title: Phosphoregulation of the human SMN complex.
abstract: The survival motor neuron (SMN) complex is a macromolecular machine comprising 9 core proteins: SMN, Gemins2-8 and unrip in vertebrates. It performs tasks in RNA metabolism including the cytoplasmic assembly of spliceosomal small nuclear ribonucleoprotein particles (snRNPs). The SMN complex also localizes to the nucleus, where it accumulates in Cajal Bodies (CB) and may function in transcription and/or pre-mRNA splicing. The SMN complex is subject to extensive phosphorylation. Detailed understanding of SMN complex regulation necessitates a comprehensive analysis of these post-translational modifications. Here, we report on the first comprehensive phosphoproteome analysis of the intact human SMN complex, which identify 48 serine/threonine phosphosites in the complex. We find that 7 out of 9 SMN components of the intact complex are phosphoproteins and confidently place 29 phosphorylation sites, 12 of them in SMN itself. By the generation of multi non-phosphorylatable or phosphomimetic variants of SMN, respectively, we address to which extent phosphorylation regulates SMN complex function and localization. Both phosphomimetic and non-phosphorylatable variants assemble into intact SMN complexes and can compensate the loss of endogenous SMN in snRNP assembly at least to some extent. However, they partially or completely fail to target to nuclear Cajal bodies. Moreover, using a mutant of SMN, which cannot be phosphorylated on previously reported tyrosine residues, we provide first evidence that this PTM regulates SMN localization and nuclear accumulation. Our data suggest complex regulatory cues mediated by phosphorylation of serine/threonine and tyrosine residues, which control the subcellular localization of the SMN complex and its accumulation in nuclear CB.
authors.affiliation: Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH), DKFZ-ZMBH Alliance, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany. Electronic address: alma.husedzinovic@nct-heidelberg.de.
authors.forename: Alma
authors.initials: A
authors.lastname: Husedzinovic
authors.affiliation: Evotec AG, Am Klopferspitz 19a, D-82152 Martinsried, Germany.
authors.forename: Felix
authors.initials: F
authors.lastname: Oppermann
authors.affiliation: Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH), DKFZ-ZMBH Alliance, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Draeger-Meurer
authors.affiliation: Theodor Boveri Institute, Biocenter of the University of Wuerzburg, D-97074 Wuerzburg, Germany.
authors.forename: Ashwin
authors.initials: A
authors.lastname: Chari
authors.affiliation: Theodor Boveri Institute, Biocenter of the University of Wuerzburg, D-97074 Wuerzburg, Germany.
authors.forename: Utz
authors.initials: U
authors.lastname: Fischer
authors.affiliation: Evotec AG, Am Klopferspitz 19a, D-82152 Martinsried, Germany.
authors.forename: Henrik
authors.initials: H
authors.lastname: Daub
authors.affiliation: Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH), DKFZ-ZMBH Alliance, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany. Electronic address: o.gruss@zmbh.uni-heidelberg.de.
authors.forename: Oliver J
authors.initials: OJ
authors.lastname: Gruss
mesh_terms: D000595:Amino Acid Sequence; D020541:Coiled Bodies / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D020543:Proteome / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism; D055532:SMN Complex Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000737:chemistry / Q000378:metabolism; D013912:Threonine / Q000737:chemistry / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D010750:Phosphoproteins; D020543:Proteome; D017411:Ribonucleoproteins, Small Nuclear; D055532:SMN Complex Proteins; D013912:Threonine; D014443:Tyrosine; D012694:Serine
keywords: Functional regulation; Phosphorylation; Proteomics; SMA; SMN