## 3 Results and Discussion
### Groups NR-III and NR-IV
#### 3.3.3 Fungal PKSs Producing Reduced Polyketides  
The Diversity of R-PKS Sequences  
R-PKS genes are involved in synthesizing various reduced, usually linear polyketides. Many are precursors of toxins that are active in animals (e.g., lovastatin, citrinin, and fumonisin) or plants (e.g., T-toxin and PM-toxin) [96; 97; 98; 99]. We follow this classification in our discussion whenever possible, while also highlighting deviations. Compared to NR-PKSs, less is known about the possible roles of R-PKSs. In our phylogeny, R-PKSs formed a large clade sister to NR-PKSs with many well-supported subclades. Unlike what has been shown in previous studies, all studied R-PKSs here lacked a second ACP domain, and in some cases, ACP was missing. According to [PERSON] et al. [14], four major groups (R-I to R-IV) of R-PKS genes can be distinguished based on their overall domain structure and produced compounds. This classification was later extended to groups R-V to R-VIII by [PERSON] et al. [72]. Over 70% of the studied fungal sequences could be assigned to groups designated by [PERSON] et al. [14] and [PERSON] et al. [72] with the general domain content: KS-AT-DH-[CMeT]-ER-KR-[ACP]. Two clades recovered by [PERSON] et al. [72] are missing from our tree, namely R-VI and R-VIII. Clade VI contained uncommon PKS-NRPS hybrids with four additional domains after ACP, viz. condensation (C), adenylation (A), thiolation (T), and reductase (R) [72]. As our study focused on the TI-PKSs, it did not include hybrid NRPS-PKS genes. The sequences from R-VIII were not present in our phylogeny, and are thus not feasible to identify with confidence. Based on our phylogenetic results, we identified two R-PKS clades that did not match any of the R-PKS groups reported in previous studies. Sequences in those two clades had characteristic domain structures and were recovered as distinct orthogroups in our Orthofinder analysis. This leads us to propose these clades as new reducing PKS groups, R-IX and R-X, following the numbering scheme by [PERSON] et al. [14] and [PERSON] et al. [72]. Group R-I is the largest group of R-PKSs in our tree. It contains one of the R-PKS genes synthesizing the diketide portion of lovastatin and citrinin, T-toxin, and PM-toxin [14]. Our results show that the genes of this group, recovered in a monophyletic clade by [PERSON] et al. [14], were nested in different clades in our analysis; therefore, R-I assignment is not complete. Group R-II forms a well-supported clade and it was previously characterized by the absence of an ER domain [14]. In our phylogeny, however, ER domain was present in all lichen-forming fungi. The group includes many previously published sequences and orthologs from _Cladonia_ and _Letharia_ species, _Xanthoria elegans_, _Pseudevernia furfuracea_, and _Ramalina internedia_. The _R. internedia_ orthologos was assigned to PKS19. It was predicted to be involved in the production of heptaketides with a role in a lesion of rice leaves formation in _Magnaporte oryzae_[100]. Two characterized proteins from _A. terreus_ lovB and _P. citriumum_ mlcA synthesize the cyclic nonaketide portion of lovastatin and citrinin. Group R-III forms a clade comprised of four sequences previously assigned to this group by [PERSON] et al. [14], and a sequence from _Cyanodemella asteris_. PKSs from this clade do not possess a CMeT domain. The experimentally characterized gene in group R-III is _C. heterostrophilus_ PKS2, which, along with PKS1, is required for the synthesis of T-toxin [14]. This group contains several sequences from [PERSON] et al. [14] interspersed with sequences from various lichen-forming fungi. Genes belonging to R-IV may contain a conserved CMeT domain. Thus, their domain configuration somewhat resembles that of groups R-I and R-II. However, N-terminal domains ER, KR, and ACP can be present or absent in R-IV. The only characterized PKS in this group is _G. moniliformis_ FUM1 (Gm_FUM1_AAD43562), which makes the linear polyketide precursor of the toxin fumonisin [98].