Quantitative live imaging of Venus::BMAL1 in a mouse model reveals complex dynamics of the master circadian clock regulator

Autoři: Nan Yang aff001;  Nicola J. Smyllie aff003;  Honor Morris aff001;  Cátia F. Gonçalves aff001;  Michal Dudek aff001;  Dharshika Pathiranage aff001;  Johanna E. Chesham aff003;  Antony Adamson aff002;  David Spiller aff002;  Egor Zindy aff002;  James Bagnall aff002;  Neil Humphreys aff002;  Judith Hoyland aff002;  Andrew S. I. Loudon aff002;  Michael H. Hastings aff003;  Qing-Jun Meng aff001;  Dharshika R. J. Pathiranage aff001;  David G. Spiller aff002
Působiště autorů: Wellcome Centre for Cell Matrix Research, University of Manchester, Manchester, United Kingdom aff001;  Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom aff002;  Division of Neurobiology, Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge, United Kingdom aff003;  NIHR Manchester Musculoskeletal Biomedical Research Centre, Manchester University NHS Foundation Trust, Manchester Academic Health Science Centre, Manchester, United Kingdom aff004
Vyšlo v časopise: Quantitative live imaging of Venus::BMAL1 in a mouse model reveals complex dynamics of the master circadian clock regulator. PLoS Genet 16(4): e32767. doi:10.1371/journal.pgen.1008729
Kategorie: Research Article
doi: 10.1371/journal.pgen.1008729


Evolutionarily conserved circadian clocks generate 24-hour rhythms in physiology and behaviour that adapt organisms to their daily and seasonal environments. In mammals, the suprachiasmatic nucleus (SCN) of the hypothalamus is the principal co-ordinator of the cell-autonomous clocks distributed across all major tissues. The importance of robust daily rhythms is highlighted by experimental and epidemiological associations between circadian disruption and human diseases. BMAL1 (a bHLH-PAS domain-containing transcription factor) is the master positive regulator within the transcriptional-translational feedback loops (TTFLs) that cell-autonomously define circadian time. It drives transcription of the negative regulators Period and Cryptochrome alongside numerous clock output genes, and thereby powers circadian time-keeping. Because deletion of Bmal1 alone is sufficient to eliminate circadian rhythms in cells and the whole animal it has been widely used as a model for molecular disruption of circadian rhythms, revealing essential, tissue-specific roles of BMAL1 in, for example, the brain, liver and the musculoskeletal system. Moreover, BMAL1 has clock-independent functions that influence ageing and protein translation. Despite the essential role of BMAL1 in circadian time-keeping, direct measures of its intra-cellular behaviour are still lacking. To fill this knowledge-gap, we used CRISPR Cas9 to generate a mouse expressing a knock-in fluorescent fusion of endogenous BMAL1 protein (Venus::BMAL1) for quantitative live imaging in physiological settings. The Bmal1Venus mouse model enabled us to visualise and quantify the daily behaviour of this core clock factor in central (SCN) and peripheral clocks, with single-cell resolution that revealed its circadian expression, anti-phasic to negative regulators, nuclear-cytoplasmic mobility and molecular abundance.

Klíčová slova:

Cartilage – Circadian oscillators – Circadian rhythms – Fluorescence imaging – Fluorescence recovery after photobleaching – Chondrocytes – Chronobiology – Mice


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