P loop-NTPases in developmental processes of Coprinopsis cinerea

Shanta Subba Weeradej Khonsuntia Ursula Kües
Molecular Wood Biotechnology and Technical Mycology, University of Goettingen, Goettingen, Germany

The self-compatible homokaryotic strain AmutBmut with defects in both mating-type loci is a mutant of Coprinopsis cinerea for studying fruiting body development. Proto159 is a derived mutant of AmutBmut with a defect in the formation of primary hyphal knots (PK) as a first step of light-regulated fruiting body development and in sclerotia formation in the dark. Proto159 shows a somewhat slower growth rate and pigments the mycelium and the agar beneath dark-brown. The mutation in the defective strain Proto159 was found to be suppressed by transformation of a specific gene that belongs to the NWD2 family. NWD2 genes encode proteins with an N-terminal NACHT domain which is an evolutionary conserved domain that serves in signal transduction and is named after four different types of P-loop NTPases (NAIP, CIITA, HET-E and TP1). The NACHT domain contains a typical NTP binding site (P-loop, Walker A motif), a Walker B motif, a charged amino acid at a specific conserved position, a GRRxE motif, and a GxP motif of STAND NTPases. NWD2 genes exist only in some Agaricales (e.g. Amanita muscaria, Agaricus bisporus, Moniliophthora roreri, and Laccaria bicolor) including C. cinerea where they have been multiplied to represent in total 36 genes, most of which are seen to locate in transposon-rich and telomere regions of chromosomes. All the encoded proteins have the N-terminal NACHT domains while these have variably been fused to different C-terminal protein halves (e.g. tandem WD40 repeats). More NWD2 genes were subcloned and used in transformations of Proto159. Introduction of some genes altered mycelial properties, blocked the brown staining of mycelium and the agar, and induced primary hyphal knot and sclerotia formation and sometimes fruiting.