Polymerase II–Associated Factor 1 Complex-Regulated FLOWERING LOCUS C-Clade Genes Repress Flowering in Response to Chilling

Abstract

RNA polymerase II–associated factor 1 complex (PAF1C) regulates the transition from the vegetative to the reproductive phase primarily by modulating the expression of FLOWERING LOCUS C (FLC) and FLOWERING LOCUS M [FLM, also known as MADS AFFECTING FLOWERING1 (MAF1)] at standard growth temperatures. However, the role of PAF1C in the regulation of flowering time at chilling temperatures (i.e., cold temperatures that are above freezing) and whether PAF1C affects other FLC-clade genes (MAF2–MAF5) remains unknown. Here, we showed that Arabidopsis thaliana mutants of any of the six known genes that encode components of PAF1C [CELL DIVISION CYCLE73/PLANT HOMOLOGOUS TO PARAFIBROMIN, VERNALIZATION INDEPENDENCE2 (VIP2)/EARLY FLOWERING7 (ELF7), VIP3, VIP4, VIP5, and VIP6/ELF8] showed temperature-insensitive early flowering across a broad temperature range (10°C–27°C). Flowering of PAF1C-deficient mutants at 10°C was even earlier than that in flc, flm, and flc flm mutants, suggesting that PAF1C regulates additional factors. Indeed, RNA sequencing (RNA-Seq) of PAF1C-deficient mutants revealed downregulation of MAF2–MAF5 in addition to FLC and FLM at both 10 and 23°C. Consistent with the reduced expression of FLC and the FLC-clade members FLM/MAF1 and MAF2–MAF5, chromatin immunoprecipitation (ChIP)-quantitative PCR assays showed reduced levels of the permissive epigenetic modification H3K4me3/H3K36me3 and increased levels of the repressive modification H3K27me3 at their chromatin. Knocking down MAF2–MAF5 using artificial microRNAs (amiRNAs) in the flc flm background (35S::amiR-MAF2–5 flc flm) resulted in significantly earlier flowering than flc flm mutants and even earlier than short vegetative phase (svp) mutants at 10°C. Wild-type seedlings showed higher accumulation of FLC and FLC-clade gene transcripts at 10°C compared to 23°C. Our yeast two-hybrid assays and in vivo co-immunoprecipitation (Co-IP) analyses revealed that MAF2–MAF5 directly interact with the prominent floral repressor SVP. Late flowering caused by SVP overexpression was almost completely suppressed by the elf7 and vip4 mutations, suggesting that SVP-mediated floral repression required a functional PAF1C. Taken together, our results showed that PAF1C regulates the transcription of FLC and FLC-clade genes to modulate temperature-responsive flowering at a broad range of temperatures and that the interaction between SVP and these FLC-clade proteins is important for floral repression.

Distinct role of subunits of the Arabidopsis RNA polymerase II elongation factor PAF1C in transcriptional reprogramming

“…With elf7-3 plants, the presence of NaCl resulted in distinctly lower transcriptional response, indicating that ELF7 plays an important role in the response to elevated NaCl concentrations. In agreement with our findings, a recent study concluded that Col-0 and cdc73 Arabidopsis plants grown at 23°C have a similar transcriptome, and that elf7 and vip3/4/5/6 share a large number of DEGs ( Nasim et al., 2022 ). In yeast as well as in mammals, PAF1C subunit mutants are also associated with a range of phenotypes suggesting (partial) subunit specificity ( Betz et al., 2002 ; Akanuma et al., 2007 ; Chu et al., 2007 ; Cheung et al., 2008 ; Wang et al., 2008 ).…”

“…Elongator, SPT4-SPT5, TFIIS) exhibit differential expression of only subsets of genes ( Nelissen et al., 2005 ; Grasser et al., 2009 ; Dürr et al., 2014 ). Likewise, relative to wild type under normal growth conditions, based on microarray hybridization experiments ( Park et al., 2010 ) or RNA-seq of total mRNA ( Nasim et al., 2022 ), several hundreds of DEGs were observed for mutants deficient in PAF1C subunits, except for cdc73 , whose transcriptome was similar to that of wildtype. Consistently, using high-throughput sequencing of nuclear mRNAs, we identified 12 and 722 DEGs in cdc73-2 and elf7-3 , respectively.…”

“…Recently, PAF1C (i.e. VIP3, VIP5, VIP6) was also identified as a critical factor in the plant response to repeated mechanic stimulation ( Jensen et al., 2017 ) and in temperature-responsive flowering ( Nasim et al., 2022 ). We have comparatively exposed different PAF1C subunit mutants to elevated concentrations of NaCl to provoke transcriptional response.…”

“…H3K4me3, H3K27me3, H3K36me2) ( He et al., 2004 ; Oh et al., 2008 ; Xu et al., 2008 ; Park et al., 2010 ; Yu and Michaels, 2010 ). PAF1C also modulates the temperature-responsive transition to flowering ( Nasim et al., 2022 ). Beyond that PAF1C is required for plant response to repeated touch stimuli, as Arabidopsis mutants deficient in VIP3, VIP5 and VIP6 did not react to the mechanical stimulation as the wild type.…”

The SMC5/6 complex recruits the PAF1 complex to facilitate DNA double‐strand break repair in Arabidopsis

“…The roles of PAF1C in transcription regulation have been well‐studied (Moniaux et al , 2006; Jaehning, 2010; Antosz et al , 2017; Vos et al , 2018; Nasim et al , 2022). Here we found that PAF1C plays a critical role in DSB repair.…”