Lack of the GTPase RHO-4 in Neurospora crassa causes a reduction in numbers and aberrant stabilization of microtubules at hyphal tips

Rasmussen, Carolyn G.; Morgenstein, Randy M.; Peck, S.; Glass, N. Louise

doi:10.1016/j.fgb.2008.02.006

Cited by 14 publications

(11 citation statements)

References 71 publications

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“…For example, genes encoding Rho GTPases (rho-2, NCU08683; rho-4, NCU03407) and a component of the Arp2/3 complex (sop2, NCU02781) showed maximal expression at the colony periphery in both N. crassa and A. niger. In N. crassa, mutations in rho-4 result in strains that lack septa, show slow growth, and have altered microtubule dynamics (52)(53)(54). Homologs of these proteins in animal cells are involved in the establishment of cell polarity and migration (43), and in plant cells, in proper polar cell expansion (3,25,81).…”

Section: Resultsmentioning

confidence: 99%

Dissecting Colony Development ofNeurospora crassaUsing mRNA Profiling and Comparative Genomics Approaches

Kasuga

Glass

2008

Eukaryot Cell

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Colony development, which includes hyphal extension, branching, anastomosis, and asexual sporulation, is a fundamental aspect of the life cycle of filamentous fungi; genetic mechanisms underlying these phenomena are poorly understood. We conducted transcriptional profiling during colony development of the model filamentous fungus Neurospora crassa, using 70-mer oligonucleotide microarrays. Relative mRNA expression levels were determined for six sections of defined age excised from a 27-h-old N. crassa colony. Functional category analysis showed that the expression of genes involved in cell membrane biosynthesis, polar growth, and cellular signaling was enriched at the periphery of the colony. The relative expression of genes involved in protein synthesis and energy production was enriched in the middle section of the colony, while sections of the colony undergoing asexual development (conidiogenesis) were enriched in expression of genes involved in protein/peptide degradation and unclassified proteins. A cross-examination of the N. crassa data set with a published data set of Aspergillus niger revealed shared patterns in the spatiotemporal regulation of gene orthologs during colony development. At present, less than 50% of genes in N. crassa have functional annotation, which imposes the chief limitation on data analysis. Using an evolutionary approach, we observed that the expression of phylogenetically conserved groups of genes was enriched in the middle section of an N. crassa colony whereas expression of genes unique to euascomycete species and of N. crassa orphan genes was enriched at the colony periphery and in the older, conidiating sections of a fungal colony.

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Section: Resultsmentioning

confidence: 99%

Dissecting Colony Development ofNeurospora crassaUsing mRNA Profiling and Comparative Genomics Approaches

Kasuga

Glass

2008

Eukaryot Cell

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“…Similarly, the PP2A-B= regulatory subunit Par1 is also a suppressor of the SIN; loss of par1 rescues defects of cdc11, cdc7, and spg1 mutants but is unable to rescue other SIN mutants (17,22). PP2A-Pab1-or PP2A-Par1p-mediated dephosphorylation likely inhibits the SIN activity, whereas Cdc11, Cdc7, and Spg1, which act as protein kinases and the GTPase, activate the SIN by phosphorylation, such that they may function antagonistically on the same substrate (65,66). Cdc11, Cdc7, and Spg1 are positive regulators that activate the SIN during septation and cytokinesis, whereas PP2A-Pab1 and PP2A-Par1 are negative regulators that deactivate the SIN (67).…”

