Genome-wide Binding Site Analysis Of Far-red Elongated Hypocotyl 3 Reveals Its Novel Function In Regulating Chloroplast Division Of Arabidopsis

Light is one of the most important environmental cues that control plant growth and development. As sessile organisms, higher plants have evolved a network of multiple photoreceptors, including phytochromes, cryptochromes and phototropins, to sense changes in the ambient light environment in order to undergo adaptive growth and development. Phytochrome A (phyA) is a very important photoreceptor are required for senseing far-red light.FAR-RED ELONGATED HYPOCOTYL 3 (FHY3) and its homolog FAR-RED IMPAIRED RESPONSE 1 (FAR1) are two transposase-derived transcription factors initially identified as the key components in phytochrome A signaling. FAR-RED ELONGATED HYPOCOTYL 1 (FHY1) and its homolog FHY1-LIKE (FHL) two small plant-specific proteins, are required for nuclear accumulation of light-activated phyA since phyA is localized only in the cytosol of fhyl fhl double mutants. FHY3 and FAR1 are the key transcription factors directly activating the transcription of FHY1 and FHL, and thus indirectly control phyA nuclear accumulation and phyA responses.Recently, several reports have shed light on the functional diversities and regulatory modes of these transposase-derived transcription factors. Firstly, FHY3 was found to be involved in independently gating phytochrome signaling to the circadian clock. FHY3 and FAR1 are involved in maintaining the rhythmic expression of EARLY FLOWERING 4 (ELF4), a key player of the central oscillator of Arabidopsis circadian clock. FHY3 and FAR1 were shown to work together with CIRCADIAN CLOCK ASSOCIATED 1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY), two MYB-related transcription factors being as the key components of the central oscillator, to generate the rhythmic expression of ELF4. However, whether FHY3 and FAR1 are involved in other processes of plant development remains largely unknown. Here, we explored chromatin immunoprecipitation-based sequencing (ChIP-seq) analysis, mircoarray, yeast one hybrid and EMSA and got the results as following:1 We used ChIP-seq to identify 1559 and 1009 FHY3 direct binding target genes in darkness and far-red light conditions, respectively in the Arabidopsis thaliana genome. This analysis revealed that FHY3 preferentially binds to the gene promoters through the previously identified typical FHY3/FAR1 binding motif;2 Interestingly, FHY3 also binds to two novel motifs in the 178-bp repeats of the Arabidopsis centromere regions in vivo;3 Comparison between the ChIP-seq and microarray data indicates that FHY3 regulates the expression of 197 and 86 genes in dark and far-red respectively by directly binding to their promoters. FHY3 also co-regulates a number of common target genes with PHYTOCHROME INTERACTING FACTOR 3-LIKE 5 (PIL5) and ELONGATED HYPOCOTYL 5 (HY5)4 Our genome-wide identification of FHY3 direct target genes ultimately led to the discovery and validation of a new role of FHY3 in controlling chloroplast development, by directly activating the expression of ACCUMULATION AND REPLICATION OF CHLOROPLASTS5 (ARC5), a key gene regulating chloroplast constriction and division.Taken together, we used ChIP-seq to identify FHY3 target genes and used FHY3p:FHY3-GR/fhy3 transgenic materrals to find FHY3 directly regulated target genes. In addition, we found FHY3 can control the chloroplast division though target gene ARC5. So our data suggest that FHY3 is involved in regulating multiple facets of plant development, thus providing new insights into the functions of this type of transposase-derived transcription factors.