title: Two microtubule-plus-end binding proteins LIS1-1 and LIS1-2, homologues of human LIS1 in Neurospora crassa.
abstract: LIS1 is a microtubule (Mt) plus-end binding protein that interacts with the dynein/dynactin complex. In humans, LIS1 is required for proper nuclear and organelle migration during cell growth. Although gene duplication is absent from Neurospora crassa, we found two paralogues of human LIS1. We named them LIS1-1 and LIS1-2 and studied their dynamics and function by fluorescent tagging. At the protein level, LIS1-1 and LIS1-2 were very similar. Although, the characteristic coiled-coil motif was not present in LIS1-2. LIS1-1-GFP and LIS1-2-GFP showed the same cellular distribution and dynamics, but LIS1-2-GFP was less abundant. Both LIS1 proteins were found in the subapical region as single fluorescent particles traveling toward the cell apex, they accumulated in the apical dome forming prominent short filament-like structures, some of which traversed the Spitzenkörper (Spk). The fluorescent structures moved exclusively in anterograde fashion along straight paths suggesting they traveled on Mts. There was no effect in the filament behavior of LIS1-1-GFP in the Δlis1-2 mutant but the dynamics of LIS1-2-GFP was affected in the Δlis1-1 mutant. Microtubular integrity and the dynein-dynactin complex were necessary for the formation of filament-like structures of LIS1-1-GFP in the subapical and apical regions; however, conventional kinesin (KIN-1) was not. Deletion mutants showed that the lack of lis1-1 decreased cell growth by ∼75%; however, the lack of lis1-2 had no effect on growth. A Δlis1-1;Δlis1-2 double mutant showed slower growth than either single mutant. Conidia production was reduced but branching rate increased in Δlis1-1 and the Δlis1-1;Δlis1-2 double mutants. The absence of LIS1-1 had a strong effect on Mt organization and dynamics and indirectly affected nuclear and mitochondrial distribution. The absence of LIS1-1 filaments in dynein mutants (ropy mutants) or in benomyl treated hyphae indicates the strong association between this protein and the regulation of the dynein-dynactin complex and Mt organization. LIS1-1 and LIS1-2 had a high amino acid homology, nevertheless, the absence of the coiled-coil motif in LIS1-2 suggests that its function or regulation may be distinct from that of LIS1-1.
authors.affiliation: Departamento de Microbiología, Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE), Ensenada, B.C., Mexico.
authors.forename: Olga A
authors.initials: OA
authors.lastname: Callejas-Negrete
authors.affiliation: School of Biological Sciences, University of Missouri-Kansas City, Kansas City, MO, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Plamann
authors.affiliation: School of Biological Sciences, University of Missouri-Kansas City, Kansas City, MO, USA.
authors.forename: Robert
authors.initials: R
authors.lastname: Schnittker
authors.affiliation: Departamento de Microbiología, Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE), Ensenada, B.C., Mexico.
authors.forename: Salomon
authors.initials: S
authors.lastname: Bartnicki-García
authors.affiliation: School of Life Sciences, Arizona State University, Tempe, AZ, USA.
authors.forename: Robert W
authors.initials: RW
authors.lastname: Roberson
authors.affiliation: Departamento de Microbiología, Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE), Ensenada, B.C., Mexico.
authors.forename: Genaro
authors.initials: G
authors.lastname: Pimienta
authors.affiliation: Departamento de Microbiología, Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE), Ensenada, B.C., Mexico. Electronic address: rmourino@cicese.mx.
authors.forename: Rosa R
authors.initials: RR
authors.lastname: Mouriño-Pérez
mesh_terms: D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000378:metabolism; D000072159:Dynactin Complex; D004398:Dyneins / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009492:Neurospora crassa / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment
chemical_list: D000072159:Dynactin Complex; D005656:Fungal Proteins; D008869:Microtubule-Associated Proteins; D011993:Recombinant Fusion Proteins; D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; C084102:PAFAH1B1 protein, human; D004398:Dyneins
keywords: +TIP proteins; Dynein–dynactin; LIS1; Microtubules; Neurospora crassa