Section: Discussionmentioning

confidence: 99%

Protein Phosphatase 2A (PP2A) Regulatory Subunits ParA and PabA Orchestrate Septation and Conidiation and Are Essential for PP2A Activity in Aspergillus nidulans

et al. 2014

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In recent decades, tremendous progress has been made in understanding serine/threonine protein phosphatase-dependent pathways, which are involved in the regulation of numerous processes, including secretion, cell motility, cell cycle, gene transcription, and cell metabolism (1-7). Serine/threonine phosphorylation, which is regulated by many kinases, can be reversed by a few phosphatases that are targeted to substrates via dozens of regulatory subunits (7,8). Currently, serine/threonine protein phosphatases (PPs) are grouped into two structurally distinct families: the PPP family (PP1, PP2A, and PP2B) and the PPM family (PP2C and pyruvate dehydrogenase phosphatase) (9). The PP2A heterotrimeric protein complex, which consists of a cascade of three subunits, namely, a catalytic subunit (C) and a structural subunit (A) associated with a third, hypothetically competitive and variable regulatory subunit (B), represents a highly conserved eukaryotic signal transduction system that is present in many organisms, from yeasts to humans. The activity of PP2A, along with its subcellular localization, is determined by B-family subunits (10-14). In mammals, PP2A is a major intracellular protein phosphatase that regulates multiple aspects of cell growth and metabolism (15). Therefore, PP2A is one of a few serine/threonine-specific phosphatases in the cell, and its complex structure and regulation system guarantee its various functions. The ability of this widely distributed heterotrimeric enzyme to function on a diverse array of substrates is largely controlled by the nature of its regulatory Bfamily subunits. Thus, multiple isoforms of the B-family regulatory subunit have been isolated from different organisms and grouped into three classes (B, B=, and BЉ) based on their structural similarities (16,17). The structural variations between these families support the hypothesis that the B-family regulatory subunit controls enzyme activity and specificity, such that different activities of PP2A and subcellular localization are determined by Bfamily regulatory subunits. However, one challenge we face is the potential for redundancy in the regulatory subunits' function of assigning PP2A function in mammals (18)(19)(20). Therefore, it is difficult to approach the function of PP2A globally by knocking out one of the regulatory subunits. In contrast, simple eukaryotic organisms, such as Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast), which contain only a few isoforms of B-family regulatory subunits, are excellent model systems for studying how B-family regulatory subunits function in regulating PP2A activity. In fission yeast, there is only one B subunit of PP2A-Pab1 and two of the B=-encoding genes (par1 and par2) (21-24). Moreover, mutational studies in fission yeast have demonstrated that B regulatory subunits play crucial roles during the processes of cytokinesis, cell morphogenesis, and cell wall synthesis (25,26). Similarly, in budding yeast, studies have shown that the B class has only one ...

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“…Based on the elongated phenotype of the nuclei observed in this study, along with the effect of benomyl on nuclear morphology in the mutants, we concluded that in N. crassa, impairment of DBF-2 confers a block in cell cycle progression, as has been shown in other systems. Stretched/broken nuclear morphology has been observed in the N. crassa rho-4 (encoding a monomeric GTPase) mutant, which also exhibits septation defects and cytoplasmic leakage (50,51). However, in addition to the morphological differences between rho-4 mutant nuclei and those observed in the dbf-2/ glycogen pathway mutants, in the rho-4 mutant the altered nuclear morphology was accompanied by a variance in nuclear distribution along the hyphal cell which was attributed to aberrant actin and microtubule cytoskeleton assembly shown to occur in that mutant (51).…”

Section: Discussionmentioning

confidence: 99%

“…Stretched/broken nuclear morphology has been observed in the N. crassa rho-4 (encoding a monomeric GTPase) mutant, which also exhibits septation defects and cytoplasmic leakage (50,51). However, in addition to the morphological differences between rho-4 mutant nuclei and those observed in the dbf-2/ glycogen pathway mutants, in the rho-4 mutant the altered nuclear morphology was accompanied by a variance in nuclear distribution along the hyphal cell which was attributed to aberrant actin and microtubule cytoskeleton assembly shown to occur in that mutant (51). The fact that we did not observe fragmented nuclei in the mutants studied here and that the benomyl treatment did not affect nuclear positioning is indicative that the elongated nuclear morphology was not due to a general defect in microtubule dynamics in these mutants.…”

