Nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) are intracellular receptors that control innate immunity and other biotic interactions in animals and plants. NLRs have been characterized in plant and animal lineages, but in fungi, this gene family has not been systematically described. There is however previous indications of the involvement of NLR-like genes in nonself recognition and programmed cell death in fungi. We have analyzed 198 fungal genomes for the presence of NLRs and have annotated a total of 5,616 NLR candidates. We describe their phylogenetic distribution, domain organization, and evolution. Fungal NLRs are characterized by a great diversity of domain organizations, suggesting frequently occurring combinatorial assortments of different effector, NOD and repeat domains. The repeat domains are of the WD, ANK, and TPR type; no LRR motifs were found. As previously documented for WD-repeat domains of fungal NLRs, TPR, and ANK repeats evolve under positive selection and show highly conserved repeats and repeat length polymorphism, suggesting the possibility of concerted evolution of these repeats. We identify novel effector domains not previously found associated with NLRs, whereas others are related to effector domains of plant or animals NLRs. In particular, we show that the HET domain found in fungal NLRs may be related to Toll/interleukin-1 receptor domains found in animal and plant immune receptors. This description of fungal NLR repertoires reveals both similarities and differences with plant and animals NLR collections, highlights the importance of domain reassortment and repeat evolution and provides a novel entry point to explore the evolution of NLRs in eukaryotes.

The ability of Anopheles gambiae mosquitoes to transmit Plasmodium parasites is highly variable between individuals. However, the genetic basis of this variability has remained unknown. We combined genome-wide mapping and reciprocal allele-specific RNA interference (rasRNAi) to identify the genomic locus that confers resistance to malaria parasites, and demonstrate that polymorphisms in a single gene encoding the antiparasitic thioester-containing protein 1 (TEP1) explain a substantial part of the variability in parasite killing. The link between TEP1 alleles and resistance to malaria may offer new tools for controlling malaria transmission. The successful application of rasRNAi in Anopheles suggests that it could also be applied to other organisms where RNA interference is feasible to dissect complex phenotypes to the level of individual quantitative trait alleles.Anopheles gambiae mosquitoes are major vectors of Plasmodium falciparum, a protozoan parasite causing the most severe form of human malaria in Africa. The fact that mosquito strains completely wide variety of parasite species (1). Previous genetic analyses of crosses between this strain and the susceptible 4Arr strain infected with two simian parasite species focused on resistant to malaria parasites can be selected (1,2) indicates that genetic factors in mosquitoes control the level of parasite transmission. Understanding the genetic basis of this resistance has been a long-standing question. The L3-5 resistant strain kills and melanises a the melanotic encapsulation phenotype and identified several quantitative trait loci (QTLs), whose relative contributions varied with parasite species and between F2 generation families (3,4). Recently it became clear that melanisation occurs after parasite killing, as a means to dispose of dead parasites in some strains, while in others, killed parasites are only cleared by lysis (Fig. S2A) (5-7). In this study, we aimed at mapping the genomic regions and identifying genes that control resistance (i.e., the absence of live parasites) of mosquitoes to the rodent malaria parasite Plasmodium berghei.We set up reciprocal crosses of the resistant L3-5 and susceptible 4Arr strains. F1 mosquitoes were intercrossed and individual females were isolated to lay eggs, yielding 10 F2 families. ‡ To whom correspondence should be addressed. (9)), indicating that the killing of P. berghei and the mode of clearance of dead parasites are complex traits that are each likely to result from the segregation of several alleles.To map loci controlling resistance to parasites, we genotyped 39 informative markers spanning the entire genome in 206 selected F2 individuals with extreme phenotypes (Fig. S1 and supporting online text). Linkage analysis comparing resistant and non-resistant mosquitoes identified a single region on chromosome 3L (Fig. 1C). We interpreted this region, covering approximately 19Mb, as a major locus responsible for resistance to P. berghei, and named it Pbres1, for P. berghei resistance locus 1. We further c...

Recognition and response to non self is essential to development and survival of all organisms. It can occur between individuals of the same species or between different organisms. Fungi are established models for conspecific non self recognition in the form of vegetative incompatibility (VI), a genetically controlled process initiating a programmed cell death (PCD) leading to the rejection of a fusion cell between genetically different isolates of the same species. In Podospora anserina VI is controlled by members of the hnwd gene family encoding for proteins analogous to NOD Like Receptors (NLR) immune receptors in eukaryotes. It was hypothesized that the hnwd controlled VI reaction was derived from the fungal innate immune response. Here we analyze the P. anserina transcriptional responses to two bacterial species, Serratia fonticola to which P. anserina survives and S. marcescens to which P. anserina succumbs, and compare these to the transcriptional response induced under VI conditions. Transcriptional responses to both bacteria largely overlap, however the number of genes regulated and magnitude of regulation is more important when P. anserina survives. Transcriptional responses to bacteria also overlap with the VI reaction for both up or down regulated gene sets. Genes up regulated tend to be clustered in the genome, and display limited phylogenetic distribution. In all three responses we observed genes related to autophagy to be up-regulated. Autophagy contributes to the fungal survival in all three conditions. Genes encoding for secondary metabolites and histidine kinase signaling are also up regulated in all three conditions. Transcriptional responses also display differences. Genes involved in response to oxidative stress, or encoding small secreted proteins are essentially expressed in response to bacteria, while genes encoding NLR proteins are expressed during VI. Most functions encoded in response to bacteria favor survival of the fungus while most functions up regulated during VI would lead to cell death. These differences are discussed in the frame of a multilayered response to non self in fungi.

Anopheles gambiae mosquitoes are the major vectors of human malaria parasites. However, mosquitoes are not passive hosts for parasites, actively limiting their development in vivo. Our current understanding of the mosquito antiparasitic response is mostly based on the phenotypic analysis of gene knockdowns obtained by RNA interference (RNAi), through the injection or transfection of long dsRNAs in adult mosquitoes or cultured cells, respectively. Recently, RNAi has been extended to silence specifically one allele of a given gene in a heterozygous context, thus allowing to compare the contribution of different alleles to a phenotype in the same genetic background.