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

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.

Characterization of EMU, the Arabidopsis homolog of the yeast THO complex member HPR1

“…(Hugouvieux et al 2001;Bezerra et al 2004;Papp et al 2004). These observations raise the possibility that EMU is involved in connecting SE and CBC, which are involved in miRNA biogenesis but do not appear to interact directly with each other (Gregory et al 2008;Kim et al 2008;Laubinger et al 2008). Although we were unable to detect the two hybrid interactions among ABH1, EMU, SE, and AtTHO2 observed in yeast (C. Furumizu and Y. Komeda, unpubl.…”

“…Article and publication date are at http://www.rnajournal.org/cgi/doi/10.1261/rna.2265710. miRNA biogenesis pathways has been recently indicated in Arabidopsis, and this has been extended to RNA silencing in animals (Gregory et al 2008;Kim et al 2008;Laubinger et al 2008;Gruber et al 2009;Sabin et al 2009;Büssing et al 2010). Connecting these two processes permits more robust and accurate regulation of gene expression, and further inquiry into the crosstalk is expected to reveal additional roles.…”

The role of decapping proteins in the miRNA accumulation inArabidopsis thaliana

“…Previous research showed that cap-binding proteins are involved in the processing of primiRNAs, so some pri-miRNAs increased in these genes mutants. 38,39 Similarly, the low level of miRNA accumulation in the decapping mutants may be due to a defect of the processing of pri-miRNAs. To examine whether DCP1, DCP2 and VCS are involved in primiRNAs processing, we examined the levels of class I and class II pri-miRNAs in the decapping mutants by qRT-PCR.…”

The crosstalk between plant microRNA biogenesis factors and the spliceosome

“…21 It has also been shown that the lack of either of these proteins impairs the biogenesis of certain miRNAs. 22,23 A very specific feature of pri-miRNAs is the formation of a stem-loop structure in which miRNA and complementary miRNA* are embedded. DCL1 interacts with SERRATE (SE) and HYPONASTIC LEAVES 1 (HYL1), forming the plant microprocessing complex.…”

“…[24][25][26] SE cooperates with CBC to ensure the proper processing of pri-miRNAs. [22][23][24] Interestingly, the accuracy of primiRNA processing requires HYL1 dephosphorylation that is triggered by a C-TERMINAL DOMAIN PHOSPHATASE-LIKE 1 (CPL1). 27 Recently it has been shown that the efficiency of primiRNA recruitment to DCL1-HYL1-SE complex is also improved by a RNA binding protein TOUGH (TGH).…”