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Chapter 2 : Hyphal Structure
Category: Microbial Genetics and Molecular Biology; Fungi and Fungal Pathogenesis
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The ability to understand cytoplasmic structure can provide powerful insights into the biology of cells and organisms. This chapter has briefly reviewed the diversity of hyphal structures and presented examples of how bioimaging has contributed to a broader understanding of hyphal biology and phylogenetic relationships between fungal taxa. At the heart of polarized growth is the secretory pathway in which vesicles are targeted to sites of growth and subsequently fuse with the plasma membrane. In mature hyphae of the septate fungi, these events have given rise to the Spitzenkörper, a complex and dynamic structure that clearly influences hyphal growth and morphogenesis. The presence or absence of certain morphological characters (e.g., septa and Woronin bodies) already has been useful in defining higher taxa, especially since evolutionary polarity often can be established using stable phylogenetic trees based on DNA sequences. Ever-enlarging molecular databases, especially those of whole genomes, are allowing us to look for the genetic basis of many structural features, such as the presence or absence of Woronin body matrix proteins. This capability will allow us to understand the basis of these features not only in an evolutionary sense but also in a functional one. Collaboration among different types of fungal biologists including systematists is essential to understanding structure and how it applies to the study of the Fungi.
Cytoplasmic order of hyphal tip. (a) Phase-contrast light micrograph of N. crassa with Spitzenkörper (white arrow), mitochondria (arrowheads), and nucleus (asterisk). Scale bar, 4 μm. (b) Near-median TEM section through S. rolfsii cryofixed hypha illustrating Spitzenkörper (white arrow), mitochondria (white arrowheads), rough ER (black arrowheads), vacuoles (black arrows), and nucleus (asterisk). Scale bar, 2 μm. Reprinted from Protoplasma ( Roberson and Fuller, 1988 ) with permission from the publisher.
Phase-contrast light micrograph of N. crassa hyphal tip illustrating the phase-dark region (arrow) and the subtending phase-light core of Spitzenkörper. Mitochondria are indicated by arrowheads. Scale bar, 1.5 μm.
Near-median TEM section through hyphal tip of Botrytis cinerea (Ascomycota) illustrating structural components of the Spitzenkörper and surrounding cytoplasm. Shown are apical vesicles (white arrows); microvesicles (white arrowheads); central core (asterisk); MTs (black arrows); MFs (small white arrows); plasma membrane at sites of exocytosis (black arrowheads); ribosomes (small black arrows); cell wall (CW); cell wall inclusions (black asterisks). Scale bar, 200 nm.
Phase-contrast light micrograph of A. macrogynus hyphal tip illustrating phase-light core of Spitzenkörper subtended by mitochondria (asterisk). Scale bar, 4 μm. Reprinted from Fungal Genetics and Biology ( McDaniel and Roberson, 2000 ) with permission from the publisher.
Near-median TEM section through hyphal tip of A. macrogynus illustrating structural components of the Spitzenkörper and surrounding cytoplasm: central core (asterisk); microvesicles (small arrows); MTs (arrows); and mitochondria (arrowheads). Scale bar, 1.7 μm. Reprinted from Fungal Genetics and Biology ( McDaniel and Roberson, 2000 ) with permission from the publisher.
TEM of nuclei of S. rolfsii illustrating prominent nucleolus (asterisk), nuclear pore complexes (white arrowheads), SPB (white arrow), mitochondrion (m), rough ER (black arrows), and vacuole (V). Scale bar, 350 nm.
