Department of Plant Pathology, University of Minnesota, St. Paul, MN 55108, USA.
Fungal Genet Biol. 2011 Jul;48(7):725-30. doi: 10.1016/j.fgb.2011.03.001. Epub 2011 Mar 21.
The ortholog of the human gene NPC1 was identified in the plant pathogenic, filamentous fungus Fusarium graminearum by shared amino acid sequence, protein domain structure and cellular localization of the mature fungal protein. The FusariumNpc1 gene shares 34% amino acid sequence identity and 51% similarity to the human gene, has similar domain structure and is constitutively expressed, although up-regulated in ungerminated macroconidia and ascospores. GFP-tagged Npc1p localizes to the fungal vacuolar membrane. Cultures derived from a Δnpc1 mutant strain contain significantly more ergosterol than cultures of the wildtype. Staining with the fluorescent, sterol binding dye filipin, shows that ergosterol accumulates in vacuoles of the Δnpc1 mutant but not the wildtype strain. The Δnpc1 mutant has a temperature dependent reduction in growth and greater sensitivity to the ergosterol synthesis inhibiting fungicide tebuconazole compared with the wildtype strain or the mutant complemented with wildtype Npc1. The mutant also is significantly reduced in pathogenicity to wheat. Our results are consistent with the interpretation that Npc1p is important for normal transport of ergosterol from the vacuole and is essential for proper membrane function under particular environmental conditions.
通过氨基酸序列、成熟真菌蛋白的结构域结构和细胞定位的共享，在植物病原丝状真菌禾谷镰刀菌中鉴定出人类 NPC1 基因的直系同源物。禾谷镰刀菌 Npc1 基因与人类基因的氨基酸序列同一性为 34%，相似性为 51%，具有相似的结构域结构，并且是组成型表达的，尽管在未萌发的大分生孢子和子囊孢子中上调。GFP 标记的 Npc1p 定位于真菌液泡膜上。与野生型相比，来自 Δnpc1 突变株的培养物中含有明显更多的麦角固醇。用荧光固醇结合染料 filipin 染色表明，ergosterol 在Δnpc1 突变体的液泡中积累，但在野生型菌株中没有。与野生型菌株或用野生型 Npc1 互补的突变体相比，Δnpc1 突变体在生长方面具有温度依赖性降低，并且对麦角固醇合成抑制剂特比萘芬的敏感性更高。与野生型相比，该突变体对小麦的致病性也显著降低。我们的结果与以下解释一致，即 Npc1p 对于麦角固醇从液泡的正常运输很重要，并且在特定环境条件下对于适当的膜功能是必不可少的。
