Dynamic Visualization Made Of Simplest Circadian Clock

A variety of organisms have evolved endogenous timing systems called a circadian clock that allows them to regulate metabolic activities in a day/night cycle. The simplest organisms known to possess a circadian oscillator are the cyanobacteria, better known as blue-green algae. The essential components of the circadian oscillator in cyanobacteria are the three clock proteins KaiA, KaiB and KaiC, all of which are expressed in the cyanobacterium S. elongatus.

Considerable research has implicated the phosphorylation cycle of KaiC as the central pacemaker in cyanobacteria and has demonstrated that the Kai proteins are repeatedly assembled and disassembled into heteromultimeric complexes, termed periodosomes. The crystal structure of each clock protein has also been determined and analyzed.

"Despite the substantial progress in structural characterization, the relationship between the assembly/disassembly dynamics and the circadian phosphorylation of KaiC is still poorly understood, mainly because of the difficulty in unraveling the underlying mechanisms solely from the static molecular pictures of individual clock components," explains Dr. Akiyama from the Japan Science and Technology Agency.

To obtain a more complete visualization of the cyanobacterial circadian oscillator, Dr. Akiyama and colleagues used small-angle X-ray scattering (SAXS) to follow the assembly/disassembly dynamics of the S. elongatus heteromultimeric Kai complexes in real time. The researchers found that the assembly/disassembly processes are crucial for phase entrainment in the early synchronizing stage but are passively driven by the phosphorylation status of KaiC in the late oscillatory stage. Further, KaiA and KaiB are recruited to KaiC in a phosphorylation-dependent manner.

"Our findings demonstrate that the initial phase of the cyanobacterial oscillator is determined predominantly by the assembly/disassembly communication of the clock components, and that the period is essentially resistant to intracellular noise such as collisions, cytoplasmic viscosity and crowding. These resistances are achieved in the binary and ternary complexes by recruiting KaiA homodimers, KaiB homotetramers and KaiC homohexamers in a phosphorylation-dependent manner," concludes Dr. Akiyama.

This research is published online March 13th by Cell Press in the journal Molecular Cell.

The researchers include Shuji Akiyama, PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama, Japan; RIKEN SPring-8 Center, Harima Institute; Atsushi Nohara, Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Japan; Kazuki Ito, RIKEN SPring-8 Center, Harima Institute; and Yuichiro Maeda, ERATO Actin Filament Dynamics Project, Japan Science and Technology Agency, c/o RIKEN, Harima SPring-8 Center, Structural Biology Research Center, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Japan.