IntegratingELF4into the circadian system through combined structural and functional studies

Abstract

The circadian clock is a timekeeping mechanism that enables anticipation of daily environmental changes. In the plant Arabidopsis thaliana, the circadian system is a multiloop series of interlocked transcription-translation feedbacks. Several genes have been arranged in these oscillation loops, but the position of the core-clock gene ELF4 in this network was previously undetermined. ELF4 lacks sequence similarity to known domains, and functional homologs have not yet been identified. Here we show that ELF4 is functionally conserved within a subclade of related sequences, and forms an alpha-helical homodimer with a likely electrostatic interface that could be structurally modeled. We support this hypothesis by expression analysis of new elf4 hypomorphic alleles. These weak mutants were found to have expression level phenotypes of both morning and evening clock genes, implicating multiple entry points of ELF4 within the multiloop network. This could be mathematically modeled. Furthermore, morning-expression defects were particular to some elf4 alleles, suggesting predominant ELF4 action just preceding dawn. We provide a new hypothesis about ELF4 in the oscillator-it acts as a homodimer to integrate two arms of the circadian clock.

EARLY FLOWERING3 Redundancy Fine-Tunes Photoperiod Sensitivity

“…In legumes, however, NOX genes are not present, and a null mutant for the single LUX ortholog in pea, SN, has a very strong early flowering phenotype and is completely insensitive to photoperiod (Liew et al, 2014), similar to the double ELF3 mutant hr ppd. Arabidopsis ELF4 is a member of a small gene family (Khanna et al, 2003) in which the closest paralog, ELF4-LIKE1 (EFL1), appears also to have ELF4 activity but three other ELF4-like genes do not (Kolmos et al, 2009). In legumes, ELF4 is also represented by small gene family (Liew et al, 2009), and the fact that in pea an apparent null mutant for the ELF4 ortholog DNE has only a minor effect on flowering when the other EC genes are intact (Liew et al, 2014) again implies a probable redundancy with one or more other members of the family.…”

Abiotic stress and the plant circadian clock

“…Research efforts in this direction have placed EARLY FLOWERING 4 (ELF4) within the oscillator as a key circadian repressor. 17,18,24,25 Other studies have shown that post-translational modifications, including protein targeted degradation and protein phosphorylation are essential for the function of the plant circadian clock. As an example, it has been shown that ZEITLUPE (ZTL), a light-sensitive protein, binds to TOC1 during the night, directing TOC1 to degradation by the proteasome.…”

“…16 Though these three elements have been considered the core of the plant clock, this model cannot fully explain all experimental data. [17][18][19][20][21] Mathematical models have been formalized to comprise a clock with three interconnected feedback loops, which harbors unknown components within them. In a three-loop model, [20][21][22] besides their described role in the core oscillator, CCA1 and LHY also participate in a morning loop to induce the expression of the PSEUDO RESPONSE REGULATOR 7 and 9 (PRR7 and PPR9), whose encoded proteins in turn bind and repress the transcription of the formers.…”

Quantitative Circadian Phosphoproteomic Analysis of Arabidopsis Reveals Extensive Clock Control of Key Components in Physiological, Metabolic, and Signaling Pathways

“…These results show that phospho-oscillation at S45 is independent of protein levels and light/dark cycles, and suggest that ELF4 phosphorylation is clock-regulated, possibly through clock-gated kinase or phosphatase activity. To understand the significance of phosphorylation of S45 we took advantage of the elf4 -211 allele, which bears an S45L transition (129). elf4 null mutants are very poorly rhythmic, flower early in short days and have long hypocotyls (130 -132).…”