Catabolism of non-canonical nucleosides from RNA turnover in plants
Plant Physiology
Final Report Abstract
In addition to the four canonical nucleosides adenosine (A), guanosine (G), cytidine (C) and uridine (U), ribonucleic acids (RNA) contain several chemically modified nucleosides, for example N6-methyl adenosine (m6A), pseudouridine (Ψ), 5-methyl cytidine (m5C) and 5-methyl uridine (m5U). These play a role in RNA stability and regulation. Degradation of RNA releases modified nucleosides and nucleotides (monophosphorylated nucleosides) whose metabolic fate is unclear. How are these degraded and building blocks possibly recycled? These were the overarching questions of this research proposal, focusing in particular on the degradation of Ψ, m5C, and m5U in Arabidopsis thaliana. Two enzymes required for the degradation of free pseudouridine were biochemically characterized and their biological role was investigated by targeted metabolite analysis of plant expression variants of these enzymes. It was shown that pseudouridine, which is released during degradation of Ψ-containing RNA, is initially phosphorylated to ΨMP. The atomic structure of the pseudouridine kinase that catalyzes this reaction was also elucidated to better understand the specificity of the enzyme for pseudouridine. It was shown that ΨMP is highly toxic and is promptly metabolized to uracil and 5-phosphoribose, which enter central metabolism. These transformations occur in the peroxisome, probably to limit the toxic effect of ΨMP. For the degradation of free 5-methylcytidine (5mC) and 5-methyluridine (5mU), the cell has no specific enzymes, but these modified nucleosides are metabolized by the same enzymes that degrade the non-methylated variants. In this process, Cytidine Deaminase converts 5mC to 5mU, which is then metabolized by Nucleoside Hydrolase 1 to thymine (5-methyl uracil) and ribose. Much of the 5mU comes directly from RNA degradation and is not generated from 5mC because RNA contains substantial amounts of m5U (about 1% m5U/U). Therefore, the nucleobase thymine arises in metabolism primarily from RNA degradation and not from DNA degradation, where this base occurs instead of uracil. The bases uracil and thymine are then further degraded in pyrimidine ring catabolism, although exactly how this degradation pathway merges with central metabolism remains unclear today. A defect in the degradation of 5mU resulted in accumulation of m5U in RNA and growth retardation in early development. Thus, degradation of 5mU is also in protecting RNA from spurious methylation. In summary, this project elucidated the degradation pathways for pseudouridine, 5mC, and 5mU. For the methylated pyrimidines in particular, this is the first study in eukaryotes, indicating that in the metabolism of other eukaryotes, for example human, there is still a significant gap in knowledge, although their RNA also contains m5C and m5C.
Publications
-
Functions and Dynamics of Methylation in Eukaryotic mRNA. RNA Technologies, 333-351. Springer International Publishing.
Chen, Mingjia & Witte, Claus-Peter
-
A Kinase and a Glycosylase Catabolize Pseudouridine in the Peroxisome to Prevent Toxic Pseudouridine Monophosphate Accumulation. The Plant Cell, 32(3), 722-739.
Chen, Mingjia & Witte, Claus-Peter
-
Structural basis for the substrate specificity and catalytic features of pseudouridine kinase from Arabidopsis thaliana. Nucleic Acids Research, 49(1), 491-503.
Kim, Sang-Hoon; Witte, Claus-Peter & Rhee, Sangkee
-
N4-acetylation of cytidine in mRNA plays essential roles in plants. The Plant Cell, 35(10), 3739-3756.
Wang, Wenlei; Liu, Huijie; Wang, Feifei; Liu, Xiaoye; Sun, Yu; Zhao, Jie; Zhu, Changhua; Gan, Lijun; Yu, Jinping; Witte, Claus-Peter & Chen, Mingjia
-
Pyrimidine catabolism is required to prevent the accumulation of 5-methyluridine in RNA. Nucleic Acids Research, 51(14), 7451-7464.
Gao, Shangyu; Sun, Yu; Chen, Xiaoguang; Zhu, Changhua; Liu, Xiaoye; Wang, Wenlei; Gan, Lijun; Lu, Yanwu; Schaarschmidt, Frank; Herde, Marco; Witte, Claus-Peter & Chen, Mingjia
