Potential regulators of CpATFL1 initiate the floral
transition in C. pallens
Global expression analysis of the leaves collected during the summer
inductive conditions from tillers that subsequently flowered shows
upregulation of crucial genes including CpFTIP1 and CpFDPthat are involved in the transport of florigen-like molecules from the
leaves to the apex to subsequently activate the floral meristem gene(s)
through formation of the florigen activation complex (FAC)
(Kaneko-Suzuki et al., 2018). Calcium is also required to catalyse the
formation of the florigen activation complex (Kawamoto, Sasabe, Endo,
Machida, & Araki, 2015).The expression of CpCPK6 , a kinase
required for calcium signalling, was also upregulated in the tillers
that flowered in the next season. It is important to acknowledge that
formation of FAC occurs at the shoot apical meristem. Even though the
differential expression of these genes was observed in the leaves, it
may suggest that CpATFL1 may form the FAC through the activity of
CpFTIP1 and CpFDP, catalysed by CpCPK6 at the shoot apical meristem.
Further investigation involving a yeast two hybrid assay or bimolecular
fluorescence assay would reveal whether CpATFL1 and CpFTIP1 do indeed
interact and could, therefore, transport ATFL1 to the apex.
Vernalisation responses to mediate flowering-time control in temperate
monocots are controlled by the VERNALISATION (VRN) loci (Reams et
al., 2014). The VRN2 locus encodes a CCT-domain protein
that blocks the floral transition. Exposure to cold temperatures
increases the expression of VRN1 , a repressor of VRN2 (Yan
et al., 2004) and maintains the repressive state of VRN2even after vernalisation. Repression of VRN2 leads to activation
of VRN3 , a homologue of FT and Hd3a, after plants
are exposed to warm spring temperatures (Preston & Kellogg, 2008;
Shimada et al., 2009; Woods, et al., 2016). Transcriptomic analysis
showed an elevated expression of CpVRN1 in the tillers that
flowered during the increase in summer temperature, which is also
observed in other temperate cereals during the process of floral
transition (Trevaskis, 2010). On the other hand, expression ofCpVRN2 , the repressor of FT -like genes (Alexandre &
Hennig, 2007), was downregulated in the same samples. Greater expression
of CpVRN1 during the spring season and which remained elevated
through the summer may have blocked repressive signals fromCpVRN2 . This repression was potentially maintained until the next
summer resulting in the activated transcription of CpATFL1 to
induce flowering.
Transcriptomic analysis also revealed an increase in the expression of
thermosensory genes, including CpPIF4 and CpPIF5 in the
tillers associated with flowering in the next season. PIF genes
are known to regulate responses to high temperatures (Choi & Oh, 2016)
and are involved in the activation of the flowering process along with
similar bHLH floral promoting proteins such as bHLH76 and bHLH80 (Ito et
al., 2012). In addition to PIF -family genes, homologues ofbHLH76 and bHLH80 in C. pallens , were also
upregulated in the above samples, which may interact with CpPIF4 to
activate ATFL1 . Transient assays are further required to confirm
this interaction. The summer temperature cue may also have blocked the
expression of several floral repressors including homologues ofAP2-LIKE genes and SVP (Capovilla, Schmid, & Pose, 2015;
Mateos et al., 2015).
Emphasis was also placed on the role of epigenetic modifiers known to
regulate the reproductive transition in plants. Ambient temperatures
have been shown to regulate the expression of floral repressors via
epigenetic modification including either through deposition of
repressive histone marks (to suppress gene expression) such as H3K27me3
or by removing acetyl groups from the histone tails at the gene loci
(Bratzel & Turck, 2015; He, 2012). In contrast, high temperatures have
been shown to lead to deposition of active histone marks (H3K4me3 or
H3K36me3) at the loci of floral promoters (Avramova, 2015).CpREF6 and CpFLD , homologues of REF6 and FLDwhich are reported to promote flowering (He, 2012; Lu, Cui, Zhang,
Jenuwein, & Cao, 2011), were upregulated in the leaves of plants that
flowered in the next season. CpFLD may then interact with CpHDA6, a
histone deacetylase complex, activating the floral promoting genes, a
process well established in Arabidopsis (Yu, Chang, & Wu, 2016).
Homologues of CpFLK , CpFVE , and CpFY , genes known
to repress the expression of floral repressors epigenetically (Cho,
Yoon, & An, 2017; He & Amasino, 2005), were upregulated in the tillers
that subsequently flowered. Interestingly, homologues of epigenetic
editors, including CpVEL1 and CpMSI1 , which are known to
be involved in the activation of VRN1 and SOC1 ,
respectively through deposition of active histone marks (Higgins,
Bailey, & Laurie, 2010; Oliver & Finnegan, 2011) were also upregulated
in the tillers that flowered in the next season. This may suggest that
an external signal such as summer temperatures may lead to certain
epigenetic changes enabling the transcription of CpATFL1 to
promote flowering.