Determination of cell polarity in germinated spores and hyphal tips of the filamentous ascomycete Ashbya gossypii requires a rhoGAP homolog

2000 ◽  
Vol 113 (9) ◽  
pp. 1611-1621 ◽  
Author(s):  
J. Wendland ◽  
P. Philippsen
Keyword(s):  
Filamentous Fungi ◽  
Cell Polarity ◽  
Germ Cells ◽  
Rho Gtpases ◽  
Protein Domain ◽  
Ashbya Gossypii ◽  
Cortical Actin ◽  
Filamentous Ascomycete ◽  
Bud Emergence ◽  
Hyphal Tip

In the filamentous ascomycete Ashbya gossypii, like in other filamentous fungi onset of growth in dormant spores occurs as an isotropic growth phase generating spherical germ cells. Thereafter, a switch to polarized growth results in the formation of the first hyphal tip. The initial steps of hyphal tip formation in filamentous fungi, therefore, resemble processes taking place prior to and during bud emergence of unicellular yeast-like fungi. We investigated whether phenotypic similarities between these distinct events extended to the molecular level. To this end we isolated and characterized the A. gossypii homolog of the Saccharomyces cerevisiae BEM2 gene which is part of a network of rho-GTPases and their regulators required for bud emergence and bud growth in yeast. Here we show that the AgBem2 protein contains a GAP- (GTPase activating protein) domain for rho-like GTPases at its carboxy terminus, and that this part of AgBem2p is required for complementation of an Agbem2 null strain. Germination of spores resulted in enlarged Agbem2 germ cells that were unable to generate the bipolar branching pattern found in wild-type germ cells. In addition, mutant hyphae were swollen due to defects in polarized cell growth indicated by the delocalized distribution of chitin and cortical actin patches. Surprisingly, the complete loss of cell polarity which lead to spherical hyphal tips was overcome by the establishment of new cell polarities and the formation of multiple new hyphal tips. In conclusion these results and other findings demonstrate that establishment of cell polarity, maintenance of cell polarity, and polarized hyphal growth in filamentous fungi require members of Ρ-GTPase modules.

scholarly journals Cell Polarity and Hyphal Morphogenesis Are Controlled by Multiple Rho-Protein Modules in the Filamentous Ascomycete Ashbya gossypii

Genetics ◽  
2001 ◽  
Vol 157 (2) ◽  
pp. 601-610
Author(s):  
J Wendland ◽  
P Philippsen
Keyword(s):  
Cell Growth ◽  
Filamentous Fungi ◽  
Cell Polarity ◽  
Hyphal Growth ◽  
Ashbya Gossypii ◽  
Rho Proteins ◽  
Filamentous Ascomycete ◽  
Polarized Cell Growth ◽  
Rho Protein ◽  
Protein Modules

Abstract Polarized cell growth requires a polarized organization of the actin cytoskeleton. Small GTP-binding proteins of the Rho-family have been shown to be involved in the regulation of actin polarization as well as other processes. Hyphal growth in filamentous fungi represents an ideal model to investigate mechanisms involved in generating cell polarity and establishing polarized cell growth. Since a potential role of Rho-proteins has not been studied so far in filamentous fungi we isolated and characterized the Ashbya gossypii homologs of the Saccharomyces cerevisiae CDC42, CDC24, RHO1, and RHO3 genes. The AgCDC42 and AgCDC24 genes can both complement conditional mutations in the S. cerevisiae CDC42 and CDC24 genes and both proteins are required for the establishment of actin polarization in A. gossypii germ cells. Agrho1 mutants show a cell lysis phenotype. Null mutant strains of Agrho3 show periodic swelling of hyphal tips that is overcome by repolarization and polar hyphal growth in a manner resembling the germination pattern of spores. Thus different Rho-protein modules are required for distinct steps during polarized hyphal growth of A. gossypii.

