WL
authors.lastname: Chang
authors.affiliation: Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
authors.forename: Chia-Chen
authors.initials: CC
authors.lastname: Lu
authors.affiliation: Program in Molecular Medicine, National Yang-Ming University and Academia Sinica, Taipei, Taiwan Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan wtarn@ibms.sinica.edu.tw.
authors.forename: Woan-Yuh
authors.initials: WY
authors.lastname: Tarn
mesh_terms: D000409:Alanine; D017398:Alternative Splicing; D022003:Cell Nucleus Structures / Q000737:chemistry*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D009687:Nuclear Proteins / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D010455:Peptides; D021382:Protein Sorting Signals; D016601:RNA-Binding Proteins / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid
chemical_list: D009687:Nuclear Proteins; D010455:Peptides; D021382:Protein Sorting Signals; C473692:RBM4 protein, human; D016601:RNA-Binding Proteins; C019529:polyalanine; D000409:Alanine