ROLE OF FUNGAL DYNEIN IN HYPHAL GROWTH, MICROTUBULE ORGANIZATION, SPINDLE POLE BODY MOTILITY AND NUCLEAR MIGRATION

Cytoplasmic dynein is a microtubule-associated motor protein with seve ral putative subcellular functions. Sequencing of the gene (DHC1) for cytoplasmic dynein heavy chain of the filamentous ascomycete, Nectria haematococca, revealed a 4,349-codon open reading frame (interrupted b y two introns) with four highly conserved P-loop motifs, typical of cy toplasmic dynein heavy chains, The predicted amino acid sequence is 78 .0% identical to the cytoplasmic dynein heavy chain of Neurospora cras sa, 70.2% identical to that of Aspergillus nidulans and 24.8% identica l to that of Saccharomyces cerevisiae, The genomic copy of DHC1 in N. haematococca wild-type strain T213 was disrupted by inserting a select able marker into the central motor domain. Mutants grew at 33% of the wildtype rate, forming dense compact colonies composed of spiral and h ighly branched hyphae. Major cytological phenotypes included (1) absen ce of aster-like arrays of cytoplasmic microtubules focused at the spi ndle pole bodies of post-mitotic and interphase nuclei, (2) limited po stmitotic nuclear migration, (3) lack of spindle pole body motility at interphase, (4) failure of spindle pole bodies to anchor interphase n uclei, (5) nonuniform distribution of interphase nuclei and (6) small or ephemeral Spitzenkorper at the apices of hyphal tip cells. Microtub ule distribution in the apical region of tip cells of the mutant was e ssentially normal. The nonuniform distribution of nuclei in hyphae res ulted primarily from a lack of both post-mitotic nuclear migration and anchoring of interphase nuclei by the spindle pole bodies. The result s support the hypothesis that DHC1 is required for the motility and fu nctions of spindle pole bodies, normal secretory vesicle transport to the hyphal apex and normal hyphal tip cell morphogenesis.