Studies on the evolutionary pathway of the insulin gene family suggest that insulin and insulin-like growth factor (IGF) became distinct molecules only after the vertebrates arose. A single molecule with identity to both insulin and IGF was reported in amphioxus. To study the origin of insulin, we selected tunicates because their ancestors are thought to be a nodal point in the evolution of vertebrates. This is the first report of separate insulin and IGF molecules from invertebrates. Two cDNAs were isolated from the tunicateChelyosoma productum: One cDNA encodes a distinct preproinsulin with B, C, and A domains, whereas the other encodes tunicate preproIGF, including all five domains in their proper sequence. Both mRNAs are expressed in the nervous system, digestive tract, heart, and possibly the gonad but not in branchial basket or tunic. Hence,insulinandigfgenes have similar expression patterns.In situmethods confirm the polymerase chain reaction evidence that tunicateinsulinandigfmRNAs are expressed in cortical cells of the neural ganglion. We conclude that insulin and IGF have maintained separate gene lineages in both vertebrate and protochordate evolution and, thus, a distinct evolutionary history of more than 600 million years.
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