Flunarizine affects F-actin pattern in Mucor rouxii germlings

1994 ◽  
Vol 40 (9) ◽  
pp. 730-735 ◽  
Author(s):  
Jiří Hašek ◽  
Eva Streiblová
Keyword(s):  
Plasma Membrane ◽  
Germ Tube ◽  
Tip Growth ◽  
Apical Growth ◽  
Mucor Rouxii ◽  
Yeast Form ◽  
Tube Emergence

The effect of the Ca2+ antagonist flunarizine on Mucor rouxii germlings was analyzed. In sporangiospores cultivated in aerobiosis, the drug blocked germ tube emergence, but enlarged spheres continued to grow isodiametrically. The formation of broad protuberances and irregular buds was observed. Calcofluor staining indicated aberrations on the surface of the flunarizine-treated cells. This was not a simple switch to the yeast form, since labelling with rhodarmine-tagged phalloidin revealed various ring-shaped structures of F-actin, indicating furrowing of the plasma membrane. Flunarizine apparently did not interfere with nuclear divisions or with the integrity of microtubules. In young germlings, flunarizine treatment induced cessation of tip growth and promoted septation associated with the rearrangement of F-actin. Apical growth was reestablished after either the drug was removed or an excess of Ca2+ ions was added to the medium. The results suggest that tip-related arrangement of F-actin requires flunarizine-sensitive Ca2+ entry.Key words: Mucor rouxii, F-actin, Ca2+ antagonist, flunarizine, germination.

scholarly journals Endocytic Machinery Protein SlaB Is Dispensable for Polarity Establishment but Necessary for Polarity Maintenance in Hyphal Tip Cells of Aspergillus nidulans

2010 ◽  
Vol 9 (10) ◽  
pp. 1504-1518 ◽  
Author(s):  
América Hervás-Aguilar ◽  
Miguel A. Peñalva
Keyword(s):  
Plasma Membrane ◽  
Aspergillus Nidulans ◽  
Germ Tube ◽  
Fluorescent Protein ◽  
Hyphal Growth ◽  
Gene Replacement ◽  
Actin Binding ◽  
Nitrate Medium ◽  
Content Type ◽  
Tube Emergence

ABSTRACT The Aspergillus nidulans endocytic internalization protein SlaB is essential, in agreement with the key role in apical extension attributed to endocytosis. We constructed, by gene replacement, a nitrate-inducible, ammonium-repressible slaB1 allele for conditional SlaB expression. Video microscopy showed that repressed slaB1 cells are able to establish but unable to maintain a stable polarity axis, arresting growth with budding-yeast-like morphology shortly after initially normal germ tube emergence. Using green fluorescent protein (GFP)-tagged secretory v-SNARE SynA, which continuously recycles to the plasma membrane after being efficiently endocytosed, we establish that SlaB is crucial for endocytosis, although it is dispensable for the anterograde traffic of SynA and of the t-SNARE Pep12 to the plasma and vacuolar membrane, respectively. By confocal microscopy, repressed slaB1 germlings show deep plasma membrane invaginations. Ammonium-to-nitrate medium shift experiments demonstrated reversibility of the null polarity maintenance phenotype and correlation of normal apical extension with resumption of SynA endocytosis. In contrast, SlaB downregulation in hyphae that had progressed far beyond germ tube emergence led to marked polarity maintenance defects correlating with deficient SynA endocytosis. Thus, the strict correlation between abolishment of endocytosis and disability of polarity maintenance that we report here supports the view that hyphal growth requires coupling of secretion and endocytosis. However, downregulated slaB1 cells form F-actin clumps containing the actin-binding protein AbpA, and thus F-actin misregulation cannot be completely disregarded as a possible contributor to defective apical extension. Latrunculin B treatment of SlaB-downregulated tips reduced the formation of AbpA clumps without promoting growth and revealed the formation of cortical “comets” of AbpA.

