Abstract
In plant mitochondria, transcription proceeds well beyond the region that will become mature 3' extremities of mRNAs, and the mechanisms of 3' maturation are largely unknown. Here, we show the involvement of two exoribonucleases, AtmtPNPase and AtmtRNaseII, in the 3' processing of atp9 mRNAs in Arabidopsis thaliana mitochondria. Down-regulation of AtmtPNPase results in the accumulation of pretranscripts of several times the size of mature atp9 mRNAs, indicating that 3' processing of these transcripts is performed mainly exonucleolytically by AtmtPNPase. This enzyme is however not sufficient to completely process atp9 mRNAs, because with down-regulation of another mitochondrial exoribonuclease, AtmtRNaseII, about half of atp9 transcripts exhibit short 3' nucleotide extensions compared with mature mRNAs. These short extensions can be efficiently removed by AtmtRNaseII in vitro. Taken together, these results show that 3' processing of atp9 mRNAs in Arabidopsis mitochondria is, at least, a two-step phenomenon. First, AtmtPNPase is involved in removing 3' extensions that may reach several kilobases. Second, AtmtRNaseII degrades short nucleotidic extensions to generate the mature 3'-ends.
Highlights
In plant mitochondria, transcription proceeds well beyond the region that will become mature 3 extremities of mRNAs, and the mechanisms of 3 maturation are largely unknown
A polynucleotide phosphorylase (PNPase)-like Protein Is Present in A. thaliana Mitochondria—In order to identify candidate genes involved in plant mitochondrial mRNA degradation and/or processing, we searched the nuclear genome of Arabidopsis for genes encoding putative proteins that would fulfill two criteria
Such a gene (At5g14580) encodes a putative protein harboring significant homologies with bacterial and chloroplastic PNPase. This gene is expressed in Arabidopsis as shown by the availability of four expressed sequence tags (ESTs) in databases (AU238738, AV528692, AU229971, BE526020), its level of expression is relatively low as the corresponding mRNAs could not be detected by Northern blot experiments. cDNAs were amplified by RTPCR and cloned (GenBankTM accession number: Y14685)
Summary
Transcription proceeds well beyond the region that will become mature 3 extremities of mRNAs, and the mechanisms of 3 maturation are largely unknown. AtmtPNPase is not sufficient to process all atp9 mRNAs as precursors harboring short nucleotidic extensions accumulate in a mutant affected in the expression of a second mitochondrial ribonuclease, AtmtRNaseII.
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