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Transcript profiling demonstrates absence of dosage compensation in Arabidopsis following loss of a single RPL23a paralog

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Abstract

Translation of nucleus-encoded messages in plants is conducted by the cytoplasmic ribosome, an enzyme that is comprised of two RNA/protein subunits. In Arabidopsis thaliana, the 81 different ribosomal proteins (r-proteins) of the cytosolic ribosome belong to gene families with multiple expressed members. Given that ribosomes generally contain only one copy of each r-protein, regulatory mechanisms must exist to ensure their stoichiometric accumulation. These mechanisms must be dynamic, allowing for adjustments to ribosome biogenesis to fulfill biological requirements for protein synthesis during development, and following stress induction of global changes in gene expression. In this study, we investigated whether r-protein paralogs are feedback regulated at the transcript level by obtaining a T-DNA knockout of one member, RPL23aB, from the two-member RPL23a family. Expression of the lone functional paralog in this line, RPL23aA, was compared to the expression of both paralogs in wildtype plants under non-stressed, low temperature-, and high light stresses. RPL23aA expression was not upregulated in RPL23aB knockouts to compensate for paralog-loss, and consequently knockouts showed reduced total abundance of RPL23a transcripts. However, no phenotype developed in RPL23aB knockouts, suggesting that this paralog is dispensable under experimental conditions examined, or that compensation by RPL23aA may occur post-transcriptionally. Patterns of RPL23aA and RPL23aB transcript accumulation in wildtype plants suggest that paralogs respond coordinately to developmental and stress stimuli.

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Abbreviations

r-protein:

Ribosomal protein

rRNA:

Ribosomal RNA

SSU:

Small subunit

LSU:

Large subunit

RNP:

Ribonucleoprotein

mRNA:

Messenger RNA

BKO:

RPL23aB knockout

WT:

Wildtype

PSII:

Photosystem II

PCNA:

Proliferating cell nuclear antigen

CBF:

CRT/DRE binding factor

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Acknowledgments

We thank Jacqueline Hulm, Donna Lindsay and Heather Wakely for technical assistance. This work was supported by the Natural Sciences and Engineering Research Council of Canada.

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  1. Department of Biology, University of Saskatchewan, Saskatoon, SK, Canada, S7N 5E2

    Rory F. Degenhardt & Peta C. Bonham-Smith

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  1. Rory F. Degenhardt
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Correspondence to Rory F. Degenhardt.

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MOESM1 Table S1 (DOCX 12 kb)

425_2008_765_MOESM2_ESM.tif

MOESM2 Fig. S1 supplementary RT-PCR analysis of homozygous T-DNA insertion lines. Gene specific primers were used to amplify RPL23aA or RPL23aB (Supplemental Table 1; RPL23A1F and RPL23A1R for RPL23aA, RPL23A2F and RPL23A2R for RPL23aB) from total RNA extracts of wildtype plants (WT), putative RPL23aA T-DNA knockout lines SAIL-258-C12 (A1) and SALK-091329.46.50 (A2), and putative RPL23aB T-DNA knockout line SAIL-597-B08 (BKO). For the SAIL-258-C12 image, intervening lanes have been removed. Thirty cycles were used for PCR and amplicons were 437 and 535 bp for RPL23aA and RPL23aB, respectively. Agarose gels were stained with EtBr. (TIFF 845 kb)

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Degenhardt, R.F., Bonham-Smith, P.C. Transcript profiling demonstrates absence of dosage compensation in Arabidopsis following loss of a single RPL23a paralog. Planta 228, 627–640 (2008). https://doi.org/10.1007/s00425-008-0765-6

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