Department of Plant Pathology1 and Department of Mathematics2, University of California, Riverside, CA 92521-0122, USA
Author for correspondence: Salomon Bartnicki-García. Tel: +1 909 787 4135. Fax: +1 909 787 4294. e-mail: bart{at}ucrac1.ucr.edu
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ABSTRACT |
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Keywords: ropy mutants, Spitzenkörper, Neurospora crassa
Abbreviations: Spk, Spitzenkörper; VSC, vesicle supply centre
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INTRODUCTION |
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The Spk is a dynamic body whose structure varies widely among fungi; it usually consists of an outer vesicle cloud and an inner core (López-Franco & Bracker, 1996 ). In addition to vesicles (Girbardt, 1969
; Grove & Bracker, 1970
), other components have been detected in the Spk, including amorphous or granular material of undefined nature in the core region as well as components of the cytoskeleton and ribosomes (McClure et al., 1968
; Turian, 1978
; Bourett & Howard, 1991
; Roberson & Vargas, 1994
; López-Franco & Bracker, 1996
). The morphology of the fungal cells is determined by the way the cell wall is assembled (Bartnicki-García, 1968
, 1973
). In hyphae, cell wall growth occurs mainly at the tip by polarized secretion of enzymes and cell wall precursors (Bartnicki-García & Lippman, 1969
; Trinci, 1978
; Harold, 1990
). There is a sizeable body of evidence that the Spk plays a central role in apical growth and morphogenesis (Girbardt, 1957
; López-Franco & Bracker, 1996
; Reynaga-Peña & Bartnicki-García, 1997
). According to the vesicle supply centre (VSC) model for fungal morphogenesis (Bartnicki-García et al., 1989
), the Spk functions as a vesicle distribution centre. Vesicles generated in distal parts of the hypha congregate in the Spk and from there migrate to the cell surface. The linear displacement of the Spk creates a sharp gradient of exocytosis responsible for hyphal morphogenesis.
There is mounting evidence that the position of the Spk governs the growth direction of a hypha (Girbardt, 1957 ; Bracker et al., 1997
; Riquelme et al., 1998
). Our previous work with microtubule inhibitors implicated the microtubular cytoskeleton in the positioning and movement of the Spk in hyphae of N. crassa (Riquelme et al., 1998
), but its exact role in apical growth has yet to be elucidated. It has long been proposed that cytoplasmic microtubules participate in the transport of secretory vesicles to the hyphal apex (Howard & Aist, 1977
, 1980
; Howard, 1981
; Gooday, 1983
; Gow, 1989
; Hasek & Bartnicki-García, 1994
; McKerracher & Heath, 1987
; Heath, 1994
). The discovery of microtubule-associated motor proteins (Hirokawa, 1982
; Paschal et al., 1987
) has helped us understand how organelles move inside the cell. Cytoplasmic dyneins and members of the kinesin superfamily are the main motor enzymes involved in vesicle translocation along microtubules (Haimo & Thaler, 1994
; Hirokawa, 1998
). There is now growing evidence that both kinesins and cytoplasmic dyneins are involved in the traffic of secretory vesicles in fungal hyphae (Seiler et al., 1997
; Wu et al., 1998
; Inoue et al., 1998
). Cytoplasmic dyneins are multisubunit enzymes involved in transport of membranous organelles towards the minus end of microtubules (Paschal et al., 1987
; Schroer & Sheetz, 1991
; Hirokawa, 1998
). Kinesins are motor proteins involved in membrane transport towards the plus end of the microtubules (Vale et al., 1985
; Hirokawa, 1982
; Steinberg & Schliwa, 1996
).
