Localization and role of MYO-1, an endocytic protein in hyphae of Neurospora crassa

• Published in: Fungal genetics and biology 2016-03, Vol.88, p.24-34
• Main Authors: Lara-Rojas, Fernando, Bartnicki-García, Salomón, Mouriño-Pérez, Rosa R.
• Format: Article
• Language: English
• Subjects: Actin Cytoskeleton - metabolism; Endocytosis; Fungal Proteins - genetics; Fungal Proteins - metabolism; Genome, Fungal; Green Fluorescent Proteins; Hyphae - growth & development; Hyphae - metabolism; Hyphae - ultrastructure; Morphogenesis; Mutation; MYO-1; Myosin; Myosins - genetics; Myosins - metabolism; Neurospora crassa; Neurospora crassa - genetics; Neurospora crassa - growth & development; Neurospora crassa - metabolism; Phenotype; Polarized growth; Spores, Fungal - physiology
• ISSN: 1087-1845; 1096-0937
• DOI: 10.1016/j.fgb.2016.01.009

•MYO-1 is a component of the endocytic patches of N. crassa.•MYO-1 is close to the cortex and is recruited in earlier stages of endocytosis.•First phenotype description of Δmyo-1 mutant that is essential for Aspergillus spp.•MYO-1-deprived cells have a compromised actin cytoskeleton affecting the Spk. The subapical endocytic collar is a prominent feature of hyphae of Neurospora crassa. It comprises a dynamic collection of actin patches associated with a number of proteins required for endocytosis, namely, ARP-2/3 complex, fimbrin, coronin, etc. We presently show that MYO-1 is another key component of this endocytic collar. A myo-1 sequence was identified in the genome of N. crassa and used it to generate a strain with a myo-1-sgfp allele under the ccg1 promoter. Examination of living hyphae by confocal microscopy, revealed MYO-1-GFP located mainly as a dynamic collection of small patches arranged in collar-like fashion in the hyphal subapex. Dual tagging showed MYO-1-GFP partially colocalized with two other endocytic proteins, fimbrin and coronin. MYO-1 was also present during septum formation. By recovering a viable strain, albeit severely inhibited, after deletion of myo-1, it was possible to investigate the phenotypic consequences of the elimination of MYO-1. Deletion of myo-1 caused a severe reduction in growth rate (95%), near absence of aerial mycelium and no conidiation. A reduced uptake of the lipophilic dye FM4-64 indicated a deficiency in endocytosis in the Δmyo-1 mutant. Hyphae were produced by the Δmyo-1 mutant but their morphogenesis was severely affected; hyphal morphology was distorted displaying irregular periods of isotropic and polarized growth. The morphological alterations were accompanied, and presumably caused, by a disruption in the organization and dynamics of a myosin-deprived actin cytoskeleton that, ultimately, compromised the stability and function of the Spitzenkörper as a vesicle supply center.