A PAK-like protein kinase is required for maturation of young hyphae and septation in the filamentous ascomycete Ashbya gossypii

2000 ◽  
Vol 113 (24) ◽  
pp. 4563-4575 ◽  
Author(s):  
Y. Ayad-Durieux ◽  
P. Knechtle ◽  
S. Goff ◽  
F. Dietrich ◽  
P. Philippsen
Keyword(s):  
Protein Kinase ◽  
Filamentous Fungi ◽  
Fluorescent Protein ◽  
Ashbya Gossypii ◽  
Polarized Growth ◽  
Fast Growing ◽  
Filamentous Ascomycete ◽  
Green Fluorescent ◽  
Actin Cables ◽  
Developmental Switch

Filamentous fungi grow by hyphal extension, which is an extreme example of polarized growth. In contrast to yeast species, where polarized growth of the tip of an emerging bud is temporally limited, filamentous fungi exhibit constitutive polarized growth of the hyphal tip. In many fungi, including Ashbya gossypii, polarized growth is reinforced by a process called hyphal maturation. Hyphal maturation refers to the developmental switch from slow-growing hyphae of young mycelium to fast-growing hyphae of mature mycelium. This process is essential for efficient expansion of mycelium. We report for the first time on the identification and characterization of a fungal gene important for hyphal maturation. This novel A. gossypii gene encodes a presumptive PAK (p21-activated kinase)-like kinase. Its closest homolog is the S. cerevisiae Cla4 protein kinase; the A. gossypii protein is therefore called AgCla4p. Agcla4 deletion strains are no longer able to perform the developmental switch from young to mature hyphae, and GFP (green fluorescent protein)-tagged AgCla4p localizes with much higher frequency in mature hyphal tips than in young hyphal tips. Both results support the importance of AgCla4p in hyphal maturation. AgCla4p is also required for septation, indicated by the inability of Agcla4 deletion strains to properly form actin rings and chitin rings. Despite the requirement of AgCla4p for the development of fast-growing hyphae, AgCla4p is not necessary for actin polarization per se, because tips enriched in cortical patches and hyphae with a fully developed network of actin cables can be seen in Agcla4 deletion strains. The possibility that AgCla4p may be involved in regulatory mechanisms that control the dynamics of the actin patches and/or actin cables is discussed.

scholarly journals A Ras-like GTPase Is Involved in Hyphal Growth Guidance in the Filamentous Fungus Ashbya gossypii

2004 ◽  
Vol 15 (10) ◽  
pp. 4622-4632 ◽  
Author(s):  
Yasmina Bauer ◽  
Philipp Knechtle ◽  
Jürgen Wendland ◽  
Hanspeter Helfer ◽  
Peter Philippsen
Keyword(s):  
Fluorescent Protein ◽  
Hyphal Growth ◽  
Time Lapse ◽  
Growth Direction ◽  
Ashbya Gossypii ◽  
Growth Guidance ◽  
Characteristic Features ◽  
Green Fluorescent ◽  
Hyphal Tip

Characteristic features of morphogenesis in filamentous fungi are sustained polar growth at tips of hyphae and frequent initiation of novel growth sites (branches) along the extending hyphae. We have begun to study regulation of this process on the molecular level by using the model fungus Ashbya gossypii. We found that the A. gossypii Ras-like GTPase Rsr1p/Bud1p localizes to the tip region and that it is involved in apical polarization of the actin cytoskeleton, a determinant of growth direction. In the absence of RSR1/BUD1, hyphal growth was severely slowed down due to frequent phases of pausing of growth at the hyphal tip. During pausing events a hyphal tip marker, encoded by the polarisome component AgSPA2, disappeared from the tip as was shown by in vivo time-lapse fluorescence microscopy of green fluorescent protein-labeled AgSpa2p. Reoccurrence of AgSpa2p was required for the resumption of hyphal growth. In the Agrsr1/bud1Δ deletion mutant, resumption of growth occurred at the hyphal tip in a frequently uncoordinated manner to the previous axis of polarity. Additionally, hyphal filaments in the mutant developed aberrant branching sites by mislocalizing AgSpa2p thus distorting hyphal morphology. These results define AgRsr1p/Bud1p as a key regulator of hyphal growth guidance.

scholarly journals The BNIP-2 and Cdc42GAP Homology (BCH) Domain of p50RhoGAP/Cdc42GAP Sequesters RhoA from Inactivation by the Adjacent GTPase-activating Protein Domain