TEM of SPB and mitosis. (a and b) Sections 3 and 5 of a complete series of 6 through a longitudinally sectioned late interphase SPB of Helicobasidium mompa showing the disk form (arrowheads) of the SPB characteristic of Ascomycota and Pucciniomycotina, Basidiomycota. Shown are the middle piece of SPB (white arrows), nuclear envelope (black arrow), and the nucleus (N). Scale bar, 0.2 μm. Reprinted from the Canadian Journal of Botany ( Bourett and McLaughlin, 1986 ) with permission of the publisher. (c and d) Sections 3 and 4 of a complete series of 8 through a late prophase SPB of Auricularia auricula-judae (Agaricomycotina, Basidiomycota) showing the two globular elements (asterisks) and middle piece (white arrows) surrounded by a ribosome-free zone; an intranuclear element (black arrowhead) consisting of two layers lies within the nucleus (N) adjacent to the nuclear envelope. The intranuclear element is characteristic of many members of Ascomycota and Basidiomycota. Scale bar, 0.2 μm. Reprinted from the Canadian Journal of Botany (Lu and McLaughlin, 1995) with permission of the publisher. (e) The metaphase SPB of H. mompa is a disk (asterisk) set in a pore in the nuclear envelope surrounded by a membranous cap (MC). S, spindle. Scale bar, 0.2 μm. Reprinted from the Canadian Journal of Botany ( Bourett and McLaughlin, 1986 ) with permission of the publisher. (f) The metaphase SPB of Auriscalpium vulgare (Agaricomycotina, Basidiomycota) is a globular element (G) with a dense inclusion lying in a large polar fenestra (delimited by arrows) and a mostly continuous nuclear envelope with a few gaps (arrowhead). Scale bar, 0.5 μm. Reprinted from Mycologia ( Celio et al., 2007 ) with permission of the publisher.
Differential interference contrast LM of A. macrogynus illustrating saltatory motility of a mitochondrion (arrowhead) presented over a 1.5-s period. The Spitzenkörper is indicated by an asterisk. Bar, 5 μm. Reprinted from Fungal Genetics and Biology ( McDaniel and Roberson, 2000 ) with permission from the publisher.
Elements of the endomembrane system and organelles viewed with TEM. (a) S. rolfsii hyphal region III showing vacuoles (V), inflated cisternae of Golgi apparatus (white arrows), rough ER (arrowheads), MVBs (black arrows), and mitochondria (M). Bar, 1.0 μm. Reprinted from Protoplasma (Roberson and Fuller, 2000) with permission from the publisher. (b through e) Serial cross sections through Golgi equivalent of N. crassa showing apical vesicle (arrowheads), MT (arrows), and a mitochondrion (M). Bar, 0.1 μm. (f) Filasome (arrow) in cortex of N. crassa hypha. Bar, 0.12 μm. (g) MVB (arrowhead) juxtaposed to MT (arrow) in S. rolfsii. Bar, 0.1 μm. From Roberson and Fuller (1988), reproduced with permission from Springer. (h) Flattened vacuolar cisternae (arrows). White arrowheads indicate coated surfaces of vacuoles. Hypha is cut in cross section. Scale bar, 0.2 μm. (i) High magnification of panel h showing coated surfaces of vacuole (black arrow) and cross section of two closely associated MTs (white arrows). Scale bar, 75 nm.
Immunofluorescence of MTs in hyphal cells of S. rolfsii visualized with confocal microscopy. (a) Reconstruction of six longitudinal optical sections through hypha. MTs are primarily oriented parallel to the growing hyphal axis exhibiting helical curvatures forming a loosely braided meshwork. (b through g) Optical serial sections. MTs are in close proximity to Spitzenkörper but at the time of cell fixation were not traversing Spitzenkörper (asterisk in panel e). MT bundles are indicated by arrows. Scale bar, 3 μm.
Immunofluorescence of actin in hyphal cells of S. rolfsii visualized with wide-field epifluorescence microscopy. Actin is localized as cortical patches in high numbers beneath apical domes (arrowheads). Actin spot was not observed at the apex (arrow) in these cells. Scale bar, 7 μm. Reprinted from Mycologia ( Roberson, 1992 ) reproduced with permission from the publisher.
TEM of septal pore complexes. (a) Simple septal pore in a conidiation mutant of Fusarium verticillioides (Ascomycota). Woronin bodies are bound by a single-unit membrane (arrow). Note one of the Woronin bodies (asterisk) plugging the septal pore and the density difference between the cellular compartments above (apical) and below (distal) the cross wall. Scale bar, 300 nm. Courtesy of Beth Richardson and Tony Glenn. (b) Dolipore septal pore and septal pore cap (i.e., parenthosomes) (arrows) of A. vulgare (Basidiomycota) with the cap extending along the cross wall (arrowheads). Scale bar, 125 nm. Reprinted from Mycologia ( Celio et al., 2007 ) with permission of the publisher.