Functional characterization of Rho GTPases in Aspergillus niger uncovers conserved and diverged roles of Rho proteins within filamentous fungi

Abstract

SummaryRho GTPases are signalling molecules regulating morphology and multiple cellular functions including metabolism and vesicular trafficking. To understand the connection between polarized growth and secretion in the industrial model organism Aspergillus niger, we investigated the function of all Rho family members in this organism. We identified six Rho GTPases in its genome and used loss-of-function studies to dissect their functions. While RhoA is crucial for polarity establishment and viability, RhoB and RhoD ensure cell wall integrity and septum formation respectively. RhoC seems to be dispensable for A. niger. RacA governs polarity maintenance via controlling actin but not microtubule dynamics, which is consistent with its localization at the hyphal apex. Both deletion and dominant activation of RacA (Rac G18V ) provoke an actin localization defect and thereby loss of polarized tip extension. Simultaneous deletion of RacA and CftA (Cdc42) is lethal; however, conditional overexpression of RacA in this strain can substitute for CftA, indicating that both proteins concertedly control actin dynamics. We finally identified NoxR as a RacA-specific effector, which however, is not important for apical dominance as reported for A. nidulans but for asexual development. Overall, the data show that individual Rho GTPases contribute differently to growth and morphogenesis within filamentous fungi.

The Genomes of Three Uneven Siblings: Footprints of the Lifestyles of Three Trichoderma Species

“…According to their primary amino acid sequence and biochemical properties, the Ras superfamily is divided into different families: RAS (rat sarcoma), RHO (RAS homologs), RAB/ YPT (rat brain), ARF/SAR (ADP ribosylation factors), RAN (RAS-related nuclear) (649), ERA-like (Escherichia coli RASlike protein), SPG1/TEM1 (septum-promoting GTPase), and MIRO-like (mitochondrial Rho-GTPases) (650,651). In T. reesei, T. atroviride, and T. virens there are 105 genes encoding proteins of the RAS superfamily that are distributed in the different families mentioned above (36 genes in T. atroviride, 34 in T. reesei, and 35 in T. virens) ( Fig.…”

“…In A. niger, six GTPases were identified that participate in diverse mechanisms, with sometimes overlapping functions (651). RhoA (homolog to RHO1; TR_119871, TA_298907, and TV_89761) is involved in the establishment of cell polarity and viability, while RacA (homolog to RAC1; TR_47055, TA_300919, and TV_81671) contributes to asexual development, septation, and branching mechanisms (651). Asexual reproduction in A. niger is also regulated by RhoB (homolog to RHO2; TR_21294, TA_156444, and TV_71661), RhoD (homolog to RHO4; TR_53562, TA_143775, and TV_31417), and CftA (homolog to CDC42; TR_50335, TA_298630, and TV_111456).…”

“…On the other hand, CftA and RacA share overlapping functions, such as in cell polarization growth and cell wall biosynthesis (651), while RhoD has an essential role in septum formation. Deletion of rho3 (rhoC) in A. fumigatus caused no discernible phenotypic changes, and deletion of rho2 and rho4 (rhoB and rhoD, respectively) are related to cell wall stress and integrity (651). In A. fumigatus, Rho1 seems to be essential, and deletion of rho2 and rho4 alters the cell wall integrity (655).…”

“…RacA, the homolog of RAC1, plays a crucial role in activation of Nox family NADPH oxidases by their interaction in both animals and fungi (651,(669)(670)(671)(672). ROS play a significant role in morphogenesis and in fungal plant colonization (673,674).…”

Hyphal Morphology and Elongation Alterations in <i>Aspergillus nidulans</i> Provoked by the Diterpene Kaurenoic Acid

“…The n-hexane-soluble fraction was chromatographed over Si gel 60 (0.063 -0.200 mm) using vaccum liquid chromatography [14] with n-hexane and increasing amounts of ethyl acetate as eluents (250 mL each fraction). The second fraction (1730.0 mg) was washed with cold methanol, to afford pure KA (ent-kaur-16(17)-en-19-oic acid; 800.0 mg), which was identified by spectroscopic analysis and comparison with literature data [15].…”