To examine the relationship among the Spk, the microtubular cytoskeleton and hyphal morphogenesis, we chose two ropy mutants of N. crassa, ro-1 and ro-3. Both belong to the group of true colonial morphological mutants (Garnjobst & Tatum, 1967 ; Vierula, 1996
) and have been characterized at the molecular level. Mutant ro-1 is deficient in one of the heavy chains of cytoplasmic dynein (Plamann et al., 1994
). Mutant ro-3 is deficient in p150Glued, the largest subunit of the dynactin (dynein activator) complex (Plamann et al., 1994
; Tinsley et al., 1996
). Dynactin is a multisubunit complex required for cytoplasmic dynein to efficiently transport vesicles along microtubules in vitro (Gill et al., 1991
; Schroer & Sheetz, 1991
).
Most studies on ropy mutants have primarily focused on the aberrant distribution of nuclei in ro-1 and ro-3 hyphae and on the molecular characterization of those loci (Plamann et al., 1994 ; Bruno et al., 1996
; Tinsley et al., 1996
). Since hyphal and, ultimately, colony morphology are largely established at the hyphal apex, we have focused this study on the impact of the ropy mutations on apical events, where most of the growth process is concentrated.
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METHODS |
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Video microscopy.
Growing hyphae were observed with an Olympus Vanox-S microscope. For low-magnification images, the fungus was grown in 8·5 cm plastic Petri dishes on a thin layer (10 ml) of VCM agar at 21 °C and observed with bright-field optics (10x objective and 10x WF eyepiece). For high-resolution work, the fungus was grown on a modified slide culture chamber (López-Franco, 1992 ; Riquelme et al., 1998
) and observed with a phase-contrast 100x oil-immersion objective (n.a. 1·25) and a 25x WF eyepiece (American Optical).
Video images were produced with a Hamamatsu C2400-07 high-resolution camera (Hamamatsu Photonic Systems), enhanced with an Argus-10 image processor (a real-time digital contrast and low light enhancement system), and displayed on a black and white, 12-inch, high-resolution monitor (Sony; model PVM-122). Sequences were videotaped in real time with an S-VHS recorder (JVC model BR-S822U).
Growth rate and cell parameters measurements.
Videotaped sequences were played on a variable-tracking player (JVC model BR-S525U) and observed on a Sony Trinitron model monitor. Individual images were captured from the videotaped sequences in 8-bit greyscale with an Imascan/Chroma frame grabber (Imagraph). With Image Pro Plus Software for Windows (Media Cybernetics), we traced the cell profiles of the images captured by video microscopy. xy coordinate values were automatically collected into a text file with a Windows application program interfaced with the Argus-10 analyser (Bartnicki et al., 1994 ). The text files were then imported into Microsoft Excel spreadsheets and analysed.
Growth rate was measured in terms of cell area increase. Typically, measurements were made during growth periods of 14 min depending on growth rate and availability of well-defined profiles. For convenience and accuracy, only images within the same field of view were compared. The area delimited by the cell profiles was computed by using Greens formula (Marsden et al., 1993 ). Spk diameter was measured directly on the monitor screen with the line command of the measure option in the Argus-10 menu.
Spk trajectory analysis.
The centre of the Spk was mapped at 2 s intervals from the videotaped sequences. To quantitate the erratic behaviour of the advancing Spk, we calculated a steadiness index (S), namely the ratio of the minimal distance between the initial and final position of the Spk in a given videotaped sequence over the total distance travelled by the Spk. A steadiness index of 1 would correspond to a perfectly straight path. The smaller the steadiness index the less steady the trajectory of the Spk.
Computer simulation.
From videotaped sequences of hyphae of wild-type and ropy mutants, we mapped the Spk position (every 2 s) and traced the cell profiles at various intervals. These data were fed to the Fungus Simulator [a Windows program for fungal morphogenesis (Bartnicki et al., 1994 ) available through the Internet (http://boyce3427.ucr.edu)]. The Fungus Simulator generates hyphal shapes by a process that mimics a vesicle-based mechanism for cell wall growth.
