LAUSR

Simple Summary Understanding the evolutionary association between immune cells and the blood–brain barrier (BBB) is vital to develop therapeutic approaches. In Drosophila, glial cells form the BBB that regulates the access of hemocytes to the brain. It is still not known which diapedesis route hemocytes cells follow. In vertebrates, paracellular migration is dependent on PECAM1, while transcellular migration is dependent on the expression of CAV1. The drosophila genome lacks both genes. The Tre1 family (Tre1, moody, and Dmel_CG4313) contribute to regulating transepithelial migration in Drosophila. However, its evolutionary history is not known. We performed phylogenetic analysis to reconstruct the evolutionary history of the Tre1 family. We found Dmel_CG4313 only in insects. Tre1 exists only in invertebrates and is highly conserved. moody evolutionary history is more spread as it appears from Cnidaria up to mammals and is less conserved. The Tre1 family origin seems to be related to opsins. We have identified an SH3 motif in Tre1, moody, and Dmel_CG4313. SH3 regulates actin movement in a Rho-dependent manner in PECAM1. Our results suggest that the Tre1 family could be playing an important role in paracellular diapedesis in Drosophila. Thus, targeting the Tre1 family could help us regulate access to the brain. Abstract Understanding the evolutionary relationship between immune cells and the blood–brain barrier (BBB) is important to devise therapeutic strategies. In vertebrates, immune cells follow either a paracellular or a transcellular pathway to infiltrate the BBB. In Drosophila, glial cells form the BBB that regulates the access of hemocytes to the brain. However, it is still not known which diapedesis route hemocytes cells follow. In vertebrates, paracellular migration is dependent on PECAM1, while transcellular migration is dependent on the expression of CAV1. Interestingly Drosophila genome lacks both genes. Tre1 family (Tre1, moody, and Dmel_CG4313) play a diverse role in regulating transepithelial migration in Drosophila. However, its evolutionary history and origin are not yet known. We performed phylogenetic analysis, together with HH search, positive selection, and ancestral reconstruction to investigate the Tre1 family. We found that Tre1 exists in Mollusca, Arthropoda, Ambulacraria, and Scalidophora. moody is shown to be a more ancient protein and it has existed since Cnidaria emergence and has a homolog (e.g., GPCR84) in mammals. The third family member (Dmel_CG4313) seems to only exist in insects. The origin of the family seems to be related to the rhodopsin-like family and in particular family α. We found that opsin is the nearest receptor to have a common ancestor with the Tre1 family that has diverged in sponges. We investigated the positive selection of the Tre1 family using PAML. Tre1 seems to have evolved under negative selection, whereas moody has evolved during positive selection. The sites that we found under positive selection are likely to play a role in the speciation of function in the case of moody. We have identified an SH3 motif, in Tre1 and, moody and Dmel_CG4313. SH3 is known to play a fundamental role in regulating actin movement in a Rho-dependent manner in PECAM1. Our results suggest that the Tre1 family could be playing an important role in paracellular diapedesis in Drosophila.