bHLH-PAS protein RITMO1 regulates diel biological rhythms in the marine diatom Phaeodactylum tricornutum

Abstract

Periodic light–dark cycles govern the timing of basic biological processes in organisms inhabiting land as well as the sea, where life evolved. Although prominent marine phytoplanktonic organisms such as diatoms show robust diel rhythms, the mechanisms regulating these processes are still obscure. By characterizing aPhaeodactylum tricornutumbHLH-PAS nuclear protein, hereby named RITMO1, we shed light on the regulation of the daily life of diatoms. Alteration of RITMO1 expression levels and timing by ectopic overexpression results in lines with deregulated diurnal gene expression profiles compared with the wild-type cells. Reduced gene expression oscillations are also observed in these lines in continuous darkness, showing that the regulation of rhythmicity by RITMO1 is not directly dependent on light inputs. We also describe strong diurnal rhythms of cellular fluorescence in wild-type cells, which persist in continuous light conditions, indicating the existence of an endogenous circadian clock in diatoms. The altered rhythmicity observed in RITMO1 overexpression lines in continuous light supports the involvement of this protein in circadian rhythm regulation. Phylogenetic analysis reveals a wide distribution of RITMO1-like proteins in the genomes of diatoms as well as in other marine algae, which may indicate a common function in these phototrophs. This study adds elements to our understanding of diatom biology and offers perspectives to elucidate timekeeping mechanisms in marine organisms belonging to a major, but under-investigated, branch of the tree of life.

The Aureochrome Photoreceptor PtAUREO1a Is a Highly Effective Blue Light Switch in Diatoms

“…Overall, the studied bZIP transcription factors like PtbZIP10 and PtbZIP18 revealed a rather high transcript abundance. PtbHLH1a (RITMO1), which recently has been shown to be involved in the circadian regulation of P. tricornutum ( Annunziata et al., 2019 ), is strongly transcribed after 10 min, and to a lesser extent after 60 min, which is mirrored much weaker in the mutant lines. Interestingly, PtbHLH1b, which is related to RITMO1, shows a very similar but even more pronounced expression pattern.…”

“…Recently, it was shown that the light acclimation ability in diatoms instead is strongly dependent on the aureochrome photoreceptor system ( Schellenberger Costa et al., 2013 ), which is lacking in the green lineage. PtbHLH1a (also called RITMO1) is involved in regulation of circadian rhythm of diatoms ( Annunziata et al., 2019 ). Transcription of RITMO1 in our experiment is increased by about 30-fold upon blue light exposure for 10 min, suggesting that its rhythmicity might be triggered by activation of aureochromes and/or other photoreceptors.…”

The Vertical Migratory Rhythm of Intertidal Microphytobenthos in Sediment Depends on the Light Photoperiod, Intensity, and Spectrum: Evidence for a Positive Effect of Blue Wavelengths

“…The fact that N. spartinetensis shows a much smaller size than G. fasciola, and probably a different migratory behavior (see Underwood et al, 2005) is an additional asset. The complementary use of the epipelic model Seminavis robusta (Blommaert et al, 2017(Blommaert et al, , 2018, which genome has been sequenced, will allow to search for molecular players, such as timekeepers (Annunziataa et al, 2019), photoreceptors (Jaubert et al, 2017), and diurnal-responsive genes (see Meyer et al, 2003;Blommaert et al, 2020), of the rhythmic migratory light-tuned response of intertidal epipelic MPB. JL performed samples and data analyses.…”

“…The endogenously controlled rhythmic MPB migration, modeled onto intertidal flat environmental conditions, would allow MPB to perform surface and depth biological processes in the most optimal conditions. Although endogenous circadian clocks, their components and regulation, have been well detailed in several algal lineages, they remain uncharacterized in diatoms (Noordally and Millar, 2015), with the exception of the recent discovery of a time-keeping molecular component, which controls the diel rhythmic expression of many genes (Annunziataa et al, 2019).…”

PhaeoNet: A Holistic RNAseq-Based Portrait of Transcriptional Coordination in the Model Diatom Phaeodactylum tricornutum

“…A total of 188 candidates, from 18 TF families (HSF, Myb, Zn_finger_C2H2, bZIP, Zn_finger_CCCH, bHLH, Sigma-70, Zn_finger_TAZ, CBF/NF, E2F-DP, CSF, Aureochrome, TRF, CCAAT-binding, AP2-EREBP, TAF9, CXC, Homeobox) corresponded to genes assigned to a PhaeoNet merged module ( Figure 5 and Supplementary Table 4). Given that the regulation of gene expression by transcription factors play a key role in the growth and progression of the cell cycle, the distribution within merged modules genes implicated in the cell cycle (cyclins) and in light perception events (e.g., phytochrome, cryptochrome) were additionally investigated, as well as genes implied in transcription and histone-related processes ( Figure 5 and Supplementary Table 4; Huysman et al, 2013;Annunziata et al, 2019).…”