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RESULTS |
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Another common difference was in vacuolation. The cytoplasm of both ropy mutants was more vacuolated than in wild-type. In ropy hyphae, large, phase-light and phase-dark structures of irregular shape (probably vacuoles and lipid bodies) accumulated at a shorter distance (as close as 1015 µm) from the tip. Occasionally, one or more small phase-dark globular bodies were seen immediately behind the Spk (Fig. 3d, g
; arrowheads). These bodies moved continuously within the area surrounding the Spk but appeared to be physically attached to the Spk by thin filaments and remained behind the Spk for the entire observation period. These granules were usually more conspicuous in the ropy mutants but were some times seen in tips of wild-type hyphae. Similar structures have been seen in other fungi (López-Franco & Bracker, 1996
) but their function remains unknown.
There was a pronounced difference in overall movement of cytoplasmic structures between wild-type and ropy mutants. To judge differences in organelle movement, the videotaped growth sequences were examined in real time and at three times the original speed. The cytoplasm in the hyphae of ropy mutants was ostensibly less dynamic than that in wild-type hyphae. In the latter, cytoplasmic organelles were in constant motion over the entire length of the subapical area observed (at least 2025 µm from the apex). In ropy hyphae, motion was less active and was usually restricted to the proximal subapex (515 µm from the apex); beyond that, the cytoplasm became static and extensively vacuolated (Fig. 3; see also video in http://boyce3427.ucr.edu/cytoskel.htm). In both ropy mutants, the movement of cytoplasmic structures appeared less organized than that in wild-type hyphae. In growing wild-type hyphae, the mitochondria moved continuously in a back and forth fashion roughly parallel to the longitudinal axis of growth. In the ropy mutants, mitochondria did not move preferentially along the longitudinal axis but in a more erratic manner.
Growth rate and Spk size
Hyphal elongation is traditionally used as a measurement of the growth rate. This method is accurate provided the hypha maintains a relatively constant shape during elongation. For the ropy mutants, with their distorted hyphal morphology, this method was clearly not valid. Instead, we measured growth rate as increase in cell area per unit of time (Table 2) and calculated that the mean growth rate of hyphae of ro-1 and ro-3 was 45 and 34%, respectively, of that in wild-type hyphae. There was no significant difference in growth rate between the two ropy mutants (Table 2
).
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The appearance and size of the Spk varied greatly during growth, particularly in the ropy mutants. For instance, during the observation of a hyphal tip of N. crassa ro-1 growing at 0·60·7 µm2 s-1, a smaller and less phase-dark Spk was visible for about 220 s (Fig. 5), then the Spk disappeared, coinciding with a sharp decrease of the growth rate to 0·1 µm2 s-1. When the growth rate returned to almost the original values (0·5 µm2 s-1), the Spk became visible again.
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Computer simulation
To analyse the effect of Spk movement on hyphal morphogenesis, the traced Spk trajectories and cell profiles of wild-type and ropy mutants were fed to the Fungus Simulator program. The VSC of the simulator was programmed to follow actual Spk trajectories. Cell profiles were used to calculate the amount of vesicles to be released by the VSC at each point in the trajectory. With these two sets of input data, the simulator duplicated the morphologies of wild-type hyphae and the highly distorted shapes of ropy mutants hyphae (Fig. 6).
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DISCUSSION |
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Impairment of dynein or dynactin caused major effects at different levels: reduced growth rate, distorted morphology, aberrant cytoplasmic organization, disrupted organelle motility, and a smaller Spk with erratic trajectory. The effects of these mutations on nuclear distribution and mitosis have been previously documented for this and other fungi (Xiang et al., 1994 , 1995
; Plamann et al., 1994
; Tinsley et al., 1996
; Bruno et al., 1996
; Inoue et al., 1998
). Apparently, the reduction in the overall movement of intracellular components along the microtubular cytoskeleton, caused by dynein/dynactin deficiencies, had a general effect on all cytoplasmic activities and hence led to an overall reduction in growth rate. But diminished growth rate need not result in distorted morphology; it should result in slower-growing hyphae with similar morphology. We believe the morphogenetic effects caused by these mutations can be directly ascribed to their impact on Spk formation and behaviour. Invariably, the Spk of ropy hyphae was smaller and lacked the stability of the wild-type Spk. The fact that the ropy mutants grew poorly and produced deformed hyphae on Petri dish cultures in the dark eliminates the possibility that the Spk deficiencies we observed were caused by stresses imposed on the mutants during microscopy.
