Ice-Binding Proteins of the Arctic Chloromonas induced by low temperature: its rarity and the association with the cold-signaling pathway

Abstract

Polar microalgae survive in extreme environments and play an important role as a primary producer in the regions. Chloromonas is abundantly found in the cryosphere but the cryoprotective mechanism and the cold signaling pathway of this genus are still obscure. Ice-Binding Proteins (IBPs) have been characterized by binding affinity to the ice surface, but the structure and mechanical activity of the proteins are diverse. In Chlamydomonadales, two kinds of IBPs are present: the general form (Type I IBP), which is conserved from prokaryotes to eukaryotes, and the rare form (Type II IBP), which limited to some Chloromonas species. In this study, we revealed six IBP genes (Type II IBP) in the Arctic Chloromonas sp. KNF0032 (called CmIBPs) were progressively expressed by temperature decreasing. These CmIBP proteins showed the effective ice recrystallization inhibition (IRI) activities in vitro as well as in planta expression system. It is the first example in which plant improves freezing tolerance by expressing the heterologous IBP proteins instead of cold acclimation. In addition, the cis-element analysis on the gene regulatory regions suggested that CmIBP protein induction could be regulated similarly to the plant cold-signaling pathway. Point to the rarity of these proteins, interestingly, the homologous sequences with CmIBPs were not found in any database, but we found that the AidA domain of Archaeal adhesin protein has a similar motif structure to CmIBPs (the repeated Thr-X-Thr), and these motifs also have effective IRI activity. This result gives an insight into that the engineering of this motif structure can fine-tune the strength of IRI activity. Possibilities of both CmIBP protein expression in planta system and fin-tuning of IRI activity are expected to be applicable especially crops and food industries.