Mutation of the gene encoding the ribonuclease P RNA in the hyperthermophilic archaeon Thermococcus kodakarensis causes decreased growth rate and impaired processing of tRNA precursors
| Autor: | Sonoko Ishino, Makoto Kimura, Yoshizumi Ishino, Toshifumi Ueda, Kotaro Suematsu, Yoshimitsu Kakuta, Takashi Nakashima, Yutaka Kawarabayasi |
|---|---|
| Rok vydání: | 2015 |
| Předmět: |
biology
RNase P Mutant Biophysics RNA Cell Biology biology.organism_classification Biochemistry Molecular biology RNase PH Ribonuclease P Thermococcus kodakarensis Enzyme Activation Thermococcus RNase MRP Structure-Activity Relationship RNA Transfer Transfer RNA Mutation Mutagenesis Site-Directed Molecular Biology Gene Cell Proliferation |
| Zdroj: | Biochemical and biophysical research communications. 468(4) |
| ISSN: | 1090-2104 |
| Popis: | Ribonuclease P (RNase P) catalyzes the processing of 5' leader sequences of tRNA precursors in all three phylogenetic domains. RNase P also plays an essential role in non-tRNA biogenesis in bacterial and eukaryotic cells. For archaeal RNase Ps, additional functions, however, remain poorly understood. To gain insight into the biological function of archaeal RNase Ps in vivo, we prepared archaeal mutants KUWΔP3, KUWΔP8, and KUWΔP16, in which the gene segments encoding stem-loops containing helices, respectively, P3, P8 and P16 in RNase P RNA (TkopRNA) of the hyperthermophilic archaeon Thermococcus kodakarensis were deleted. Phenotypic analysis showed that KUWΔP3 and KUWΔP16 grew slowly compared with wild-type T. kodakarensis KUW1, while KUWΔP8 displayed no difference from T. kodakarensis KUW1. RNase P isolated using an affinity-tag from KUWΔP3 had reduced pre-tRNA cleavage activity compared with that from T. kodakarensis KUW1. Moreover, quantitative RT-PCR (qRT-PCR) and Northern blots analyses of KUWΔP3 showed greater accumulation of unprocessed transcripts for pre-tRNAs than that of T. kodakarensis KUW1. The current study represents the first attempt to prepare mutant T. kodakarensis with impaired RNase P for functional investigation. Comparative whole-transcriptome analysis of T. kodakarensis KUW1 and KUWΔP3 should allow for the comprehensive identification of RNA substrates for archaeal RNase Ps. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect