Roles of Arabidopsis Cyclin-Dependent Kinase C Complexes in Cauliflower Mosaic Virus Infection, Plant Growth, and Development

Abstract

The C-terminal domain (CTD) of RNA polymerase II is phosphorylated during the transcription cycle by three cyclindependent kinases (CDKs): CDK7, CDK8, and CDK9. CDK9 and its interacting cyclin T partners belong to the positive transcription elongation factor b (P-TEFb) complexes, which phosphorylate the CTD to promote transcription elongation. We report that Arabidopsis thaliana CDK9-like proteins, CDKC;1 and CDKC;2, and their interacting cyclin T partners, CYCT1;4 and CYCT1;5, play important roles in infection with Cauliflower mosaic virus (CaMV). cdkc;2 and cyct1;5 knockout mutants are highly resistant and cdkc;2 cyct1;5 double mutants are extremely resistant to CaMV. The mutants respond normally to other types of plant viruses that do not replicate by reverse transcription. Expression of a reporter gene driven by the CaMV 35S promoter is markedly reduced in the cdkc;2 and cyct1;5 mutants, indicating that the kinase complexes are important for transcription from the viral promoter. Loss of function of CDKC;1/CDKC;2 or CYCT1;4/CYCT1;5 results in complete resistance to CaMV as well as altered leaf and flower growth, trichome development, and delayed flowering. These results establish Arabidopsis CDKC kinase complexes as important host targets of CaMV for transcriptional activation of viral genes and critical regulators of plant growth and development. INTRODUCTIONThe transcription of protein-coding genes in eukaryotes is performed by RNA polymerase II (RNAP II) and forms a so-called transcription cycle that includes preinitiation, initiation, promoter clearance, elongation, and termination (Sims et al., 2004). The transcription cycle starts with the assembly of the preinitiation complex at the promoter, which includes the general transcription factors TFIID, TFIIB, TFIIE, and TFIIH as well as RNAP II. Assembly of the preinitiation complex is followed by ATPdependent melting of the double-stranded DNA (dsDNA) template and formation of an open complex between RNAP II and the DNA template. Once the open complex is established, transcription initiation occurs and RNAP II is allowed to clear the promoter and engaged to make the transition from initiation to elongation. Transcription elongation is a dynamic and highly regulated process that coordinates downstream events such as capping and splicing of primary transcripts. The final step in the cycle is transcription termination. At this stage, the mRNA is cleaved, polyadenylated, and transported to the cytoplasm for translation.The transcription cycle involving RNAP II is accompanied by another cycling event: phosphorylation of the C-terminal domain (CTD) of the largest subunit of RNAP II (Sims et al., 2004). The RNAP II CTD contains multiple repeats of the heptapeptide sequence YSPTSPS (26 such repeats in yeast, 32 in Caenorhabditis elegans, 45 in Drosophila, and 52 in mammals). RNAP II in the preinitiation complex is unphosphorylated or hypophosphorylated, whereas transcription-competent RNAP II is heavily phosphorylated on its CTD, with Ser-2 and Ser-5 as the major...

CDKF;1 and CDKD Protein Kinases Regulate Phosphorylation of Serine Residues in the C-Terminal Domain of Arabidopsis RNA Polymerase II

“…In contrast with a previous report (Shimotohno et al, 2004), our data demonstrate that CDKD;1, a third member of the CDKD family, is also an active kinase and, similarly to CDKD;2 and CDKD;3, phosphorylates all Ser residues of Arabidopsis RNAPII CTD. In the cdkd double and d123* triple mutants, reduction of the CTD S 2 P mark during seedling development showed slower kinetics compared with the depletion of S 5 P, suggesting a possible involvement of other S 2 -specific CTD kinases, including candidate CDKC homologs of Ctk1/CDK9 (Fülöp et al, 2005;Cui et al, 2007). Throughout seedling development, the S 2 P levels were nonetheless lower in cdkf;1 compared with the cdkd mutants, suggesting that CDKF;1 is probably also required for maintenance of the CDKDindependent CTD S 2 -kinase activity.…”

Sequencing of the genus Arabidopsis identifies a complex history of nonbifurcating speciation and abundant trans-specific polymorphism

“…8) and were significantly (FDR-corrected P = 0.006; Online Methods) enriched for genes involved in response to virus, suggesting that selection may indeed have contributed to maintaining some of the ancestral polymorphisms. We identified a total of four virus-response genes with haplotypes shared across all four species: CYCT1;4 (TAIR locus AT4G19600), which is involved in transcriptional activation of viral genes; GRIK1 (TAIR locus AT3G45240), which has a role in the metabolic regulation of virus-infected cells 26,27 ; NIG (TAIR locus AT4G13350), which is involved in nucleocytoplasmic trafficking of viral proteins 28 ; and WAK1 (TAIR locus AT1G21250), which activates an immunity response to damaged cell walls 29 .…”

A critical role of autophagy in plant resistance to necrotrophic fungal pathogens

“…The resulting clones were verified by sequencing. The plasmids were introduced into Agrobacterium tumefaciens (strain GV3101), and infiltration of N. benthamiana was performed as described previously (Cui et al, 2007). Infected tissues were analyzed at approximately 24 h after infiltration.…”

“…DNA sequences for the N-terminal 173 amino acid YFP (N-YFP) and the C-terminal 64 amino acid YFP (C-YFP) fragments were PCRamplified and cloned into plasmid pFGC5941 to generate plasmids pFGC-N-YFP and pFGC-C-YFP, respectively (Cui et al, 2007). The WRKY33 and WRKY48 coding sequences were inserted into pFGC-N-YFP to generate N-terminal in-frame fusions with N-YFP, and ATG18a was introduced into pFGC-C-YFP to generate C-terminal in-frame fusions with C-YFP.…”