2010 ◽  
Vol 21 (18) ◽  
pp. 3232-3246 ◽  
Author(s):  
Yi Ting Zhou ◽  
Li Li Chew ◽  
Sheng-cai Lin ◽  
Boon Chuan Low
Keyword(s):  
Rho Gtpases ◽  
Rho Gtpase ◽  
Intramolecular Interaction ◽  
Complete Loss ◽  
Protein Domain ◽  
Binding Motif ◽  
Gtpase Activating Protein ◽  
New Paradigm ◽  
Cell Rounding ◽  
In Cis

The BNIP-2 and Cdc42GAP homology (BCH) domain is a novel regulator for Rho GTPases, but its impact on p50-Rho GTPase-activating protein (p50RhoGAP or Cdc42GAP) in cells remains elusive. Here we show that deletion of the BCH domain from p50RhoGAP enhanced its GAP activity and caused drastic cell rounding. Introducing constitutively active RhoA or inactivating GAP domain blocked such effect, whereas replacing the BCH domain with endosome-targeting SNX3 excluded requirement of endosomal localization in regulating the GAP activity. Substitution with homologous BCH domain from Schizosaccharomyces pombe, which does not bind mammalian RhoA, also led to complete loss of suppression. Interestingly, the p50RhoGAP BCH domain only targeted RhoA, but not Cdc42 or Rac1, and it was unable to distinguish between GDP and the GTP-bound form of RhoA. Further mutagenesis revealed a RhoA-binding motif (residues 85-120), which when deleted, significantly reduced BCH inhibition on GAP-mediated cell rounding, whereas its full suppression also required an intramolecular interaction motif (residues 169-197). Therefore, BCH domain serves as a local modulator in cis to sequester RhoA from inactivation by the adjacent GAP domain, adding to a new paradigm for regulating p50RhoGAP signaling.

scholarly journals Control of cortical cytoskeleton-membrane interaction by RhoA regulates peripheral nerve myelination

2021 ◽  
Author(s):  
Ana I. Seixas ◽  
Miguel R. G. Morais ◽  
Cord Brakebusch ◽  
Jo&atildeo B. Relvas
Keyword(s):  
Schwann Cell ◽  
Schwann Cells ◽  
Rho Gtpases ◽  
Intracellular Signaling ◽  
Cell Interaction ◽  
Actin Dynamics ◽  
Specific Gene ◽  
Cortical Actin ◽  
Membrane Attachment ◽  
Cytoskeleton Dynamics

Bidirectional transmission of mechanical and biochemical signals is integral to cell-environment communication and underlies the function of Schwann cells, the myelinating glia of the peripheral nervous system. As major integrators of "outside-in" signaling, Rho GTPases link actin cytoskeleton dynamics with cellular architecture to regulate adhesion and cell deformation. Using Schwann cell-specific gene inactivation, we discovered that RhoA promotes the initiation of myelination, axonal wrapping and axial spreading of Schwann cells, and is later required to restrict myelin growth in peripheral nerves. These effects are mediated by modulation of actomyosin contractility, actin dynamics and cortical actin-membrane attachment, which collectively couple tensional forces to intracellular signaling that regulate axon-Schwann cell interaction and myelin synthesis. This work establishes RhoA as an intrinsic regulator of a biomechanical response that controls the switch of Schwann cells towards the myelinating and the homeostatic states.

scholarly journals Regulation of exit from mitosis in multinucleate Ashbya gossypii cells relies on a minimal network of genes

2011 ◽  
Vol 22 (17) ◽  
pp. 3081-3093 ◽  
Author(s):  
Mark R. Finlayson ◽  
A. Katrin Helfer-Hungerbühler ◽  
Peter Philippsen
Keyword(s):  
Regulatory Networks ◽  
Temporal Constraints ◽  
Ashbya Gossypii ◽  
Metaphase Arrest ◽  
Minimal Network ◽  
Filamentous Ascomycete ◽  
Low Degree ◽  
Mitotic Exit Network ◽  
Early Anaphase ◽  
Core Components

