Functional Roles Of Modifications Research Articles

Impact of methylations of m2G966/m5C967 in 16S rRNA on bacterial fitness and translation initiation

The functional centers of the ribosome in all organisms contain ribosomal RNA (rRNA) modifications, which are introduced by specialized enzymes and come at an energy cost for the cell. Surprisingly, none of the modifications tested so far was essential for growth and hence the functional role of modifications is largely unknown. Here, we show that the methyl groups of nucleosides m2G966 and m5C967 of 16S rRNA in Escherichia coli are important for bacterial fitness. In vitro analysis of all phases of translation suggests that the m2G966/m5C967 modifications are dispensable for elongation, termination and ribosome recycling. Rather, the modifications modulate the early stages of initiation by stabilizing the binding of fMet-tRNAfMet to the 30S pre-initiation complex prior to start-codon recognition. We propose that the m2G966 and m5C967 modifications help shaping the bacterial proteome, most likely by fine-tuning the rates that determine the fate of a given messenger RNA (mRNA) at early checkpoints of mRNA selection.

E-cadherin gene mutations in signet ring cell carcinoma of the stomach.

To clarify the significance of E‐cadherin gene alterations in the development of diffuse‐type adenocarcinoma of the stomach, we analyzed mutations of the E‐cadherin gene using the polymerase chain reaction single‐strand conformation polymorphism (PCR‐SSCP) method followed by direct sequencing. Twenty‐two signet ring cell carcinomas of the stomach (10 intramucosal and 12 advanced cancers) were examined. Genomic DNA was extracted separately from cancerous and non‐cancerous tissues and PCR‐SSCP analysis was performed on exons 5 to 9 and the adjacent 30‐ to 40‐base‐pair intron sequences of the E‐cadherin gene. Mobility shifts were found in 2 of the 10 intramucosal cancers. In 2 of the 12 advanced cancers, abnormalities of the E‐cadherin gene were observed in intramucosal lesions as well as in deeply invaded areas. These results indicated that E‐cadherin gene mutations are an early event in the development of signet ring cell carcinoma of the stomach. Direct sequencing revealed that the locations of mutations of the E‐cadherin gene included the branch point sequence in the intron which is responsible for RNA splicing. Reverse transcriptase‐PCR demonstrated aberrant RNA splicing in a case with a branch point mutation, suggesting that branch point mutations play an important role in functional modifications of E‐cadherin in signet ring cell carcinoma of the stomach.