Spk size
Presumably, the smaller size of the Spk of ropy hyphae is a consequence of a diminished supply of vesicles to the apical region caused by the dynein/dynactin deficiency in the mutants. This conclusion differs from that made by Seiler et al. (1999) , who described the ro-1 mutant as having a prominent Spk. However, they apparently did not take into account that the Spk of ro-1 is considerably smaller than that of the wild-type, and at times may not even be visible, as shown here in Fig. 4
(see also http://boyce3427.ucr.edu/cytoskel.htm for video). Consequently, their conclusion that apical transport was intact in this ropy mutant runs contrary to our evidence, which clearly shows that the processes responsible for Spk formation, including apical vesicle traffic, are affected by dynein deficiency. Inoue et al. (1998)
also found that dynein is required for normal secretory vesicle transport to the hyphal apex of Nectria haematococca.
Since mutations in kinesin are known to affect Spk formation (Seiler et al., 1997 ; Wu et al., 1998
), it was proposed that cytoplasmic microtubules are oriented with their plus ends towards the apex (Lehmler et al., 1997
; Seiler et al., 1997
, 1999
). By the same reasoning, our studies showing that deficiency in cytoplasmic dynein or dynactin affects Spk formation could lead us to the opposite conclusion, namely that the microtubules are oriented with their minus ends towards the apex. Clearly, none of these observations can reveal conclusively the polarity of cytoplasmic microtubules in a hypha. The similarity of phenotypic effects caused by the impairment of opposite motor proteins suggests that both motors are necessary for the maintenance of the Spk and apical growth, but whether or not both are directly involved in the apical transport of secretory vesicles remains to be seen. Possibly, a deficiency in cytoplasmic dynein or kinesin may also impair the endocytotic processes that contribute to the recycling of membranous components from apex to subapex (Hoffmann & Mendgen, 1998
), and thus affect Spk formation.
Overall, our analyses showed positive, though not necessarily linear, correlations between Spk size, hyphal growth rate and hyphal diameter in both the wild-type and the ropy mutants of N. crassa. In general, fast-growing hyphae share a tendency to have a larger hyphal diameter and a larger Spk than slow-growing hyphae. Similar tendencies were observed in other fungi (López-Franco & Bracker, 1996 ). Wu et al. (1998)
found a similar correlation between Spk size, growth rate and hyphal diameter in a kinesin-deletion mutant of Nectria haematococca. In other tip-growing cells such as pollen tubes and root hairs, the size of what would be their Spk equivalent (so-called tip body or clear cap) has also been correlated with high growth rate (Reiss & Herth, 1979
; Sievers, 1963
).
Spk behaviour and morphogenesis
The highly erratic behaviour of the Spk in ropy mutants is difficult to interpret since we do not know for sure which cellular components determine the positioning and advance of such a complex and dynamic structure as the Spk. Previously, based on comparative results with benomyl and cytochalasin, we proposed that the microtubule cytoskeleton was directly involved in maintaining the trajectory of the Spk (Riquelme et al., 1998 ). Our present observations with the dynein-deficient ropy mutants lend support to the notion that the microtubule cytoskeleton plays a major role in the formation and behaviour of the Spk. Similarly, Wu et al. (1998)
showed that kinesin was essential for normal positioning of the Spk in hyphae of N. haematococca. All this leads us to conclude that microtubule-associated motor proteins are necessary for maintenance of a high growth rate, a rather steady Spk and a near-perfect hyphoid shape.
