PT  - JOURNAL ARTICLE
AU  - Orús-Alcalde, Andrea
AU  - Lu, Tsai-Ming
AU  - Hejnol, Andreas
TI  - The evolution of the metazoan Toll receptor family and its expression during protostome development
AID  - 10.1101/2021.02.01.429095
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.02.01.429095
4099  - http://biorxiv.org/content/early/2021/02/02/2021.02.01.429095.short
4100  - http://biorxiv.org/content/early/2021/02/02/2021.02.01.429095.full
AB  - Background Toll-like receptors (TLRs) play a crucial role in immunity and development. They contain leucine-rich repeat domains, one transmembrane domain, and one Toll/IL-1 receptor domain. TLRs have been classified into V-type/scc and P-type/mcc TLRs, based on differences in the leucine-rich repeat domain region. Although TLRs are widespread in animals, detailed phylogenetic studies of this gene family are lacking. Here we aim to uncover TLR evolution by conducting a survey and a phylogenetic analysis in species across Bilateria. To discriminate between their role in development and immunity we furthermore analyzed stage-specific transcriptomes of the ecdysozoans Priapulus caudatus and Hypsibius exemplaris, and the spiralians Crassostrea gigas and Terebratalia transversa.Results We detected a low number of TLRs in ecdysozoan species, and multiple independent radiations within the Spiralia. V-type/scc and P-type/mcc type-receptors are present in cnidarians, protostomes and deuterostomes, and therefore they emerged early in TLR evolution, followed by a loss in xenacoelomorphs. Our phylogenetic analysis shows that TLRs cluster into three major clades: clade α is present in cnidarians, ecdysozoans, and spiralians; clade β in deuterostomes, ecdysozoans, and spiralians; and clade γ is only found in spiralians. Our stage-specific transcriptome and in situ hybridization analyses show that TLRs are expressed during development in all species analyzed, which indicates a broad role of TLRs during animal development.Conclusions Our findings suggest that the bilaterian TLRs likely emerged by duplication from a single TLR encoding gene (proto-TLR) present in the last common cnidarian-bilaterian ancestor. This proto-TLR gene duplicated before the split of protostomes and deuterostomes; a second duplication occurred in the lineage to the Trochozoa. While all three clades further radiated in several spiralian lineages, specific TLRs clades have been presumably lost in others. Furthermore, the expression of the majority of these genes during protostome ontogeny suggests their involvement in immunity and development.Competing Interest StatementThe authors have declared no competing interest.AMPsAntimicrobial peptidesBCIP5-bromo-4-chloro-3-indolyl phosphateBUSCOBenchmarking Universal Single-Copy OrthologsIL-1Interleukin-I receptorLRRLeucine-rich repeatLRRCTLeucine-rich repeat C-terminal domainLRRNTLeucine-rich repeat N-terminal domainMccmultiple cysteine clusterNBTnitro blue tetrazoliumNODNucleotide oligomerization domainPBSPhosphate-Buffered SalineP-typeProtostome typePTwPBS with 0.1% Tween® 20RSEMRNA-Seq by Expectation MaximizationSccSingle cysteine clusterTIRToll/IL-1 receptorTLRToll-like receptorTLR-Cαclade α type TLR geneTLR-Cβclade β type TLR geneTLR-Cγclade γ type TLR geneTLR-Cβ/γproto-TLR gene of clades β and γTLR-likeToll-like receptor-likeTMTransmembraneTMMTrimmed mean of M valuesTPMTranscripts per millionV-typeVertebrate typeWMISHWhole mount in situ hybridization