MTB-3, a Microtubule Plus-End Tracking Protein (+TIP) of Neurospora crassa

Abstract

The microtubule (MT) “plus end” constitutes the platform for the accumulation of a structurally and functionally diverse group of proteins, collectively called “MT plus-end tracking proteins” (+TIPs). +TIPs control MT dynamics and link MTs to diverse sub-cellular structures. Neurospora crassa MicroTubule Binding protein-3 (MTB-3) is the homolog of yeast EB1, a highly conserved +TIP. To address the function of MTB-3, we examined strains with mtb-3 deletions, and we tagged MTB-3 with GFP to assess its dynamic behavior. MTB-3-GFP was present as comet-like structures distributed more or less homogeneously within the hyphal cytoplasm, and moving mainly towards the apex at speeds up to 4× faster than the normal hyphal elongation rates. MTB-3-GFP comets were present in all developmental stages, but were most abundant in mature hyphae. MTB-3-GFP comets were observed moving in anterograde and retrograde direction along the hypha. Retrograde movement was also observed as originating from the apical dome. The integrity of the microtubular cytoskeleton affects the presence and dynamics of MTB-3-GFP comets, while actin does not seem to play a role. The size of MTB-3-GFP comets is affected by the absence of dynactin and conventional kinesin. We detected no obvious morphological phenotypes in Δmtb-3 mutants but there were fewer MTs in Δmtb-3, MTs were less bundled and less organized. Compared to WT, both MT polymerization and depolymerization rates were significantly decreased in Δmtb-3. In summary, the lack of MTB-3 affects overall growth and morphological phenotypes of N. crassa only slightly, but deletion of mtb-3 has strong effect on MT dynamics.

The microtubule end‐binding protein FgEB1 regulates polar growth and fungicide sensitivity via different interactors in Fusarium graminearum

“…Consistent with the phenotypic changes of ΔFgEB1, the MAL3 deletion mutants of S. pombe produces branched and curved cells instead of the typical cylindrical shape of wild‐type cells (Beinhauer et al ., ). However, in filamentous fungus N. crassa , EB1 mutants only exhibited slightly reduced linear growth and increased conidiation, but did not show an obvious change in hyphal morphology in comparison with the wild type (Mouriño‐Pérez et al ., ). These studies indicate functions of EB1 homologs are species‐specific in different fungal species.…”

“…In Saccharomyces cerevisiae, the EB1 ortholog Bim1 combining with Kar9 and Stu2 is involved not only in microtubule modulation in cytoplasm but also in the accurate positioning and elongation of spindle in anaphase (Miller et al, 2000;Meednu et al, 2008;Zimniak et al, 2009). However, the deletion of MTB-3 (encoding EB1) was found to slightly affect overall growth and morphological phenotypes of Neurospora crassa, although the mutant showed decreased polymerization and depolymerization rates of microtubule (Mouriño-P erez et al, 2013), indicating that the functions of EB1 in filamentous fungi might be different from those in the yeasts.…”

Hyphal ontogeny in Neurospora crassa: a model organism for all seasons

“…Lis1, an MT plus-end-binding protein, is involved in the regulation of the dynein/dynactin complex in humans 75 . In N. crassa , two paralogues of Lis1—LIS1-1 and LIS1-2—were identified in the hyphal apical dome as short filaments or comet-like structures decorating the terminal plus-ends of the MTs 76 , similar to the distribution of the MT plus-end-binding protein MTB-3, a homologue of EB1 77 . LIS1-1 localization was found to be dependent on the minus-end-directed MT-associated motor dynein and dynactin (p150Glued) 76, 78 .…”

Fungal Morphogenesis, from the Polarized Growth of Hyphae to Complex Reproduction and Infection Structures

“…It was shown that a MT plus-end tracking protein (ϩTIP), an EB1 orthologue, accumulates at the tip and moves in a retrograde manner. Also, bleaching experiments suggested MT polymerization starting from the tip (133,134). Whether there are MTOCs at septa is still unsolved.…”