Helical growth during the phototropic response, avoidance response, and in stiff mutants of Phycomyces blakesleeanus.

The sporangiophores of Phycomyces blakesleeanus have been used as a model system to study sensory transduction, helical growth, and to establish global biophysical equations for expansive growth of walled cells. More recently, local statistical biophysical models of the cell wall are being constructed to better understand the molecular underpinnings of helical growth and its behavior during the many growth responses of the sporangiophores to sensory stimuli. Previous experimental and theoretical findings guide the development of these local models. Future development requires an investigation of explicit and implicit assumptions made in the prior research. Here, experiments are conducted to test three assumptions made in prior research, that (a) elongation rate, (b) rotation rate, and (c) helical growth steepness, R, of the sporangiophore remain constant during the phototropic response (bending toward unilateral light) and the avoidance response (bending away from solid barriers). The experimental results reveal that all three assumptions are incorrect for the phototropic response and probably incorrect for the avoidance response but the results are less conclusive. Generally, the experimental results indicate that the elongation and rotation rates increase during these responses, as does R, indicating that the helical growth steepness become flatter. The implications of these findings on prior research, the "fibril reorientation and slippage" hypothesis, global biophysical equations, and local statistical biophysical models are discussed.

Dimensionless numbers to study cell wall deformation of stiff mutants of Phycomyces blakesleeanus.

The sporangiophores of Phycomyces blakesleeanus are large cylindrical aerial cells that elongate vertically at rates between 10 µm/min and 60 µm/min. Wild-type sporangiophores grow toward light, opposed to gravitational acceleration and away from solid barriers (tropic responses). Sporangiophores of stiff mutants C149 and C216 exhibit diminished tropic (bending) responses. Originally, it was thought that the altered genes affect the "stiffness" (elastic wall deformation) of the cell wall. Subsequent investigations employing the pressure probe demonstrated that the irreversible (plastic) wall deformation was smaller for the stiff mutants compared to wild type and could account for the diminished tropic responses. However, it was not shown whether the elastic wall deformation was altered in these stiff mutants. Recent theoretical studies have identified dimensionless numbers that can be used to quantitate the magnitudes of biophysical processes involved in expansive growth of walled cells. In this study, dimensionless numbers are used to determine the magnitudes of elastic deformation rate, plastic deformation rate, and stress relaxation rate of the cell wall during expansive growth of the stiff mutant sporangiophores. It is found that the altered genes reduce stress relaxation rates and plastic deformation rates of the wall, but do not significantly alter the magnitude of the elastic deformation rates of the wall. These results indicate that the mutant genes reduce wall loosening chemistry in these sporangiophores and the genetic mutation is not expressed in a change in "wall stiffness," but in "wall viscosity" or "wall extensibility."

Cell Wall Loosening in the Fungus, Phycomyces blakesleeanus.

A considerable amount of research has been conducted to determine how cell walls are loosened to produce irreversible wall deformation and expansive growth in plant and algal cells. The same cannot be said about fungal cells. Almost nothing is known about how fungal cells loosen their walls to produce irreversible wall deformation and expansive growth. In this study, anoxia is used to chemically isolate the wall from the protoplasm of the sporangiophores of Phycomyces blakesleeanus. The experimental results provide direct evidence of the existence of chemistry within the fungal wall that is responsible for wall loosening, irreversible wall deformation and elongation growth. In addition, constant-tension extension experiments are conducted on frozen-thawed sporangiophore walls to obtain insight into the wall chemistry and wall loosening mechanism. It is found that a decrease in pH to 4.6 produces creep extension in the frozen-thawed sporangiophore wall that is similar, but not identical, to that found in frozen-thawed higher plant cell walls. Experimental results from frozen-thawed and boiled sporangiophore walls suggest that protein activity may be involved in the creep extension.

Stiff mutant genes of phycomyces affect turgor pressure and wall mechanical properties to regulate elongation growth rate.

Regulation of cell growth is paramount to all living organisms. In plants, algae and fungi, regulation of expansive growth of cells is required for development and morphogenesis. Also, many sensory responses of stage IVb sporangiophores of Phycomyces blakesleeanus are produced by regulating elongation growth rate (growth responses) and differential elongation growth rate (tropic responses). "Stiff" mutant sporangiophores exhibit diminished tropic responses and are found to be defective in at least five genes; madD, E, F, G, and J. Prior experimental research suggests that the defective genes affect growth regulation, but this was not verified. All the growth of the single-celled stalk of the stage IVb sporangiophore occurs in a short region termed the "growth zone." Prior experimental and theoretical research indicates that elongation growth rate of the stage IVb sporangiophore can be regulated by controlling the cell wall mechanical properties within the growth zone and the magnitude of the turgor pressure. A quantitative biophysical model for elongation growth rate is required to elucidate the relationship between wall mechanical properties and turgor pressure during growth regulation. In this study, it is hypothesized that the mechanical properties of the wall within the growth zone of stiff mutant sporangiophores are different compared to wild type (WT). A biophysical equation for elongation growth rate is derived for fungal and plant cells with a growth zone. Two strains of stiff mutants are studied, C149 madD120 (-) and C216 geo- (-). Experimental results demonstrate that turgor pressure is larger but irreversible wall deformation rates within the growth zone and growth zone length are smaller for stiff mutant sporangiophores compared to WT. These findings can explain the diminished tropic responses of the stiff mutant sporangiophores. It is speculated that the defective genes affect the amount of wall-building material delivered to the inner cell wall.