Deep sequencing of the tobacco mitochondrial transcriptome reveals expressed ORFs and numerous editing sites outside coding regions

Benjamin T Grimes,Awa K Sisay,Hyrum D Carroll,A Bruce Cahoon

doi:10.1186/1471-2164-15-31

Benjamin T Grimes, Awa K Sisay + Show 2 more

Open Access

https://doi.org/10.1186/1471-2164-15-31

Copy DOI

Journal: BMC Genomics	Publication Date: Jan 17, 2014
Citations: 103	License type: cc-by

Affiliation: Middle Tennessee State University

Abstract

BackgroundThe purpose of this study was to sequence and assemble the tobacco mitochondrial transcriptome and obtain a genomic-level view of steady-state RNA abundance. Plant mitochondrial genomes have a small number of protein coding genes with large and variably sized intergenic spaces. In the tobacco mitogenome these intergenic spaces contain numerous open reading frames (ORFs) with no clear function.ResultsThe assembled transcriptome revealed distinct monocistronic and polycistronic transcripts along with large intergenic spaces with little to no detectable RNA. Eighteen of the 117 ORFs were found to have steady-state RNA amounts above background in both deep-sequencing and qRT-PCR experiments and ten of those were found to be polysome associated. In addition, the assembled transcriptome enabled a full mitogenome screen of RNA C→U editing sites. Six hundred and thirty five potential edits were found with 557 occurring within protein-coding genes, five in tRNA genes, and 73 in non-coding regions. These sites were found in every protein-coding transcript in the tobacco mitogenome.ConclusionThese results suggest that a small number of the ORFs within the tobacco mitogenome may produce functional proteins and that RNA editing occurs in coding and non-coding regions of mitochondrial transcripts.

Highlights

The purpose of this study was to sequence and assemble the tobacco mitochondrial transcriptome and obtain a genomic-level view of steady-state RNA abundance
Deep sequencing and alignment of the tobacco mitochondrial transcriptome Total RNA from tobacco leaves collected from six plants was sequenced in a single Illumina run and aligned to the tobacco mitochondrial genome as deposited in GenBank (NC_006581.1 and [6]), including repeated regions. 4,539,709 reads with an average length of 100nt aligned to the mitogenome and the resulting depth of coverage (DOC) chart revealed discrete regions with moderate to high DOC separated by spans of very low to non-existent coverage (Figures 1A & B, Additional file 1: Figure S1)
It has been suggested that mitochondrial genes between 3,000 and 8,000 nucleotides apart will most likely be transcribed as a cluster [21] which are processed and edited to form translatable mRNAs

Summary

Introduction

The purpose of this study was to sequence and assemble the tobacco mitochondrial transcriptome and obtain a genomic-level view of steady-state RNA abundance. Plant mitochondrial genomes have a small number of protein coding genes with large and variably sized intergenic spaces. Angiosperm mitochondrial genomes range from 200,000 to more than 2.6 million bp These large size differences are due to highly variable intergenic regions that lie between a relatively conserved set of protein coding genes [1,2]. Short degenerate repeats are common between genes in cucurbit mtDNA [4]. Richardson and Palmer [5] showed that the mitochondria of the dicot Amborella trichopoda contained sequences homologous to different species’ mitogenomes.

Objectives

Methods

Results

Discussion

Conclusion