Regardless of the exact mechanism controlling the position of the Spk, we have reason to conclude that the erratic trajectory of the Spk is directly responsible for the distorted morphology of ropy hyphae. As we did in studying other morphogenetic processes (Bartnicki-García et al., 1995 ; Reynaga-Peña & Bartnicki-García, 1997
; Riquelme et al., 1998
), we used the Fungus Simulator to test the correlation between Spk trajectory and cell shape. The simulator generated forms that reproduced the distorted morphology of the ropy hyphae. This indicates that the fungal Spk, operating as a VSC, is the structure that ultimately controls the shape of fungal hyphae. In wild-type hyphae, the Spk advances in a fixed direction with only minor transverse oscillations. The spatially uniform vesicle traffic emanating from such a Spk produces a smooth regular shape that approximates the ideal hyphoid shape stipulated by the hyphoid equation (Bartnicki-García et al., 1989
). In the ropy hyphae, sustained departures in the trajectory of the Spk result in corresponding distortions in the morphology of the hypha.
Barring pleiotropic effects caused by the ropy mutations, the dynein/dynactin deficiency probably causes morphogenetic effects by (1) diminishing the traffic of secretory vesicles from their synthesis site (endoplasmic reticulum) either to intermediate secretory compartments or to the hyphal apex; (2) affecting the organization and movement of the growing microtubules; and/or (3) impeding the proper recycling of material needed for normal apical growth to proceed.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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Bartnicki-García, S. (1968). Cell wall chemistry, morphogenesis, and taxonomy of fungi.Annu Rev Microbiol 22, 87-107.[Medline]
Bartnicki-García, S. (1973). Fundamental aspects of hyphal morphogenesis. In Microbial Differentiation, pp. 245-267. Edited by J. M. Ashworth & J. E. Smith. Cambridge: Cambridge University Press.
Bartnicki-García, S. & Lippman, E. (1969). Fungal morphogenesis. Cell wall construction in Mucor rouxii.Science 165, 302-304.[Medline]
Bartnicki-García, S., Hergert, F. & Gierz, G. (1989). Computer simulation of fungal morphogenesis and the mathematical basis for (hyphal tip) growth.Protoplasma 153, 46-57.
Bartnicki-García, S., Bartnicki, D. D., Gierz, G., López-Franco, R. & Bracker, C. E. (1995). Evidence that Spitzenkörper behavior determines the shape of a fungal hypha: a test of the hyphoid model.Exp Mycol 19, 153-159.[Medline]
Bourett, T. M. & Howard, R. J. (1991). Ultrastructural immunolocalization of actin in a fungus.Protoplasma 163, 199-202.
Bracker, C. E. (1995). The video-enhanced light microscope: a renaissance tool for quantitative live-cell microscopy.Zool Stud 34, 154-156.
Bracker, C. E., Murphy, D. J. & López-Franco, R. (1997). Laser microbeam manipulation of cell morphogenesis in growing fungal hyphae. In Functional Imaging and Optical Manipulation of Living Cells, pp. 6780. Edited by D. L. Farkas & B. J. Tromberg. Bellingham, WA: SPIE (International Society for Optical Engineering) (Proceedings of SPIE vol. 2893).
Bruno, K. S., Tinsley, J. H., Minke, P. F. & Plamann, M. (1996). Genetic interactions among cytoplasmic dynein, dynactin, and nuclear distribution mutants of Neurospora crassa.Proc Natl Acad Sci USA 93, 4775-4780.
Garnjobst, L. & Tatum, E. L. (1967). A survey of new morphological mutants in Neurospora crassa.Genetics 57, 579-604.
Gill, S. R., Schroer, T. A., Szilak, I., Steuer, E. R., Sheetz, M. P. & Cleveland, D. W. (1991). Dynactin, a conserved, ubiquitously expressed component of an activator of vesicle motility mediated by cytoplasmic dynein.J Cell Biol 115, 1639-1650.[Abstract]
Girbardt, M. (1957). Der Spitzenkörper von Polystictus versicolor (L.).Planta 50, 47-59.
Girbardt, M. (1969). Die Ultrastruktur der Apikalregion von Pilzhyphen.Protoplasma 67, 413-441.
Gooday, G. W. (1983). The hyphal tip. In Fungal Differentiation, pp. 315-356. Edited by J. E. Smith. New York: Marcel Dekker.
