The evolution of the vertebrate liver is a prime example of the evolution of complex organs, yet the driving genetic factors behind it remain unknown. Here we study the evolutionary genetics of liver by comparing the amphioxus hepatic caecum and the vertebrate liver, as well as examining the functional transition within vertebrates. Using in vivo and in vitro experiments, single-cell/nucleus RNA-seq data and gene knockout experiments, we confirm that the amphioxus hepatic caecum and vertebrate liver are homologous organs and show that the emergence of ohnologues from two rounds of whole-genome duplications greatly contributed to the functional complexity of the vertebrate liver. Two ohnologues, kdr and flt4, play an important role in the development of liver sinusoidal endothelial cells. In addition, we found that liver-related functions such as coagulation and bile production evolved in a step-by-step manner, with gene duplicates playing a crucial role. We reconstructed the genetic footprint of the transfer of haem detoxification from the liver to the spleen during vertebrate evolution. Together, these findings challenge the previous hypothesis that organ evolution is primarily driven by regulatory elements, underscoring the importance of gene duplicates in the emergence and diversification of a complex organ.

脊椎动物肝脏的进化是复杂器官进化的一个典型例子，但其背后的驱动遗传因素仍然未知。在这里，我们通过比较两栖动物盲肠和脊椎动物肝脏，以及检查脊椎动物内部的功能转变来研究肝脏的进化遗传学。利用体内和体外实验、单细胞/核 RNA-seq 数据和基因敲除实验，我们证实了两栖动物肝盲肠和脊椎动物肝脏是同源器官，并表明两轮全基因组重复中出现的 ohnologues 极大地促进了脊椎动物肝脏的功能复杂性。kdr 和 flt4 这两个 ohnologues 在肝窦内皮细胞的发育中起重要作用。此外，我们发现肝脏相关功能（如凝血和胆汁产生）是逐步进化的，其中基因复制起着至关重要的作用。我们重建了脊椎动物进化过程中血红素解毒从肝脏转移到脾脏的遗传足迹。总之，这些发现挑战了先前的假设，即器官进化主要由调节元件驱动，强调了基因复制在复杂器官的出现和多样化中的重要性。