The expression of MpPRIA1encoding a putative aegerolysin, 4SC-202 cell line NVP-LDE225 chemical structure decreased in the yellow- and reddish pink-mycelium phases, and also before stress, but increased 4.3-fold in mycelia with primordia, and about 90-fold in the basidiomata, compared to the white mycelium stage (Figure 6A). The expression of the putative hemolysin-encoding gene MpPRIA1 increased 17-fold at the reddish pink mycelium stage, but decreased 11-fold before stress, 4-fold in stressed mycelia,

and 47.4-fold in mycelia with primordia. The transcripts of MpPRIA2 increased 23-fold in basidiomata, but were lower in mycelia with primordia (Figure 6B). The transcripts of gene MpPLYB, corresponding to a pleurotolysin B, increased 1.4-fold in the yellow mycelium stage, 15.2-fold in reddish pink mycelia, and remained at high levels in the mycelia before stress (11.7-fold), when stressed (11.2-fold) and in mycelia with primordia (10.1-fold), but decreased in basidiomata, where it was only 1.6 times higher than in

white mycelia (Figure 6C). Hemolysins, already identified in some bacteria and fungi, comprise a cytolytic protein family, Proteasome inhibitors in cancer therapy whose members appear abundantly during primordia and basidiomata formation [47, 58, 61, 62]. MpPRIA1 and MpPRIA2 have homologous regions but seem to correspond to two individual genes whose expression coincides with the morphological differentiation of primary hyphal nodules from primordia. These hemolysins may contribute to the process of hyphal aggregation

[61] as their expression occurred, although at low levels, before the appearance of primordia, when hyphae became globose for the formation of the “”initials”". This stage coincides with the reddish pink mycelium stage, where hyphal nodules are detectable. The exact function of these proteins remains unclear, but their Non-specific serine/threonine protein kinase involvement in programmed cell death (PCD), as proposed by Kues and Liu [17], seems rather unlikely because ostreolysins have lytic function, acting in cholesterol- and sphingomyelin-containing membranes [63] at a pH between 7 and 8 [64], which is not usually found in fungal cells. The known fungal hemolysins have some variations in amino acid sequences, but all share the conserved domain aegerolysin (code PF06355 by Pfam database [65]). Aegerolysin Aa-Pri1 from A. aegerita has the same molecular weight as the 16 kDa ostreolysin of P. ostreatus and is mainly expressed in the initial stage of primordium formation. PriA (or pleurotolysin or PlyA) of P. ostreatus forms a subfamily with the aegerolysin superfamily, which includes the Asp-hemolysins of Aspergillus fumigatus, and some hypothetical proteins of Clostridium bifermentans, P. aeruginosa and Neurospora crassa. P.