The Candida albicans pH-regulated KER1 gene encodes a lysine/glutamic-acid-rich plasma-membrane protein that is involved in cell aggregation

Microbiology ◽  
2004 ◽  
Vol 150 (8) ◽  
pp. 2641-2651 ◽  
Author(s):  
Amparo Galán ◽  
Manuel Casanova ◽  
Amelia Murgui ◽  
Donna M. MacCallum ◽  
Frank C. Odds ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Plasma Membrane ◽  
Glutamic Acid ◽  
Germ Tube ◽  
Cell Surface Hydrophobicity ◽  
Cell Aggregation ◽  
Surface Hydrophobicity ◽  
Amino Acid Sequences ◽  
Calcofluor White

Immunoscreening of a Candida albicans cDNA library with a polyclonal germ-tube-specific antibody (pAb anti-gt) resulted in the isolation of a gene encoding a lysine/glutamic-acid-rich protein, which was consequently designated KER1. The nucleotide and deduced amino acid sequences of this gene displayed no significant homology with any other known sequence. KER1 encodes a 134 kDa lysine (14·5 %)/glutamic acid (16·7 %) protein (Ker1p) that contains two potential transmembrane segments. KER1 was expressed in a pH-conditional manner, with maximal expression at alkaline pH and lower expression at pH 4·0, and was regulated by RIM101. A Δker1/Δker1 null mutant grew normally but was hyperflocculant under germ-tube-inducing conditions, yet this behaviour was also observed in stationary-phase cells grown under other incubation conditions. Western blotting analysis of different subcellular fractions, using as a probe a monospecific polyclonal antibody raised against a highly antigenic domain of Ker1p (pAb anti-Ker1p), revealed the presence of a 134 kDa band in the purified plasma-membrane fraction from the wild-type strain that was absent in the homologous preparation from Δker1/Δker1 mutant. The pattern of cell-wall protein and mannoprotein species released by digestion with β-glucanases, reactive towards pAbs anti-gt and anti-Ker1p, as well as against concanavalin A, was also different in the Δker1/Δker1 mutant. Mutant strains also displayed an increased cell-surface hydrophobicity and sensitivity to Congo red and Calcofluor white. Overall, these findings indicate that the mutant strain was affected in cell-wall composition and/or structure. The fact that the ker1 mutant had attenuated virulence in systemic mouse infections suggests that this surface protein is also important in host–fungus interactions.

Carbon dioxide induces endotrophic germ tube formation in Candida albicans

1990 ◽  
Vol 36 (4) ◽  
pp. 249-253 ◽  
Author(s):  
Ruth C. Mock ◽  
Jordan H. Pollack ◽  
Tadayo Hashimoto
Keyword(s):  
Carbon Dioxide ◽  
Candida Albicans ◽  
Sodium Bicarbonate ◽  
Key Words ◽  
Germ Tube ◽  
Tube Formation ◽  
Chemical Inducers ◽  
Tube Emergence ◽  
Ph Range ◽  
Germ Tubes

Candida albicans formed germ tubes when exposed to air containing 5 to 15% carbon dioxide (CO2). The CO2-mediated germ tube formation occurred optimally at 37 °C in a pH range of 5.5 to 6.5. No germ tubes were produced at 25 °C, even when the optimal concentration of CO2 (10%) was present in the environment. The requirement of CO2 for germ tube formation could be partially substituted by sodium bicarbonate but not by N2. Carbon dioxide was required to be present throughout the entire course of germ tube emergence suggesting that its role is not limited to an initial triggering of morphogenic change. We suggest that carbon dioxide may be a common effector responsible for the germ tube promoting activity of certain chemical inducers for C. albicans. Key words: Candida albican germ tubes, CO2-induced germ tube formation, endotrophic germ tube formation.

Two divergent plasma membrane syntaxin-like SNAREs, nsyn1 and nsyn2, contribute to hyphal tip growth and other developmental processes in Neurospora crassa

2003 ◽  
Vol 40 (3) ◽  
pp. 271-286 ◽  
Author(s):  
Gagan D Gupta ◽  
Stephen J Free ◽  
Natalia N Levina ◽  
Sirkka Keränen ◽  
I.Brent Heath
Keyword(s):  
Plasma Membrane ◽  
Neurospora Crassa ◽  
Tip Growth ◽  
Developmental Processes ◽  
Hyphal Tip Growth ◽  
Hyphal Tip

Ultrastructure of freeze-substituted appressoria produced by aeciospore germlings of the rust fungus Arthuriomyces peckianus