Gow, N. A. R. (1989). Control of the extension of the hyphal apex.Curr Top Med Mycol 3, 109-152.[Medline]
Grove, S. N. & Bracker, C. E. (1970). Protoplasmic organization of hyphal tips among fungi.J Bacteriol 104, 989-1009.[Medline]
Haimo, L. T. & Thaler, C. D. (1994). Regulation of organelle transport: lessons from color change in fish.BioEssays 16, 727-733.
Harold, F. M. (1990). To shape a cell: an inquiry into the causes of morphogenesis of microorganisms.Microbiol Rev 54, 381-431.
Hasek, J. & Bartnicki-García, S. (1994). The arrangement of F-actin and microtubules during germination of Mucor rouxii sporangiospores.Arch Microbiol 161, 363-369.[Medline]
Heath, I. B. (1994). The cytoskeleton in hyphal growth, organelle movements, and mitosis. In The Mycota I: Growth, Differentiation and Sexuality, pp. 43-65. Edited by J. G. H. Wessels & F. Meinhardt. Berlin & Heidelberg: Springer.
Hirokawa, N. (1982). Cross-linker system between neurofilaments, microtubules and membranous organelles in frog axons revealed by the quick-freeze, deep-etching method.J Cell Biol 94, 129-142.
Hirokawa, N. (1998). Kinesin and dynein superfamily proteins and the mechanism of organelle transport.Science 279, 519-526.
Hoch, H. C. & Staples, R. C. (1985). The microtubule cytoskeleton in hyphae of Uromyces phaseoli germlings: its relationship to the region of nucleation and to the F-actin cytoskeleton.Protoplasma 124, 112-122.
Hoffmann, J. & Mendgen, K. (1998). Endocytosis and membrane turnover in the germ tube of Uromyces fabae.Fungal Genet Biol 24, 77-85.
Howard, R. J. (1981). Ultrastructural analysis of hyphal tip cell growth in fungi: Spitzenkörper, cytoskeleton and endomembranes after freeze-substitution.J Cell Sci 48, 89-103.[Abstract]
Howard, R. & Aist, J. R. (1977). Effects of MBC on hyphal tip organization, growth, and mitosis of Fusarium acuminatum, and their antagonism by D2O.Protoplasma 92, 195-210.[Medline]
Howard, R. J. & Aist, J. R. (1980). Cytoplasmic microtubules and fungal morphogenesis: ultrastuctural effects of methyl benzimidazole-2-ylcarbamate determined by freeze-substitution of hyphal tip cells.J Cell Biol 87, 55-64.[Abstract]
Inoue, S., Turgeon, B. G., Yoder, O. C. & Aist, J. R. (1998). Role of fungal dynein in hyphal growth, microtubule organization, spindle pole body motility and nuclear migration.J Cell Sci 111, 1555-1566.
Lehmler, C., Steinberg, G., Snetselaar, K. M., Schliwa, M., Kahmann, R. & Bölker, M. (1997). Identification of a motor protein required for filamentous growth in Ustilago maydis.EMBO J 16, 3464-3473.
López-Franco, R. (1992). Organization and dynamics of the Spitzenkörper in growing hyphal tips. PhD dissertation, Purdue University, West Lafayette, IN.
López-Franco, R. & Bracker, C. E. (1996). Diversity and dynamics of the Spitzenkörper in growing hyphal tips of higher fungi.Protoplasma 195, 90-111.
McClure, W. K., Park, D. & Robinson, P. M. (1968). Apical organization in the somatic hyphae of fungi.J Gen Microbiol 50, 177-182.[Medline]
McKerracher, L. J. & Heath, I. B. (1987). Cytoplasmic migration and intracellular organelle movements during tip growth of fungal hyphae.Exp Mycol 11, 79-100.
Marsden, J. E., Tromba, A. J. & Weinstein, A. (1993). Basic Multivariable Calculus. New York: Springer.
