Functional specialization of cytochrome b5 prior to the invention of syringyl lignin biosynthesis

Abstract

The production of lignin represents a hallmark of vascular plant evolution. But syringyl-lignin production is lineage specific. Its emergence in angiosperms has been regarded as a recent evolution event, as is the appearance of syringyl monomer biosynthetic F5H (CYP84A1). Angiosperm F5H uniquely requires cytochrome b5 protein CB5D as an obligatory redox partner. However, it remains unclear how CB5D functionally evolved and specialized and whether it co-evolved with F5H. Here we show that plant CB5 family expanded and functionally specialized at the early stage of plant terrestrialization. CB5D was absent in green algae but emerged in bryophytes and conserved in most embryophytes, particularly proliferating in angiosperms. The innovation of CB5D is attributed to the evolution and maintenance of acidic amino residues in the helix 5 of CB5 protein. Interestingly, the syringyl-lignin producing lycophyte Selaginella lacks the functionally specialized CB5D. The Selaginella F5H (CYP788A1) solely relies on cytochrome P450 reductase as a redox partner. These results suggest that CB5 functionally diversified and specialized prior to the divergence of land plant lineages but lost in Selaginella; and the recently evolved angiosperm F5H has co-opted the anciently invented CB5D to form a modern cytochrome P450 monooxygenase system enabling the re-emergence of syringyl-lignin biosynthesis in higher plants. One Sentence SummaryCytochrome b5 D, an indispensable electron donor for F5H-catalyzed S-lignin biosynthesis, was invented prior to the divergence of land plant lineages, earlier than the emergence of vascular plant F5Hs.