Stanly B. Williams

A KaiC-Interacting Sensory Histidine Kinase, SasA, Necessary to Sustain Robust Circadian Oscillation in Cyanobacteria

Both regulated expression of the clock genes kaiA, kaiB, and kaiC and interactions among the Kai proteins are proposed to be important for circadian function in the cyanobacterium Synechococcus sp. strain PCC 7942. We have identified the histidine kinase SasA as a KaiC-interacting protein. SasA contains a KaiB-like sensory domain, which appears sufficient for interaction with KaiC. Disruption of the sasA gene lowered kaiBC expression and dramatically reduced amplitude of the kai expression rhythms while shortening the period. Accordingly, sasA disruption attenuated circadian expression patterns of all tested genes, some of which became arrhythmic. Continuous sasA overexpression eliminated circadian rhythms, whereas temporal overexpression changed the phase of kaiBC expression rhythm. Thus, SasA is a close associate of the cyanobacterial clock that is necessary to sustain robust circadian rhythms.

Structure and function from the circadian clock protein KaiA of Synechococcus elongatus: A potential clock input mechanism

In the cyanobacterium Synechococcus elongatus (PCC 7942) the proteins KaiA, KaiB, and KaiC are required for circadian clock function. We deduced a circadian clock function for KaiA from a combination of biochemical and structural data. Both KaiA and its isolated carboxyl-terminal domain (KaiA180C) stimulated KaiC autophosphorylation and facilitated attenuation of KaiC autophosphorylation by KaiB. An amino-terminal domain (KaiA135N) had no function in the autophosphorylation assay. NMR structure determination showed that KaiA135N is a pseudo-receiver domain. We propose that this pseudo-receiver is a timing input-device that regulates KaiA stimulation of KaiC autophosphorylation, which in turn is essential for circadian timekeeping.

MicroReview: Functional similarities among two‐component sensors and methyl‐accepting chemotaxis proteins suggest a role for linker region amphipathic helices in transmembrane signal transduction

Signal‐responsive components of transmembrane signal‐transducing regulatory systems include methyl‐accepting chemotaxis proteins and membrane‐bound, two‐component histidine kinases. Prokaryotes use these regulatory networks to channel environmental cues into adaptive responses. A typical network is highly discriminating, using a specific phosphoryl relay that connects particular signals to appropriate responses. Current understanding of transmembrane signal transduction includes periplasmic signal binding with the subsequent conformational changes being transduced, via transmembrane helix movements, into the sensory proteins cytoplasmic domain. These induced conformational changes bias the proteins regulatory function. Although the mutational analyses reviewed here identify a role for the linker region in transmembrane signal transduction, no specific mechanism of linker function has yet been described. We propose a speculative, mechanistic model for linker function based on interactions between two putative amphipathic helices. The model attempts to explain both mutant phenotypes and hybrid sensor data, while accounting for recognized features of amphipathic helices.

Letter to the Editor: Sequence-specific 1H, 13C and 15N resonance assignments of the N-terminal, 135-residue domain of KaiA, a clock protein from Synechococcus elongatus

An endogenous, self-sustaining circadian clock (also called an oscillator or pacemaker) modulates metabolic, physiological and behavioral activities of virtually all organisms with a period of ∼24 h. The fact that evolutionarily divergent organisms are commonly endowed with the ability to anticipate daily environmental oscillations suggests that a healthy circadian pacemaker is of fundamental importance to the survival of most life forms. In the past few years large strides have been made in elucidating the genetic and biochemical bases of the circadian clock in cyanobacteria, fungi, plants, insects, and mammals; however, the structural basis of any circadian clock remains unknown as no clock protein structures have been determined yet. The basic timing oscillator of the cyanobacterium Synechococcus elongatus consists of three proteins KaiA, KaiB and KaiC whose expression and mutual interactions drive the circadian rhythm (Ishiura et al., 1998; Xu et al., 2000; Iwasaki et al., 1999). Point mutations in these proteins are known to alter these interactions and thereby the periodicity of the circadian clock. KaiA acts as the positive element of the oscillator by enhancing kaiBC expression and therefore maintains the robustness of the oscillation. We have been able to identify a stable, independently folded domain for KaiA, which resists proteolytic cleavage by trypsin for up to 5 h. This domain,