Torralba Sara, Raudaskoski Marjatta, Pedregosa Ana María, Laborda Fernando
Departament de Microbiología y Parasitología, Universidad de Alcalá de Henares, Carretera Madrid-Barcelona, Km 33, E-28871 Alcalá de Henares, Madrid, Spain.
Division of Plant Physiology, Department of Biosciences, University of Helsinki, PO Box 56, Viikinkaari 9, FIN-00014, University of Helsinki, Finland.
Microbiology (Reading). 1998 Jan;144 ( Pt 1):45-53. doi: 10.1099/00221287-144-1-45.
The role of actin in apical growth and enzyme secretion in the filamentous fungus Aspergillus nidulans was studied by treating the hyphae with cytochalasin A (CA), which inhibits actin polymerization. Indirect immunofluorescence microscopy revealed actin at the tips of main hyphae and branches, and at the site of developing septa. CA inhibited the growth of the fungus and changed the growth pattern of hyphal tips from cylindrical tubes to spherical beads. The regions with swellings showed no actin fluorescence, and neither was actin seen in association with septa. After 4 h exposure, hyphae were able to resume the normal tip growth pattern in the presence of CA for a short period of time and new cylindrical hyphae, with actin fluorescence at the apex, emerged from the swollen tips. Later, the tips of the hyphae swelled again, which led to a beaded appearance. We also studied the effect of CA on the secretion of alpha- and beta-galactosidase. alpha-Galactosidase is secreted into the culture medium, whereas beta-galactosidase remains in the mycelium, with part of its activity bound to the cell wall. When A. nidulans mycelium was incubated in the presence of CA, a reduction in the secretion of alpha-galactosidase into the culture medium and a decrease in the alpha- and beta-galactosidase activities bound to the cell wall was detected. However, the CA dose used for the hyphae did not modify the secretion of the enzymes from protoplasts. Results described here provide evidence that a polymerized actin cytoskeleton is required for normal apical growth, hyphal tip shape and polarized enzyme secretion in A. nidulans. Cytochalasin-induced disruptions of the actin cytoskeleton could result in the alterations of apical growth and inhibition of enzyme secretion observed by blocking secretory vesicle transport to the apex.
通过用细胞松弛素A（CA）处理丝状真菌构巢曲霉的菌丝来研究肌动蛋白在其顶端生长和酶分泌中的作用，CA可抑制肌动蛋白聚合。间接免疫荧光显微镜检查显示，在主菌丝和分支的顶端以及正在形成隔膜的部位存在肌动蛋白。CA抑制了真菌的生长，并将菌丝顶端的生长模式从圆柱形管变为球形珠。出现肿胀的区域没有肌动蛋白荧光，在隔膜处也未观察到肌动蛋白。暴露4小时后，菌丝能够在CA存在的情况下短时间内恢复正常的顶端生长模式，并且从肿胀的顶端长出新的顶端有肌动蛋白荧光的圆柱形菌丝。后来，菌丝顶端再次肿胀，导致呈珠状外观。我们还研究了CA对α-和β-半乳糖苷酶分泌的影响。α-半乳糖苷酶分泌到培养基中，而β-半乳糖苷酶保留在菌丝体中，其部分活性与细胞壁结合。当构巢曲霉的菌丝体在CA存在下孵育时，检测到分泌到培养基中的α-半乳糖苷酶减少，以及与细胞壁结合的α-和β-半乳糖苷酶活性降低。然而，用于处理菌丝的CA剂量并未改变原生质体中酶的分泌。此处描述的结果提供了证据，表明聚合的肌动蛋白细胞骨架对于构巢曲霉的正常顶端生长、菌丝顶端形状和极化酶分泌是必需的。细胞松弛素诱导的肌动蛋白细胞骨架破坏可能通过阻止分泌囊泡向顶端转运而导致观察到的顶端生长改变和酶分泌抑制。