Paschal, B. M., Shpetner, H. S. & Vallee, R. B. (1987). MAP 1C is a microtubule-activated ATPase which translocates microtubules in vitro and has dynein-like properties.J Cell Biol 105, 1273-1282.[Abstract]
Plamann, M., Minke, P. F., Tinsley, J. H. & Bruno, K. S. (1994). Cytoplasmic dynein and actin-related protein Arp1 are required for normal nuclear distribution in filamentous fungi.J Cell Biol 127, 139-149.[Abstract]
Reiss, H. D. & Herth, W. (1979). Calcium gradients in tip growing plant cells visualized by chlorotetracycline fluorescence.Planta 146, 615-621.
Reynaga-Peña, C. G. & Bartnicki-García, S. (1997). Apical branching in a temperature sensitive mutant of Aspergillus niger.Fungal Genet Biol 22, 153-167.[Medline]
Riquelme, M., Reynaga-Peña, C. G., Gierz, G. & Bartnicki-García, S. (1998). What determines growth direction in fungal hyphae?Fungal Genet Biol 24, 101-109.
Roberson, R. W. & Vargas, M. M. (1994). The tubulin cytoskeleton and its sites of nucleation in hyphal tips of Allomyces macrogynus.Protoplasma 182, 19-31.
Schroer, T. A. & Sheetz, M. P. (1991). Two activators of microtubule-based vesicle transport.J Cell Biol 115, 1309-1318.[Abstract]
Seiler, S., Nargang, F. E., Steinberg, G. & Schliwa, M. (1997). Kinesin is essential for cell morphogenesis and polarized secretion in Neurospora crassa.EMBO J 16, 3025-3034.
Seiler, S., Plamann, M. & Schliwa, M. (1999). Kinesin and dynein mutants provide novel insights into the roles of vesicle traffic during cell morphogenesis in Neurospora.Curr Biol 9, 779-785.[Medline]
Sievers, A. (1963). Beteiligung des Golgi-Apparates bei der Bildung der Zellwand von Wurzelhaaren.Protoplasma 56, 188-192.
Steinberg, G. & Schliwa, M. (1996). Characterization of the biophysical and motility properties of kinesin from the fungus Neurospora crassa.J Biol Chem 271, 7516-7521.
Tinsley, J. H., Minke, P. F., Bruno, K. S. & Plamann, M. (1996). p150Glued, the largest subunit of the dynactin complex, is nonessential in Neurospora but required for nuclear distribution.Mol Biol Cell 7, 731-742.[Abstract]
Trinci, A. P. J. (1978). Wall and hyphal growth.Sci Prog 65, 75-79.
Turian, G. (1978). The Spitzenkörper, centre of the reducing power in the growing hyphal apices of two septomycetous fungi.Experientia 34, 1277-1279.
Vale, R. D., Reese, T. S. & Sheetz, M. P. (1985). Identification of a novel force-generating protein, kinesin, involved in microtubule-based motility.Cell 42, 39-50.[Medline]
Vierula, P. J. (1996). The genetics of morphogenesis in Neurospora crassa. In Patterns in Fungal Development, pp. 87-104. Edited by S.-W. Chiu & D. Moore. Cambridge: Cambridge University Press.
Vogel, H. J. (1956). A convenient growth medium for Neurospora (Medium N).Microb Genet Bull 13, 42-43.
Wu, Q., Sandrock, T. M., Turgeon, B. G., Yoder, O. C., Wirsel, S. G. & Aist, J. R. (1998). A fungal kinesin required for organelle motility, hyphal growth, and morphogenesis.Mol Biol Cell 9, 89-101.
Xiang, X., Beckwith, S. M. & Morris, N. R. (1994). Cytoplasmic dynein is involved in nuclear migration in Aspergillus nidulans.Proc Natl Acad Science USA 91, 2100-2104.[Abstract]
Xiang, X., Roghi, C. & Morris, N. R. (1995). Characterization and localization of the cytoplasmic dynein heavy chain in Aspergillus nidulans.Proc Natl Acad Sci USA 92, 9890-9894.[Abstract]
Received 5 January 2000;
revised 29 March 2000;
accepted 25 April 2000.