bims-ciryme 2022-07-17 papers

Nat Commun. 2022 Jul 09. 13(1): 3991

Decoupling PER phosphorylation, stability and rhythmic expression from circadian clock function by abolishing PER-CK1 interaction.

Yang An, Baoshi Yuan, Pancheng Xie, Yue Gu, Zhiwei Liu, Tao Wang, Zhihao Li, Ying Xu, Yi Liu.

Robust rhythms of abundances and phosphorylation profiles of PERIOD proteins were thought be the master rhythms that drive mammalian circadian clock functions. PER stability was proposed to be a major determinant of period length. In mammals, CK1 forms stable complexes with PER. Here we identify the PER residues essential for PER-CK1 interaction. In cells and in mice, their mutation abolishes PER phosphorylation and CLOCK hyperphosphorylation, resulting in PER stabilization, arrhythmic PER abundance and impaired negative feedback process, indicating that PER acts as the CK1 scaffold in circadian feedback mechanism. Surprisingly, the mutant mice exhibit robust short period locomotor activity and other physiological rhythms but low amplitude molecular rhythms. PER-CK1 interaction has two opposing roles in regulating CLOCK-BMAL1 activity. These results indicate that the circadian clock can function independently of PER phosphorylation and abundance rhythms due to another PER-CRY-dependent feedback mechanism and that period length can be uncoupled from PER stability.

DOI: https://doi.org/10.1038/s41467-022-31715-4

Cell Rep. 2022 Jul 12. pii: S2211-1247(22)00857-9. [Epub ahead of print]40(2): 111059

A guiding role of the Arabidopsis circadian clock in cell differentiation revealed by time-series single-cell RNA sequencing.

Kotaro Torii, Keisuke Inoue, Keita Bekki, Kazuya Haraguchi, Minoru Kubo, Yuki Kondo, Takamasa Suzuki, Akane Kubota, Kyohei Uemoto, Hanako Shimizu, Masato Saito, Hiroo Fukuda, Takashi Araki, Motomu Endo.

Circadian rhythms and progression of cell differentiation are closely coupled in multicellular organisms. However, whether establishment of circadian rhythms regulates cell differentiation or vice versa has not been elucidated due to technical limitations. Here, we exploit high cell fate plasticity of plant cells to perform single-cell RNA sequencing during the entire process of cell differentiation. By analyzing reconstructed actual time series of the differentiation processes at single-cell resolution using a method we developed (PeakMatch), we find that the expression profile of clock genes is changed prior to cell differentiation, including induction of the clock gene LUX ARRYTHMO (LUX). ChIP sequencing analysis reveals that LUX induction in early differentiating cells directly targets genes involved in cell-cycle progression to regulate cell differentiation. Taken together, these results not only reveal a guiding role of the plant circadian clock in cell differentiation but also provide an approach for time-series analysis at single-cell resolution.

Keywords: CP: Molecular biology; CP: Plants; cell differentiation; circadian clock; single-cell RNA sequencing

DOI: https://doi.org/10.1016/j.celrep.2022.111059

Methods Mol Biol. 2022 ;2525 321-332

Time-Lapse Bioluminescence Imaging of Hes7 Expression In Vitro and Ex Vivo.

Marina Sanaki-Matsumiya, Ryoichiro Kageyama.

Somites are formed sequentially by the segmentation of the anterior parts of the presomitic mesoderm (PSM), and such periodical somite formation is crucial to ensure the proper vertebrae. In the mouse embryo, Hes7, a segmentation clock gene, controls this periodic event with new somites forming every 2 h. Hes7 oscillations are synchronized between neighboring PSM cells and propagate from the posterior to the anterior PSM in the form of traveling waves. However, the exact mechanisms that generate these oscillatory dynamics and control synchronization are still unclear. Given that the half-life of Hes7 is too short to be monitored with most fluorescent proteins, time-lapse bioluminescence imaging (BLI) is a suitable tool to monitor the chronological Hes7 expression dynamics. In this chapter, we introduce a ubiquitinated luciferase reporter which enables the visualization of Hes7 expression dynamics with high temporal and spatial resolution in living cells and tissues.

Keywords: Embryonic stem (ES) cells; Hes7; Luciferase; Mus musculus; Oscillation; Segmentation clock; Time-lapse imaging

DOI: https://doi.org/10.1007/978-1-0716-2473-9_25