Reference: Chen C, et al. (2011) Unexpected accumulation of ncm(5)U and ncm(5)S(2) (U) in a trm9 mutant suggests an additional step in the synthesis of mcm(5)U and mcm(5)S(2)U. PLoS One 6(6):e20783

Reference Help

Abstract


Background: Transfer RNAs are synthesized as a primary transcript that is processed to produce a mature tRNA. As part of the maturation process, a subset of the nucleosides are modified. Modifications in the anticodon region often modulate the decoding ability of the tRNA. At position 34, the majority of yeast cytosolic tRNA species that have a uridine are modified to 5-carbamoylmethyluridine (ncm(5)U), 5-carbamoylmethyl-2'-O-methyluridine (ncm(5)Um), 5-methoxycarbonylmethyl-uridine (mcm(5)U) or 5-methoxycarbonylmethyl-2-thiouridine (mcm(5)s(2)U). The formation of mcm(5) and ncm(5) side chains involves a complex pathway, where the last step in formation of mcm(5) is a methyl esterification of cm(5) dependent on the Trm9 and Trm112 proteins.

Methodology and principal findings: Both Trm9 and Trm112 are required for the last step in formation of mcm(5) side chains at wobble uridines. By co-expressing a histidine-tagged Trm9p together with a native Trm112p in E. coli, these two proteins purified as a complex. The presence of Trm112p dramatically improves the methyltransferase activity of Trm9p in vitro. Single tRNA species that normally contain mcm(5)U or mcm(5)s(2)U nucleosides were isolated from trm9Δ or trm112Δ mutants and the presence of modified nucleosides was analyzed by HPLC. In both mutants, mcm(5)U and mcm(5)s(2)U nucleosides are absent in tRNAs and the major intermediates accumulating were ncm(5)U and ncm(5)s(2)U, not the expected cm(5)U and cm(5)s(2)U.

Conclusions: Trm9p and Trm112p function together at the final step in formation of mcm(5)U in tRNA by using the intermediate cm(5)U as a substrate. In tRNA isolated from trm9Δ and trm112Δ strains, ncm(5)U and ncm(5)s(2)U nucleosides accumulate, questioning the order of nucleoside intermediate formation of the mcm(5) side chain. We propose two alternative explanations for this observation. One is that the intermediate cm(5)U is generated from ncm(5)U by a yet unknown mechanism and the other is that cm(5)U is formed before ncm(5)U and mcm(5)U.

Reference Type
Journal Article | Research Support, N.I.H., Extramural | Research Support, Non-U.S. Gov't
Authors
Chen C, Huang B, Anderson JT, Byström AS
Primary Lit For
Additional Lit For
Review For

Gene Ontology Annotations


Increase the total number of rows showing on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Gene/Complex Qualifier Gene Ontology Term Aspect Annotation Extension Evidence Method Source Assigned On Reference

Phenotype Annotations


Increase the total number of rows showing on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details.

Gene Phenotype Experiment Type Mutant Information Strain Background Chemical Details Reference

Disease Annotations


Increase the total number of rows showing on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Gene Disease Ontology Term Qualifier Evidence Method Source Assigned On Reference

Regulation Annotations


Increase the total number of rows displayed on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; to filter the table by a specific experiment type, type a keyword into the Filter box (for example, “microarray”); download this table as a .txt file using the Download button or click Analyze to further view and analyze the list of target genes using GO Term Finder, GO Slim Mapper, or SPELL.

Regulator Target Direction Regulation Of Happens During Method Evidence

Post-translational Modifications


Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through its pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Site Modification Modifier Reference

Interaction Annotations


Genetic Interactions

Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details about experiment type and any other genes involved in the interaction.

Interactor Interactor Allele Assay Annotation Action Phenotype SGA score P-value Source Reference

Physical Interactions

Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details about experiment type and any other genes involved in the interaction.

Interactor Interactor Assay Annotation Action Modification Source Reference

Functional Complementation Annotations


Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through its pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Gene Species Gene ID Strain background Direction Details Source Reference