RESUMO
Current models that describe the extension of fungal hyphae and development of a mycelium either do not describe the role of vesicles in hyphal extension or do not correctly describe the experimentally observed profile for distribution of vesicles along the hypha. The present work uses the n-tanks-in-series approach to develop a model for hyphal extension that describes the intracellular transport of nutrient to a sub-apical zone where vesicles are formed and then transported to the tip, where tip extension occurs. The model was calibrated using experimental data from the literature for the extension of reproductive aerial hyphae of three different fungi, and was able to describe different profiles involving acceleration and deceleration of the extension rate. A sensitivity analysis showed that the supply of nutrient to the sub-apical vesicle-producing zone is a key factor influencing the rate of extension of the hypha. Although this model was used to describe the extension of a single reproductive aerial hypha, the use of the n-tanks-in-series approach to representing the hypha means that the model has the flexibility to be extended to describe the growth of other types of hyphae and the branching of hyphae to form a complete mycelium.
Assuntos
Aspergillus/crescimento & desenvolvimento , Hifas/crescimento & desenvolvimento , Modelos Estatísticos , Phycomyces/crescimento & desenvolvimento , Rhizopus/crescimento & desenvolvimento , Aspergillus/metabolismo , Transporte Biológico , Simulação por Computador , Hifas/metabolismo , Maltose/metabolismo , Modelos Biológicos , Phycomyces/metabolismo , Rhizopus/metabolismo , Vesículas Transportadoras/metabolismoRESUMO
The effects of the Ca(2+)/H(+) exchanger A23187 and the K(+)/H(+) exchanger nigericin, the electrogenic membrane-potential depleters valinomycin and CCCP, and the calcium channel blockers ruthenium red, nifedipine, and nitrendipine on the apical growth of Phycomyces blakesleeanus were analyzed. While all of the compounds inhibited the growth of germlings in liquid medium, the Ca(2+) channel blockers were the least effective. Chitin synthesis in vivo was also sensitive to the inhibitors; here again, the calcium channel blockers were less efficient, and their effect occurred after a lag phase, in contrast to the electroneutral ionophores whose effects were immediate. The ionophores rapidly inhibited protein secretion, and reduced the number of secretory vesicles and chitosomes in the hyphal apex of P. blakesleeanus. The results suggest that not only tip-to-base calcium gradients but also transmembrane ionic gradients and membrane potential have a role in the apical growth of P. blakesleeanus. They are probably involved in the formation, migration, and/or fusion with the plasmalemma of secretory vesicles and chitosomes.
Assuntos
Bloqueadores dos Canais de Cálcio/farmacologia , Ionóforos/farmacologia , Phycomyces/efeitos dos fármacos , Phycomyces/crescimento & desenvolvimento , Calcimicina/farmacologia , Metabolismo dos Carboidratos , Carbonil Cianeto m-Clorofenil Hidrazona/farmacologia , Quitina/biossíntese , Hifas/efeitos dos fármacos , Hifas/ultraestrutura , Nifedipino/farmacologia , Nigericina/farmacologia , Nitrendipino/farmacologia , Phycomyces/metabolismo , Phycomyces/ultraestrutura , Transporte Proteico/efeitos dos fármacos , Rutênio Vermelho/farmacologia , Vesículas Secretórias/efeitos dos fármacos , Valinomicina/farmacologia , beta-Frutofuranosidase/metabolismoRESUMO
A DNA sequence homologous to a G alpha s DNA probe, and the corresponding G alpha s protein (stimulatory alpha-subunit of GTP-binding protein) were detected in Phycomyces blakesleeanus. The protein was demonstrated in membrane fractions of dormant spores of this fungus using three different experimental approaches. Photoaffinity-labelling experiments with [alpha-32P]GTP of the membrane fraction revealed two bands, of 56 and 32 kDa. The 56 kDa GTP-binding protein was detected by this method in all the stages of early development and growth investigated. Also, a spore protein of 56 kDa was ADP-ribosylated by cholera toxin, and a 56 kDa protein was detected by Western blotting with a specific antibody against mammalian G alpha s. These results indicate that G alpha s (56 kDa) is present in dormant spores of P. blakesleeanus. Using the ADP-ribosylation and Western blotting assays, G alpha s was detected during all stages of spore germination before the hyphae became highly branched, but it was not detected in the branched hyphae that formed 18 h after the initiation of spore germination. Therefore, G alpha s is expressed differentially during Phycomyces development.
Assuntos
Proteínas de Ligação ao GTP/metabolismo , Phycomyces/fisiologia , Adenosina Difosfato Ribose/metabolismo , Marcadores de Afinidade , Toxina da Cólera/farmacologia , DNA Fúngico/genética , Proteínas de Ligação ao GTP/química , Proteínas de Ligação ao GTP/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação Fúngica da Expressão Gênica , Genes Fúngicos , Guanosina Trifosfato , Phycomyces/genética , Phycomyces/crescimento & desenvolvimento , Esporos Fúngicos/genética , Esporos Fúngicos/fisiologiaRESUMO
Chitin synthetase activity was analyzed in vitro and in vivo in two morphogenetic stages, namely, dormant spore cells and germlings of the wild type strain and the developmental mutant S356 of Phycomyces blakesleeanus. In vitro experiments showed a much higher specific activity in dormant spores of the mutant strain than in those of the wild-type. This difference was restricted to the dormant spore phase since germlings exhibited comparable levels of activity to those detected in the wild-type strain. Although no correlation was observed between chitin synthesis in vitro and in vivo in mutant spores, germination of these cells was accompanied by an earlier expression of chitin synthetase in vivo. Germination of mutant spores in liquid medium produced morphologically aberrant germlings. Contrary to the extended mycelial growth of the wild-type strain in solid medium, the mutant grew with a typical colonial morphology. Results are discussed in relation to the possible basis of the mutant phenotype.
Assuntos
Quitina Sintase/genética , Phycomyces/genética , Quitina Sintase/biossíntese , Morfogênese , Mutação , Fenótipo , Phycomyces/enzimologia , Phycomyces/crescimento & desenvolvimento , Esporos Fúngicos/fisiologia , Especificidade por SubstratoRESUMO
Diamino butanone (DAB), a competitive inhibitor of ornithine decarboxylase (ODC) a key enzyme in polyamine biosynthesis, inhibited the yeast to hyphae transition in Mucor rouxii, induced by transfer from anaerobiosis to aerobiosis, but not the opposite phenomenon. Addition of DAB to anaerobic yeast cells brought about a decrease in ODC and polyamine levels. In these conditions, the aerobic shift produced only a weak increase in ODC activity and no change in polyamine levels. DAB also blocked phorogenesis in M. rouxii and in Phycomyces blakesleeanus. At the effective concentrations DAB did not affect cell growth of either fungus. It is suggested that low, constant levels of ODC and polyamines are necessary for cell growth, and that high transient levels are required during the differentiative steps. DAB, at the concentrations used, affects this last process, but does not interfere with the maintenance level of polyamines.