1991 ◽  
Vol 69 (8) ◽  
pp. 1655-1665 ◽  
Author(s):  
E. C. Swann ◽  
C. W. Mims
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Key Words ◽  
Direct Contact ◽  
Germ Tube ◽  
Rust Fungus ◽  
Mitotic Division ◽  
Extracellular Material ◽  
Dialysis Membrane ◽  
Fungus Infection

Aeciospores of Arthuriomyces peckianus germinated readily on moist strips of dialysis membrane and developing appressoria were observed within 3 to 6 h after spores were deposited on membranes. A single germ tube typically emerged from each binucleate spore and grew until its tip contacted the dialysis membrane. The germ tube tip was then transformed into a swollen appressorium that adhered tightly to the membrane, apparently as a result of an extracellular material that surrounded the appressorium base. Virtually all the spore cytoplasm and both nuclei moved into the germ tube and developing appressorium. Following a synchronous mitotic division of the two nuclei, a septum formed to delimit the now tetranucleate appressorium from the germ tube. As the appressorium matured, an apparently wall-less region developed in the central portion of the appressorium appressed against the dialysis membrane. In this region the fungus plasma membrane appeared to make direct contact with the underlying dialysis membrane. A funnel-like or cone-like structure referred to as the appressorial cone then developed in the wall-less region. The appressorial cone extended up into the cytoplasm of the appressorium and was lined by the fungal plasma membrane. Numerous branched elaborations of the plasma membrane were associated with the inner portion of the cone. Key words: rust fungus, infection structures, electron microscopy.

Pattern of protein degradation during germination of Streptomyces antibioticus spores

1983 ◽  
Vol 29 (6) ◽  
pp. 637-643 ◽  
Author(s):  
J. A. Guijarro ◽  
J. E. Suarez ◽  
J. A. Salas ◽  
C. Hardisson
Keyword(s):  
Protein Degradation ◽  
Protease Activity ◽  
Germ Tube ◽  
Synthetic Medium ◽  
Distilled Water ◽  
Streptomyces Antibioticus ◽  
Constant Rate ◽  
Degradation Pattern ◽  
Tube Emergence ◽  
Inhibition Studies

The pattern of protein degradation during germination of Streptomyces antibioticus spores was studied by the pulse and chase technique. Two different protein fractions were found. First, a fraction of the proteins synthesized during the darkening process (20–30%) was quickly degraded in the 30 min following the labelling period. This rapid protein degradation was partially inhibited by protease inhibitors: p-chloromercuribenzoic acid, phenylmethylsulphonylfluoride, and o-phenanthroline. Second, the remaining 70–80% and the entire protein population formed during spore swelling and germ tube emergence were degraded with a lower and constant rate (3.3–6.0%/h). A stable mRNA fraction of the dormant spores was translated upon incubation of the spores in a minimal synthetic medium (MSM) or in distilled water. However, the degradation of these proteins did not occur unless the spores were then incubated in the MSM. A strong correlation between the degradation pattern of these proteins and that of those quickly degraded at the beginning of germination was observed. Protease activity in cell-free extracts of dormant spores was detected. Inhibition studies suggest the presence of serine, thiol, and metalloproteases. The protease activity, using casein as substrate, remained constant during the darkening process and started to increase progressively from the beginning of spore swelling.

Polyadenylate-containing RNA in dormant and germinating Botryodiplodia theobromae pycnidiospores

1979 ◽  
Vol 25 (3) ◽  
pp. 375-379 ◽  
Author(s):  
James L. Van Etten ◽  
Carroll D. Rawn
Keyword(s):  
Germ Tube ◽  
De Novo ◽  
Segment Length ◽  
Botryodiplodia Theobromae ◽  
Rna Molecules ◽  
Polyuridylic Acid ◽  
Tube Emergence ◽  
Dormant Spore ◽  
Average Size

