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Mitochondrial and plastid functions depend on coordinated expression of proteins encoded by genomic compartments that have radical differences in copy number of organellar and nuclear genomes. In polyploids, doubling of the nuclear genome may add challenges to maintaining balanced expression of proteins involved in cytonuclear interactions. Here, we use ribo-depleted RNA sequencing (RNA-seq) to analyze transcript abundance for nuclear and organellar genomes in leaf tissue from four different polyploid angiosperms and their close diploid relatives. We find that even though plastid genomes contain <1% of the number of genes in the nuclear genome, they generate the majority (69.9 to 82.3%) of messenger RNA (mRNA) transcripts in the cell. Mitochondrial genes are responsible for a much smaller percentage (1.3 to 3.7%) of the leaf mRNA pool but still produce much higher transcript abundances per gene compared to nuclear genome. Nuclear genes encoding proteins that functionally interact with mitochondrial or plastid gene products exhibit mRNA expression levels that are consistently more than 10-fold lower than their organellar counterparts, indicating an extreme cytonuclear imbalance at the RNA level despite the predominance of equimolar interactions at the protein level. Nevertheless, interacting nuclear and organellar genes show strongly correlated transcript abundances across functional categories, suggesting that the observed mRNA stoichiometric imbalance does not preclude coordination of cytonuclear expression. Finally, we show that nuclear genome doubling does not alter the cytonuclear expression ratios observed in diploid relatives in consistent or systematic ways, indicating that successful polyploid plants are able to compensate for cytonuclear perturbations associated with nuclear genome doubling.
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Comparison of transcriptional activity profiling by metabolic labeling or nuclear RNA sequencing
SUMMARY The application of high‐throughput sequencing to cellular transcriptome profiling (RNA‐seq) has enabled significant advances in our understanding of gene expression in plants. However, conventional RNA‐seq data reports mainly cytoplasmic transcript abundance rather than actual transcription rates. As a result, it is less sensitive to detect unstable and low‐abundance nuclear RNA species, such as long non‐coding RNAs, and is less directly connected to chromatin features and processes such as DNA replication. To bridge this gap, several protocols have been established to profile newly synthesized RNA in plants and other eukaryotes. These protocols can be technically challenging and present their own difficulties and limitations. Here we analyze newly synthesized nuclear RNA metabolically labeledin vivowith 5‐ethynyl uridine (EU‐nuclear RNA) in maize (Zea maysL.) root tips and compare it with the entire nuclear RNA population. We also compare both nuclear RNA preparations to conventional RNA‐seq analysis of cellular RNA. The transcript abundance profiles of protein‐coding genes in nuclear RNA and EU‐nuclear RNA were tightly correlated with each other (R2 = 0.767), but quite distinct from that of cellular RNA (R2 = 0.170 or 0.293). Nuclear and EU‐nuclear RNA reads are frequently mapped across entire genes, including introns, while cellular reads are predominantly mapped to mature transcripts. Both nuclear and EU‐nuclear RNA exhibited a greater ability to detect both protein‐coding and non‐coding expressed genes.
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- Award ID(s):
- 2025811
- PAR ID:
- 10645552
- Publisher / Repository:
- Wiley-Blackwell
- Date Published:
- Journal Name:
- The Plant Journal
- Volume:
- 123
- Issue:
- 3
- ISSN:
- 0960-7412
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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