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Effects of protein transport inhibitors on the distribution and secretion of the fusion protein RntA-EGFP in Aspergillus oryzae.

Bioscience, biotechnology, and biochemistry (2004-07-28)
Kumiko Masai, Jun-Ichi Maruyama, Harushi Nakajima, Katsuhiko Kitamoto
ABSTRACT

The distribution of the secreted protein ribonuclease T1 (RntA) fused with the enhanced green fluorescent protein (EGFP), RntA-EGFP, was visualized in hyphae of Aspergillus oryzae in the presence of a protein transport inhibitor, brefeldin A, cytochalasin A, or nocodazole. During treatment with the protein transport inhibitors, the distribution of RntA-EGFP changed and distinct patterns of fluorescence accumulation were observed. The addition of brefeldin A caused RntA-EGFP fluorescence to appear in reticular networks, and the disruption of the polymerization of actin filaments by cytochalasin A caused an increase in RntA-EGFP fluorescence intensity in the hyphae without accumulation in a specific cellular component. In contrast, RntA-EGFP fluorescence was distributed in different parts of a hypha during treatment with nocodazole, a compound that depolymerizes microtubules. In addition, quantitative analysis was performed using the RntA-EGFP visualization system to analyze the relative amount of RntA-EGFP secreted into the culture medium during treatment with the protein transport inhibitors.

MATERIALS
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Brand
Product Description

Sigma-Aldrich
Cytochalasin A from Drechslera dematioidea

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