tRNA is one of the main RNA molecules, and involves in protein synthesis. Among all cellular RNAs, tRNAs contain the greatest number of modified nucleosides, and some modifications are essential for the function of tRNA and lacking of these modifications could cause severe cellular deficiency and result in many human diseases. The acceptor stem of tRNA is seldom modified, while methylation of cytosine at the fifth carbon atom, 5-methylcytosine (m5C) is one of the several modifications in this region. tRNA m5C72 modification presents in higher eukaryotes and is strictly conserved, implying that this modification plays an important role in higher eukaryotes. The NSun family members are the main RNA m5C MTases in higher eukaryotes. Recently, the NSun6 was reported as the enzyme that responsible for them5C72 modification of tRNA. However, the mechanism of RNA recognition by NSun6 remains elusive.
Under the guidance of Prof. WANG Enduo and Dr. LIU Rujuan, Ph.D students LONG Tao et al. identified the tRNA recognition elements by human NSun6 (hNsun6) through mass spectrometry assays, site directed mutagenesis, and enzymatic kinetic studies. They found that hNSun6 recognizes RNA tertiary structure and specific bases or base pairs in the acceptor stem and D stem. In summary, hNSun6 recognizes substrate tRNAs based on (i) a well-folded, full-length tRNA, (ii) the CCA terminus at 3’ end and the methylation site C72, (iii) the discriminator base U73, (iv) the second and third base pairs of the acceptor stem, and (v) two base pairs (11:24 and 12:23) in the D-stem. Our results showed that hNSun6 uses a complicated network for RNA recognition and that the tertiary structure and some characteristic elements of the tRNA are all required. Other cellular RNAs are unlikely to satisfy all the elements required for hNSun6 recognition; thus, we conclude that hNSun6 is a tRNA-specific MTase.
This study entitled “Sequence-specific and Shape-selective RNA Recognition by the Human RNA m5C Methyltransferase NSun6” was published online in the Journal of Biological Chemistry on October 4th, 2016. The work is supported by grants from the National Key Basic Research Foundation of China, the National Natural Science Foundation of China, Chinese Academy of Sciences（CAS）, and the Youth Innovation Promotion Association from CAS (to R-J Liu).
CONTACT:
WANG En-Duo, Principal Investigator
Institute of Biochemistry and Cell Biology, Shanghai Institutes of Biological Sciences, Chinese Academy of Sciences
Shanghai 200031, P. R. China
Phone: 86-21-54921241
E-mail: edwang@sibcb.ac.cn
Figure: A scheme to summarize the determinant elements of tRNA recognized by NSun6. (Image provided by Prof. WANG Enduo’s group)
Key Word: RNA, RNA modification, 5-methylcytosine
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