Chesham, J. E. | Search bioRxiv

bioRxiv2020

The cell-autonomous clock of VIP receptor VPAC2 cells drives circadian behaviour

Circadian ([~]daily) rhythms pervade mammalian behaviour. They are generated by cell-autonomous, transcriptional/translational feedback loops (TTFL), active in all tissues. This distributed clock network is co-ordinated by the principal circadian pacemaker, the hypothalamic suprachiasmatic nucleus (SCN). Its robust and accurate time-keeping arises from circuit-level interactions that bind its individual cellular clocks into a coherent time-keeper. Cells that express the neuropeptide vasoactive intestinal peptide (VIP) mediate retinal entrainment of the SCN, and in the absence of VIP, or its cognate receptor VPAC2, circadian behaviour is compromised because SCN cells cannot synchronise. The contributions to SCN pacemaking and circadian behaviour of other cell types, not least the VPAC2-expressing target cells of VIP, are, however, not understood. We therefore employed intersectional genetics to manipulate the cell-autonomous TTFL of VPAC2-expressing cells, creating temporally chimaeric mice. We could then determine whether and how VPAC2-expressing cells (a minority [~]35% of SCN cells) contribute to SCN time-keeping. Lengthening of the intrinsic TTFL period of VPAC2 cells by deletion of the CK1{varepsilon}Tau allele concomitantly lengthened the period of circadian behavioural rhythms. It also increased the variability of the circadian period of bioluminescent TTFL rhythms in SCN slices recorded ex vivo. Abrogation of circadian competence in VPAC2 cells by deletion of Bmal1 severely disrupted circadian behavioural rhythms and compromised TTFL time-keeping in the corresponding SCN slices. Thus, VPAC2-expressing cells are a distinct, functionally powerful subset of the SCN circuit, contributing to computation of ensemble period and maintenance of circadian robustness. These findings extend our understanding of SCN circuit topology.

neuroscience↗

bioRxiv2020

CRYPTOCHROMES confer robustness, not rhythmicity, to circadian timekeeping

SummaryCircadian (approximately daily) rhythms are a pervasive property of mammalian cells, tissues, and behaviour, ensuring physiological and metabolic adaptation to solar time. Models of daily cellular timekeeping revolve around transcriptional feedback repression, whereby CLOCK and BMAL1 activate the expression of clock proteins PERIOD (PER) and CRYPTOCHROME (CRY), which in turn repress CLOCK/BMAL1 activity. CRY proteins are thus considered essential negative regulators of the oscillation; a function supported by behavioural arrhythmicity of CRY-deficient mice when kept under constant conditions. Challenging this interpretation, however, we find evidence for persistent circadian rhythms in mouse behaviour and cellular PER2 levels when CRY is absent. CRY-less oscillations are variable in their expression and have a shorter period than wild type controls. Importantly, we find classic circadian hallmarks such as temperature compensation and determination of period by casein kinase 1{delta}/{varepsilon} activity to be maintained. In the absence of CRY-mediated transcriptional feedback repression and rhythmic Per2 transcription, PER2 protein rhythms are sustained for several cycles, accompanied by circadian variation in protein stability. We suggest that, whereas circadian transcriptional feedback imparts robustness and functionality onto biological clocks, the core timekeeping mechanism is post-translational. Our findings suggest that PER proteins normally act as signalling hubs that transduce timing information to the nucleus, imparting daily rhythms upon the activity of transcriptional effectors. Highlights[tpltrtarr] PER/CRY-mediated negative feedback is dispensable for mammalian circadian timekeeping [tpltrtarr]Circadian variation in PER2 levels persists in the absence of rhythmic Per2 transcription [tpltrtarr]CK1 and GSK3 are plausible mechanistic components of a cytoscillator mechanism [tpltrtarr]CRY-mediated feedback repression imparts robustness to biological timekeeping In briefCircadian turnover of mammalian clock protein PERIOD2 persists in the absence of canonical transcriptional feedback repression and rhythmic clock gene activity, demanding a re-evaluation of cellular clock function and evolution.

cell biology↗