1887

Abstract

Hydrophobins are small amphipathic proteins that function in a broad range of growth and developmental processes in fungi. They are involved in the formation of aerial structures, the attachment of fungal cells to surfaces, and act in signalling in response to surface cues and pathogenesis. is an important entomopathogenic fungus used as an arthropod biological control agent. To examine the feasibility of using phage display technology to clone cDNAs encoding hydrophobins, biopanning experiments were performed using a variety of affinity resins, including ,′-diacetylchitobiose-, fucose-, lactose-, maltose- and melibiose-coupled agarose beads. After five rounds of iterative biopanning, cDNAs corresponding to two (class I) hydrophobins were selectively enriched using melibiose- or lactose-coupled agarose beads. Expression analysis revealed that the gene was expressed in all samples tested, including aerial conidia, blastospores, submerged conidia, and cells sporulating on chitin and insect cuticle, with expression peaking in growing mycelia. In contrast, the gene was not appreciably expressed in any of the single-cell types (aerial conidia, blastospores and submerged conidia), but was constitutively expressed in growing mycelia and when cells were sporulating on chitin and insect cuticle. MS fingerprinting of an ∼10 kDa protein found in boiling SDS-insoluble, trifluoroacetic acid-soluble extracts from aerial conidia identified the major component of the rodlet layer to be the gene product. These results reveal the differential regulation of the isolated hydrophobins and indicate that phage display represents a novel approach to cDNA cloning of hydrophobins.

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2007-10-01
2019-10-20
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