As these putative GPCRs represented a separate clade in the phylogenetic analysis (Figure 1), they were assigned to a new class (class XIII, Table 1) thereby extending the classification system of fungal GPCRs to 14 classes. Conclusions A thorough examination of the genomes of the two mycoparasites T. atroviride and T. virens and the saprophyte T. reesei for putative GPCRs revealed for most classes a high conservation of their number and structure within this genus. On the other hand, remarkable differences in individual classes were found among the three Trichoderma species and among Trichoderma and other filamentous fungi.

Whereas for class Ralimetinib molecular weight I to VII members, orthologous triplets with similar length and sequence are present in the genomes of the three Trichoderma species and their number is also similar to other fungi, the PAQR family has expanded especially in T. atroviride. Considering the identification of members of classes X, XI, and XII and proteins similar to the P. sojae GPR11 receptor in Trichoderma, the presented 14 classes now define the most comprehensive classification system for GPCR-like proteins of fungi. The huge diversity of GPCRs in Trichoderma spp. and especially in the mycoparasites is likely to reflect the capability of these fungi to establish various ecological niches and interactions with other organisms. It is worth mentioning that Vactosertib clinical trial with the exception

of few members, the proteins identified as putative GPCRs in this study have only been characterized in silico. Taking into account that only three α, one β and one γ subunit of heterotrimeric

G proteins are encoded in the Trichoderma genomes which face more than 55 GPCRs, studying the signaling output and identifying the respective intracellular until interaction partners of those receptors will provide interesting insights on how these fungi adapt to their different lifestyles. Methods Identification of GPCR-encoding genes of Trichoderma atroviride and Trichoderma virens Version 2 of the T. atroviride genome database [57] comprises 11,863 gene models on 29 scaffolds; version 2 of the T. virens genomic sequence [58] comprises 12,427 gene models on 93 scaffolds. For the homology-based PI3K inhibitor search of GPCR-like proteins from T. atroviride and T. virens, the genomic sequences and deduced proteomes of the following fungi were used: Trichoderma reesei[59]Aspergillus nidulans, Aspergillus fumigatus, Aspergillus oryzae[62], Neurospora crassa[63], Magnaporthe grisea[64], Podospora anserine[65], Chaetomium globosum[66], Fusarium graminearum[67], and Nectria haematococca[68]. An e-value limit of 1e-09 was applied. To identify putative GPCRs within the T. atroviride and T. virens proteomes that lack significant sequence similarity to known GPCR-like proteins and therefore may escape detection by homology search, a more sensitive database searching using hidden Markov models (HMM) was performed using the program HMMER (http://​hmmer.​janelia.