Section: Discussionmentioning

confidence: 99%

The NDR Kinase DBF-2 Is Involved in Regulation of Mitosis, Conidial Development, and Glycogen Metabolism in Neurospora crassa

et al. 2010

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Neurospora crassa dbf-2 encodes an NDR (nuclear Dbf2-related) protein kinase, homologous to LATS1, a core component of the Hippo pathway. This pathway plays important roles in restraining cell proliferation and promoting apoptosis in differentiating cells. Here, we demonstrate that DBF-2 is involved in three fundamental processes in a filamentous fungus: cell cycle regulation, glycogen biosynthesis, and conidiation. DBF-2 is predominantly localized to the nucleus, and most (approximately 60%) dbf-2 null mutant nuclei are delayed in mitosis, indicating that DBF-2 activity is required for properly completing the cell cycle. The dbf-2 mutant exhibits reduced basal hyphal extension rates accompanied by a carbon/nitrogen ratio-dependent bursting of hyphal tips, vast glycogen leakage, defects in aerial hypha formation, and impairment of all three asexual conidiation pathways in N. crassa. Our findings also indicate that DBF-2 is essential for sexual reproduction in a filamentous fungus. Defects in other Hippo and glycogen metabolism pathway components (mob-1, ccr-4, mst-1, and gsk-3) share similar phenotypes such as mitotic delay and decreased CDC-2 (cell division cycle 2) protein levels, massive hyphal swellings, hyphal tip bursting, glycogen leakage, and impaired conidiation. We propose that DBF-2 functions as a link between Hippo and glycogen metabolism pathways.The nuclear Dbf2-related (NDR) protein kinases are essential components of signaling networks that control cellular processes in various organisms, including morphogenesis, exit from mitosis, cytokinesis, proliferation, differentiation, and apoptosis (23). Based on structural and functional conservation over long evolutionary distances, NDR kinases can be ascribed to one of two subgroups. One is comprised of mammalian NDR1/2, and the other is comprised of LATS1/2 (large tumor suppressor 1/2), as well as their orthologs and related kinases in different organisms.The filamentous fungus Neurospora crassa has two NDR kinases, which representative both subgroups. These are encoded by cot-1 (colonial temperature sensitive 1; NCU07296.3) and dbf-2 (NCU09071.3). While cot-1 (an ortholog of human NDR1/2) is involved in apical hyphal cell elongation and polarity (71), the role of dbf-2 (an ortholog of human LATS1/2) is yet unknown. Additionally, significant sequence similarities (52.8%) between the catalytic domains of DBF-2 and COT-1 raise the question whether their functions or localizations overlap. COT-1 has been localized to several intracellular compartments, including the cytoplasm and nucleus, and in association with the plasma membrane and various proteins (18,19,54). The role that COT-1 plays in the formation of branched cellular structures as well as its cellular localization has been suggested to be preserved throughout evolution (54, 77). Similar conservation may also exist among DBF-2 and its orthologs, of which human LATS1 and Drosophila melanogaster WTS/LATS are, by far, the most extensively analyzed (13,45,53,74,75).Both human LATS1 and NDR1 are wide...

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Lack of the GTPase RHO-4 in Neurospora crassa causes a reduction in numbers and aberrant stabilization of microtubules at hyphal tips

Cited by 14 publications

References 71 publications

Dissecting Colony Development ofNeurospora crassaUsing mRNA Profiling and Comparative Genomics Approaches

Dissecting Colony Development ofNeurospora crassaUsing mRNA Profiling and Comparative Genomics Approaches

Protein Phosphatase 2A (PP2A) Regulatory Subunits ParA and PabA Orchestrate Septation and Conidiation and Are Essential for PP2A Activity in Aspergillus nidulans

The NDR Kinase DBF-2 Is Involved in Regulation of Mitosis, Conidial Development, and Glycogen Metabolism in Neurospora crassa

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