The Footprints of Winter

Epigenetic marks laid down during the cold months of the year allow flowering in spring and summer.

By | March 1, 2011


Many plants that grow in climates with a cold winter require growth for several months at low temperatures—a process called vernalization—to promote flowering in spring, when days lengthen and temperatures increase. Without this period of cold, plants would grow leaves in the spring, but would fail to flower. This phenomenon, familiar to every horticulturist, was difficult to explain with genetics alone; something occurred during those cold months that left a mark, which, in effect, released a switch that permitted flowering in spring. In recent years, the field has looked beyond the genome and found that vernalization is controlled by a wide range of epigenetic mechanisms.

Researchers studying the genetics of flowering found that the flowering switch was controlled by two central players—the genes FLOWERING LOCUS T (FT) and SUPPRESSOR OF CONSTANS1 (SOC1), which were held in the “off” position by the product of the gene called FLOWERING LOCUS C (FLC).[1. I. Searle et al., “The transcription factor FLC confers a flowering response to vernalization by repressing meristem competence and systemic signaling in Arabidopsis,” Genes Dev, 20:898-912, 2006. Free F1000 Evaluation] Through a process that is initiated in the winter, and is complete by spring, FLC expression is reduced to levels that release the switch, allowing expression of FT and SOC1 to drive flowering. Whatever molecular mechanism reduces FLC expression, it is clearly sensitive to environmental input: cold temperature.

When Caroline Dean and colleagues at the John Innes Centre in the United Kingdom experimentally exposed Arabidopsis plants to cold temperatures, they saw expression of FLC progressively reduced. In addition, the production of PHD-PRC2, which belongs to a family of proteins called the Polycomb Group (PcG), was upregulated and recruited to FLC. PcG proteins, which are expressed by animals and plants alike, mark histone tails with a number of different epigenetic modifications including methylation and ubiquitination, altering whether the DNA in chromatin is tightly bound or available for transcription. (See Epigenetics: A Primer.) When Dean and colleagues purified the PHD-PRC2 complex, they saw that it contained the plant equivalents of the four core components of a PcG protein complex identified in animals called Polycomb repressive complex 2 (PRC2). PRC2 acts as a histone methyltransferase, specifically catalyzing trimethylation of lysine 27 on the histone H3 tail (H3K27me3), which results in repression of the genes wound around these histones. In addition, PHD-PRC2 contains several plant-specific components which probably boost the histone methyltransferase activity. The group observed that the methylation marks on histone 3 of the FLC gene increased during cold treatments and persisted when plants were moved to warm conditions, in effect silencing the FLC gene.[2. F. De Lucia et al., “A PHD-polycomb repressive complex 2 triggers the epigenetic silencing of FLC during vernalization,” PNAS, 105:16831-36, 2008.]

Because the PcG components are mostly expressed constitutively, and seemingly lack specific DNA binding activity, it has been a major problem to explain how they are recruited to specific targets such as FLC. Numerous recent studies in animals have shown that noncoding RNA (ncRNA) plays a role in recruitment,[3. R. Margueron and D. Reinberg, “The Polycomb complex PRC2 and its mark in life,” Nature, 469, 343-49, 2011.] and two noncoding RNAs have now been implicated in vernalization-induced recruitment of PHD-PRC2. One of these, termed COLDAIR, is a long (about 1 kb), noncoding sense RNA produced from sequences within FLC—the very gene it appears to silence.[4. J.B. Heo and S. Sung, “Vernalization-mediated epigenetic silencing by a long intronic noncoding RNA,” Science, 331:76-79, 2011. Free F1000 Evaluation] Expression of COLDAIR is cold-induced and several observations suggest it indeed silences FLC. First, when the sequence within FLC that coded for the ncRNA was deleted, FLC did not stay switched off when plants were moved to a warm environment. In a sense they had become forgetful by losing their winter marks, showing that this ncRNA was required for silencing.4,[5. C.C. Sheldon et al., “Different regulatory regions are required for the vernalization-induced repression of FLOWERING LOCUS C and for the epigenetic maintenance of repression,” Plant Cell, 14:2527-37, 2002.] Secondly, researchers have shown that a component of the PHD-PRC2 complex binds the COLDAIR ncRNA and recruits the PHD-PRC2 to FLC.

A second noncoding RNA, termed COOLAIR, is also produced from FLC.[6. S. Swiezewski et al., “Cold-induced silencing by long antisense transcripts of an Arabidopsis Polycomb target,” Nature, 462:799-802, 2009. Free F1000 Evaluation] Like COLDAIR, COOLAIR expression is also strongly upregulated by cold. There is good evidence that COOLAIR also plays a role in regulating FLC expression. Two genes, FPA and FCA, promote flowering by downregulating FLC expression. Gordon Simpson and colleagues from Dundee University in Scotland showed that these genes function by regulating the processing of COOLAIR transcripts.[7. C. Hornyik et al., “The spen family protein FPA controls alternative cleavage and polyadenylation of RNA,” Dev Cell, 18:203-13, 2010. Free F1000 Evaluation] However, FPA and FCA are not required for the vernalization response, and the function of COOLAIR in vernalization is less clear. One possibility, suggested by Jae Bok Heo and Sibum Sung at the University of Texas at Austin, is that COOLAIR may reduce transcription from the normal promoter at the 5’ end of the FLC protein coding region and promote COLDAIR expression from its cryptic promoter within FLC.4

Although many plants show a vernalization response with epigenetic features, the target genes and even the mechanisms may be different than in Arabidopsis. In sugar beets, vernalization results in stable repression of a floral repressor which, although it is unrelated to FLC, acts in a similar fashion to repress an FT homologue. It is not clear yet if this is also PcG mediated. In cereals, vernalization causes the stable upregulation of a gene (VRN1) which promotes flowering. Here, the epigenetic memory is of an active state, and is correlated with H3K4me3 methylation at VRN1. It is likely, although not yet proven, that this will involve the trx-G genes as these are known to catalyze H3K4me3 methylation and work antagonistically to the PcG to mediate stable gene activation.

Ralf Müller and F1000 Member Justin Goodrich are at the Institute of Molecular Plant Sciences, University of Edinburgh.

This article is adapted from an upcoming review in F1000 Medicine Reports. It will be available for citation at access)


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