Hybridization of [3H]polyuridylic acid to RNA isolated from Botryodiplodia theobromae pycnidiospores yielded an estimate of about 6.25 × 105 polyadenylate-containing RNA (poly A(+) RNA) molecules per dormant spore. The number increased about fourfold by the time of germ tube emergence at 3 h. The average size of this presumed mRNA was about 4.1 × 105 daltons (1275 nucleotides), with an average polyadenylate segment length of 26 nucleotides. Neither of these values changed significantly during germination. The earliest detectable (first 30 min of germination) de novo synthesized mRNA's were rapidly incorporated into polyribosomes. This newly synthesized, presumably functional, mRNA was composed of both poly A(+) RNA and polyadenylate-lacking RNA. The average sizesof the two polyribosomal mRNA subpopulations and the total poly A(+) RNA population were identical.

scholarly journals AtRBOHC/RHD2 vesicular delivery to the apical plasma membrane domain during root hair development

2021 ◽  
Author(s):  
Lenka Kuběnová ◽  
Michaela Tichá ◽  
Jozef Šamaj ◽  
Miroslav Ovečka
Keyword(s):  
Plasma Membrane ◽  
Quantitative Analysis ◽  
Root Hair ◽  
Tip Growth ◽  
Root Hairs ◽  
Apical Plasma Membrane ◽  
Loss Of Function ◽  
Short Root ◽  
Early Endosomes ◽  
Root Hair Initiation

AbstractArabidopsis root hairs develop as long tubular extensions from the rootward pole of trichoblasts and exert polarized tip growth. The establishment and maintenance of root hair polarity is a complex process involving the local apical production of reactive oxygen species (ROS) generated by NADPH oxidase RESPIRATORY BURST OXIDASE HOMOLOG PROTEIN C/ROOT HAIR DEFECTIVE 2 (AtRBOHC/RHD2). It has been shown that loss-of-function rhd2 mutants have short root hairs that are unable to elongate by tip growth, and this phenotype was fully complemented by GFP-RHD2 expressed under the RHD2 promoter. However, the spatiotemporal mechanism of AtRBOHC/RHD2 subcellular redistribution and delivery to the plasma membrane (PM) during root hair initiation and tip growth are still unclear. Here, we used advanced microscopy for detailed qualitative and quantitative analysis of vesicular compartments containing GFP-RHD2 and characterization of their movements in developing bulges and growing root hairs. These compartments, identified by an independent marker such as the trans-Golgi network (TGN), deliver GFP-RHD2 to the apical PM domain, the extent of which correlates with the stage of root hair formation. Movements of TGN/early endosomes, but not late endosomes, were affected in the bulging domains of the rhd2-1 mutant. Finally, we reveal that accumulation in the growing tip, docking, and incorporation of TGN compartments containing GFP-RHD2 to the apical PM of root hairs requires structural sterols. These results help clarify the mechanism of polarized AtRBOHC/RHD2 targeting, maintenance, and recycling at the apical PM domain, coordinated with different developmental stages of root hair initiation and growth.One-sentence summaryAdvanced microscopy and quantitative analysis of vesicular TGN compartments revealed that delivering GFP-RHD2 to the apical plasma membrane domains of developing bulges and growing root hairs requires structural sterols.

scholarly journals Defects in Assembly of the Extracellular Matrix Are Responsible for Altered Morphogenesis of a Candida albicans phr1 Mutant

1998 ◽  
Vol 180 (1) ◽  
pp. 163-166 ◽  
Author(s):  
Laura Popolo ◽  
Marina Vai
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Parental Strain ◽  
Germ Tube ◽  
Null Mutant ◽  
Yeast Form ◽  
Insoluble Glucan ◽  
Cross Linker ◽  
External Ph

ABSTRACT Analysis of Candida albicans cells using antibodies directed against Gas1p/Ggp1p, Saccharomyces cerevisiaehomolog of Phr1p, revealed that Phr1p is a glycoprotein of about 88 kDa whose accumulation increases with the rise of external pH. This polypeptide is present both in the yeast form and during germ tube induction. In the Phr1− cells at pH 8 the solubility of glucans in alkali is greatly affected. In the parental strain the alkali-soluble/-insoluble glucan ratio shows a 50% decrease at pH 8 with respect to pH 4.5, whereas in the null mutant it is unchanged, indicating the lack of a polymer cross-linker activity induced by the rise of pH. The mutant has a sixfold increase in chitin level and is hypersensitive to calcofluor. Consistently with a role of chitin in strengthening the cell wall, Phr1− cells are more sensitive to nikkomycin Z than the parental strain.

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