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Plant and Cell Physiology Advance Access originally published online on September 30, 2008
Plant and Cell Physiology 2008 49(11):1634-1644; doi:10.1093/pcp/pcn146
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© The Author 2008. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oxfordjournals.org

Rapid Paper

Two Cap-Binding Proteins CBP20 and CBP80 are Involved in Processing Primary MicroRNAs

Sanghee Kim1, Jun-Yi Yang1, Jun Xu1, In-Cheol Jang1, Michael J. Prigge2 and Nam-Hai Chua1,*

1 Laboratory of Plant Molecular Biology, The Rockefeller University, 1230 York Avenue, New York, NY 10065-6399, USA
2 Department of Biology, Indiana University, 915 East Third Street, Bloomington, IN 47405, USA

*Corresponding author: E-mail, chua{at}mail.rockefeller.edu; Fax, +1-212-327-8327.


  Abstract

MicroRNAs (miRNAs) are 21 nt RNAs that regulate many biological processes in plants by mediating translational inhibition or cleavage of target transcripts. Arabidopsis mutants defective in miRNA biogenesis have overlapping and highly pleiotropic phenotypes including serrated leaves and ABA hypersensitivity. Recent evidence indicates that miRNA genes are transcribed by RNA polymerase II (Pol II). Since Pol II transcripts are capped, we hypothesized that CBP (cap-binding protein) 20 and 80 may bind to capped primary miRNA (pri-miRNA) transcripts and play a role in their processing. Here, we show that cbp20 and cbp80 mutants have reduced miRNA levels and increased pri-miRNA levels. Co-immunoprecipitation experiments revealed that pri-miRNAs 159, 166, 168 and 172 could be associated with CBP20 and CBP80. We found that CBP20 and CBP80 are stabilized by ABA by a post-translational mechanism, and these proteins are needed for ABA induction of miR159 during seed germination. The lack of miR159 accumulation in ABA-treated seeds of cbp20/80 mutants leads to increased MYB33 and MYB101 transcript levels, and presumably higher levels of these positive regulators result in ABA hypersensitivity. Genetic and molecular analyses show that CBP20 and 80 have overlapping function in the same developmental pathway as SE and HYL1. Our results identify new components in miRNA biogenesis.

Keywords: ABA - Cap-binding protein - MicroRNA - Post-translational modification - Primary microRNA

Abbreviations: CaMV, cauliflower mosaic virus; CBC, cap-binding complex; CBP, cap-binding protein; CHX, cycloheximide; GUS, β-glucuronidase; miRNA, microRNA; MS, Murashige and Skoog; Pol II, RNA polymerase II; pri-miRNA, primary microRNA; PTGS, post-transcriptional gene silencing; qRT–PCR, quantitative reverse transcription–PCR; RNAi, RNA interference; siRNA, small interfering RNA; ta-siRNA, trans-acting small interfering RNA; WT, wild-type.

(Received July 11, 2008; Accepted September 22, 2008)
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