Identification of AnFus3 [MpkB] associated proteins and AnFus3 interactions with the velvet complex components.

Abstract

<p>(A) Life cycle of <i>Aspergillus nidulans</i> and developmental functions of AnSte12 [SteA], LaeA and velvet domain proteins. Germination of spores leads to tube-like vegetative filaments (hyphae) which become competent for environmental signals after at least 12 hours of growth. Exposure of developmentally competent hyphae to light (or aeration) leads to asexual development (conidiophores and asexual spores [conidia]) in 24 hours. VosA-VelB inhibits asexual differentiation. Incubation in dark (96 hours) induces the sexual cycle with sexual fruiting bodies (cleistothecia) which are nursed by globose Hülle cells. LaeA is required for Hülle cell formation. VelB-VeA supports sexual development together with AnSte12 [SteA]. The VelB-VeA-LaeA trimeric complex coordinates development with secondary metabolism. Co; conidia, S; stalk, Cl; cleistothecium, Hc; Hülle cells. (B) A silver stain treated 5–14% gradient SDS polyacrylamide gel of AnFus3 [MpkB]::cTAP from vegetatively, asexually (on plates, under light) and sexually (on plates, in the dark) grown cultures at 30°C for 20 hours. Identified proteins from the excised lanes (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002816#pgen.1002816.s010" target="_blank">Table S1</a>). SA; SteA-AnSte12, MB; MpkB-AnFus3. (C) AnFus3-AnSte12 interaction <i>in vivo</i>. N-EYFP::AnFus3 [MpkB] fusion interacts with C-EYFP fusion of AnSte12 [SteA] in the nuclei (arrow) which were visualized by a monomeric red fluorescent protein histone 2A fusion (mRFP::Histone2A). (D) Interaction partners of AnFus3 [MpkB] in BIFC. (+) indicates AnFus3 interactions with VosA and LaeA at very early stages after germination (10–12 hours) and with VeA after 24 hours of hyphal growth. (−) indicates that VelB does not interact with AnFus3. Scale bars are 10 µm.</p

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