About 500 million years ago, a new type of adaptive immune defense emerged in basal jawed vertebrates, accompanied by morphological innovations, including the thymus. Did these evolutionary novelties arise de novo or from elaboration of ancient genetic networks? We reconstructed the genetic changes underlying thymopoiesis by comparative genome and expression analyses in chordates and basal vertebrates. The derived models of genetic networks were experimentally verified in bony fishes. Ancestral networks defining circumscribed regions of the pharyngeal epithelium of jawless vertebrates expanded in cartilaginous fishes to incorporate novel genes, notably those encoding chemokines. Correspondingly, novel networks evolved in lymphocytes of jawed vertebrates to control the expression of additional chemokine receptors. These complementary changes enabled unprecedented Delta/Notch signaling between pharyngeal epithelium and lymphoid cells that was exploited for specification to the T cell lineage. Our results provide a framework elucidating the evolution of key features of the adaptive immune system in jawed vertebrates.

Significance Lampreys and hagfishes (cyclostomes) are the only living group of jawless vertebrates and therefore are important for the study of vertebrate evolution. We have characterized Hox clusters in the Japanese lamprey ( Lethenteron japonicum ), and shown that it contains at least six Hox clusters as compared with four Hox clusters in tetrapods. This suggests that the lamprey lineage has undergone an additional round of genome duplication compared with tetrapods. Several conserved noncoding elements (CNEs) were predicted in the Hox clusters of lamprey, elephant shark, and human. Transgenic assay of CNEs demonstrated their potential to function as cis -regulatory elements. Thus, these CNEs may represent part of the core set of cis -regulatory elements that were present in the common ancestor of vertebrates.

With our increasing ability for generating whole-genome sequences, comparative analysis of whole genomes has become a powerful tool for understanding the structure, function, and evolutionary history of human and other vertebrate genomes. By virtue of their position basal to bony vertebrates, cartilaginous fishes (class Chondrichthyes) are a valuable outgroup in comparative studies of vertebrates. Recently, a holocephalan cartilaginous fish, the elephant shark, Callorhinchus milii (Subclass Holocephali: Order Chimaeriformes), has been proposed as a model genome, and low-coverage sequence of its genome has been generated. Despite such an increasing interest, the evolutionary history of the modern holocephalans-a previously successful and diverse group but represented by only 39 extant species-and their relationship with elasmobranchs and other jawed vertebrates has been poorly documented largely owing to a lack of well-preserved fossil materials after the end-Permian about 250 Ma. In this study, we assembled the whole mitogenome sequences for eight representatives from all the three families of the modern holocephalans and investigated their phylogenetic relationships and evolutionary history. Unambiguously aligned sequences from these holocephalans together with 17 other vertebrates (9,409 nt positions excluding entire third codon positions) were subjected to partitioned maximum likelihood analysis. The resulting tree strongly supported a single origin of the modern holocephalans and their sister-group relationship with elasmobranchs. The mitogenomic tree recovered the most basal callorhinchids within the chimaeriforms, which is sister to a clade comprising the remaining two families (rhinochimaerids and chimaerids). The timetree derived from a relaxed molecular clock Bayesian method suggests that the holocephalans originated in the Silurian about 420 Ma, having survived from the end-Permian (250 Ma) mass extinction and undergoing familial diversifications during the late Jurassic to early Cretaceous (170-120 Ma). This postulated evolutionary scenario agrees well with that based on the paleontological observations.