Biochemical Characterization of Arabidopsis Enzymes Involved in Inositol Pyrophosphate Biosynthesis

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Virginia Tech


To compensate for the sessile nature of plants, thousands of years of evolution have led to the development of many sophisticated signaling pathways that help plants sense and respond appropriately to different environmental cues. One such signaling pathway is called inositol phosphate signaling. This research dissertation focuses on the inositol phosphate signaling pathway in plants, with emphasis on elucidating how a new class of signaling molecules collectively referred to inositol pyrophosphates are synthesized. Inositol pyrophosphates are an emerging class of "high-energy" intracellular signaling molecules containing one or two diphosphate groups attached to an inositol ring, with suggested roles in bioenergetic homeostasis and inorganic phosphate sensing. Information regarding the biosynthesis of this unique class of signaling molecules in plants is scarce, however the enzymes responsible for their biosynthesis in other eukaryotes have been well described. This work aims to characterize the biochemical activity of the kinase domain (KD) of the Arabidopsis plant diphosphoinositol pentakisphosphate kinase enzymes (AtVIP1 and AtVIP2), and elucidate the biosynthesis pathway of inositol pyrophosphates in plants. Our data indicate that AtVIP1-KD and AtVIP2-KD function primarily as diphosphoinositol pentakisphosphate 5 kinases that phosphorylate this substrate at the 1-position. We also discovered a previously unreported inositol hexakisphosphate kinase activity for the Arabidopsis inositol(1,3,4) triphosphate 5/6kinase enzymes, that can convert InsP6 to InsP7. Together, these enzymes can function in plants to produce inositol pyrophosphates, which have been implicated in signal transduction and phosphate sensing pathways. The significance and potential application of these findings in terms of reduced phytate content and phosphate pollution, improved plant fitness, and improved nutrient use efficiency are discussed. The future outlook of inositol phosphate signaling research is also discussed.



Inositol, Inositol Pyrophosphates, Biosynthesis, Inositol phosphate signaling, Inositol phosphate kinases, PPIP5K