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Position-dependent effects of regioisomeric methylated adenine and guanine ribonucleosides on translation.

ABSTRACT: Reversible methylation of the N6 or N1 position of adenine in RNA has recently been shown to play significant roles in regulating the functions of RNA. RNA can also be alkylated upon exposure to endogenous and exogenous alkylating agents. Here we examined how regio-specific methylation at the hydrogen bonding edge of adenine and guanine in mRNA affects translation. When situated at the third codon position, the methylated nucleosides did not compromise the speed or accuracy of translation under most circumstances. When located at the first or second codon position, N1-methyladenosine (m1A) and m1G constituted robust blocks to both Escherichia coli and wheat germ extract translation systems, whereas N2-methylguanosine (m2G) moderately impeded translation. While m1A, m2G and N6-methyladenosine (m6A) did not perturb translational fidelity, O6-methylguanosine (m6G) at the first and second codon positions was strongly and moderately miscoding, respectively, and it was decoded as an adenosine in both systems. The effects of methylated ribonucleosides on translation could be attributed to the methylation-elicited alterations in base pairing properties of the nucleobases, and the mechanisms of ribosomal decoding contributed to the position-dependent effects. Together, our study afforded important new knowledge about the modulation of translation by methylation of purine nucleobases in mRNA.

SUBMITTER: You C

PROVIDER: S-EPMC5587754 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Position-dependent effects of regioisomeric methylated adenine and guanine ribonucleosides on translation.

You Changjun C Dai Xiaoxia X Wang Yinsheng Y

Nucleic acids research 20170901 15

Reversible methylation of the N6 or N1 position of adenine in RNA has recently been shown to play significant roles in regulating the functions of RNA. RNA can also be alkylated upon exposure to endogenous and exogenous alkylating agents. Here we examined how regio-specific methylation at the hydrogen bonding edge of adenine and guanine in mRNA affects translation. When situated at the third codon position, the methylated nucleosides did not compromise the speed or accuracy of translation under ...[more]

PMID: 28591780

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Project description:We have characterized the regulation of phospholipase D (PLD) in electropermeabilized HL-60 granulocytes in which endogenous phospholipids were pre-labelled with [3H]oleic acid. Treatment of these permeabilized cells with the non-hydrolysable GTP analogues guanosine 5'-[gamma-thio]triphosphate (GTP[S]) and guanosine 5'-[beta gamma-imido]triphosphate induced a sustained (near-linear for up to 60 min) accumulation of phosphatidic acid (PA). In the presence of ethanol a sustained production of phosphatidylethanol (PEt) was also observed. With increasing concentrations of ethanol, PEt formation increased, whereas PA formation declined; this indicated involvement of a PLD-type effector enzyme. The ability of GTP[S] to stimulate this PLD activity was Mg(2+)-dependent and was inhibited by GDP and its non-hydrolysable beta-thio analogue. Ca2+, at concentrations less than or equal to nM, had no effect on the GTP[S]-dependent PLD activity. However, higher concentrations of Ca2+ produced a significant potentiation of this activity. Inclusion of MgATP (greater than or equal to 0.1 mM), but not other nucleoside triphosphates, also induced a large potentiation of GTP[S]-dependent PLD activation. In the absence of guanine nucleotides, MgATP elicited no significant activation of PLD. Significantly, this effect of ATP was not mimicked by adenosine 5'-[beta gamma-methylene]triphosphate, a non-hydrolysable ATP analogue. Rather, this analogue inhibited both basal and ATP-potentiated GTP[S]-dependent PLD activity. This suggests that the ability of ATP to potentiate GTP[S]-dependent PLD activity involves phosphotransferase action rather than simple allosteric effects induced by adenine nucleotide binding. The absolute magnitude of the GTP[S]-dependent PLD activity which could be potentiated by MgATP was decreased by 90% when the permeabilized cells were preincubated for various times before addition of these stimulatory agents. This time-dependent loss of MgATP-induced potentiation was prevented when the permeabilized cells were preincubated in the presence of GTP[S]. These results demonstrate that electropermeabilized HL-60 granulocytes can be used to discriminate synergistic roles for a GTP-binding protein(s) and an ATP-dependent process (kinase?) in the regulation of phospholipase D activity.

Dataset Information

Position-dependent effects of regioisomeric methylated adenine and guanine ribonucleosides on translation.

Publications

Position-dependent effects of regioisomeric methylated adenine and guanine ribonucleosides on translation.

Similar Datasets

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