In Saccharomyces cerevisiae, mitosis is coupled to cell division by the action of the Cdc fourteen early anaphase release (FEAR) and mitotic exit network (MEN) regulatory networks, which mediate exit from mitosis by activation of the phosphatase Cdc14. The closely related filamentous ascomycete Ashbya gossypii provides a unique cellular setting to study the evolution of these networks. Within its multinucleate hyphae, nuclei are free to divide without the spatial and temporal constraints described for budding yeast. To investigate how this highly conserved system has adapted to these circumstances, we constructed a series of mutants lacking homologues of core components of MEN and FEAR and monitored phenomena such as progression through mitosis and Cdc14 activation. MEN homologues in A. gossypii were shown to have diverged from their anticipated role in Cdc14 release and exit from mitosis. We observed defects in septation, as well as a partial metaphase arrest, in Agtem1Δ, Agcdc15Δ, Agdbf2/dbf20Δ, and Agmob1Δ. A. gossypii homologues of the FEAR network, on the other hand, have a conserved and more pronounced role in regulation of the M/G1 transition. Agcdc55Δ mutants are unable to sequester AgCdc14 throughout interphase. We propose a reduced model of the networks described in yeast, with a low degree of functional redundancy, convenient for further investigations into these networks.

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

2011 ◽  
Vol 79 (5) ◽  
pp. 1151-1167 ◽  
Author(s):  
Min Jin Kwon ◽  
Mark Arentshorst ◽  
Eelke D. Roos ◽  
Cees A. M. J. J. van den Hondel ◽  
Vera Meyer ◽  
...  
Keyword(s):  
Aspergillus Niger ◽  
Filamentous Fungi ◽  
Rho Gtpases ◽  
Functional Characterization ◽  
Rho Proteins

scholarly journals The TRE17 Oncogene Encodes a Component of a Novel Effector Pathway for Rho GTPases Cdc42 and Rac1 and Stimulates Actin Remodeling

2003 ◽  
Vol 23 (6) ◽  
pp. 2151-2161 ◽  
Author(s):  
Jeffrey M. Masuda-Robens ◽  
Sara N. Kutney ◽  
Hongwei Qi ◽  
Margaret M. Chou
Keyword(s):  
Plasma Membrane ◽  
Rho Gtpases ◽  
Actin Polymerization ◽  
Dependent Manner ◽  
Cortical Actin ◽  
Actin Remodeling ◽  
Membrane Recruitment ◽  
Truncation Mutant ◽  
Effector Pathway

ABSTRACT The Rho family GTPases Cdc42 and Rac1 play fundamental roles in transformation and actin remodeling. Here, we demonstrate that the TRE17 oncogene encodes a component of a novel effector pathway for these GTPases. TRE17 coprecipitated specifically with the active forms of Cdc42 and Rac1 in vivo. Furthermore, the subcellular localization of TRE17 was dramatically regulated by these GTPases and mitogens. Under serum-starved conditions, TRE17 localized predominantly to filamentous structures within the cell. Epidermal growth factor (EGF) induced relocalization of TRE17 to the plasma membrane in a Cdc42-/Rac1-dependent manner. Coexpression of activated alleles of Cdc42 or Rac1 also caused complete redistribution of TRE17 to the plasma membrane, where it partially colocalized with the GTPases in filopodia and ruffles, respectively. Membrane recruitment of TRE17 by EGF or the GTPases was dependent on actin polymerization. Finally, we found that a C-terminal truncation mutant of TRE17 induced the accumulation of cortical actin, mimicking the effects of activated Cdc42. Together, these results identify TRE17 as part of a novel effector complex for Cdc42 and Rac1, potentially contributing to their effects on actin remodeling. The present study provides insights into the regulation and cellular function of this previously uncharacterized oncogene.

scholarly journals The Ashbya gossypii fimbrin SAC6 is required for fast polarized hyphal tip growth and endocytosis

2011 ◽  
Vol 166 (3) ◽  
pp. 137-145 ◽  
Author(s):  
Sigyn Jorde ◽  
Andrea Walther ◽  
Jürgen Wendland
Keyword(s):  
Tip Growth ◽  
Ashbya Gossypii ◽  
Hyphal Tip Growth ◽  
Hyphal Tip
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