CIRCADIAN RHYTHMS - Cyanobacteria - page 322

near the pole in presence of PsR domain, as revealed by green fl uorescent reporter, suggests that it
interacts with other components near the cell pole. But in the absence of PsR, CikA cannot localize
at cell poles but gets uniformly distributed though out the cell. These observations signify that it is
the PsR component that mediates interaction with other components of the input pathway (Zhang
et al ., 2006). Four component proteins of the input pathway working in tandem with CikA have
been identifi ed by Mackey et al . (2008). They have identifi ed four new genes nhtA (orf2160), prkE
(orf0600), ircA (orf2387) and cdpA (orf1604) that encode aminotransferase-1, phase-resetting kinase,
input-related cytochrome and cell division and phase protein, respectively. The involvement of these
four proteins in the CikA input pathway has been confi rmed by the isolation of mutants inactivated
in the respective genes by the insertion of an antibiotic resistance cassette. S . elongatus PCC 7942 with
P kaiBC :: luxAB luciferase as reporter gene was transformed with the inactivated nhtA / prkE / ircA / cdpA
gene and allowed to segregate so that mutant strains possessed corresponding inactivated genes in
question. The nhtA null mutant behaved very much in a similar fashion as that of the wild-type but
the prkE null mutant exhibited an unpredictable behavior with either the phase being advanced or
delayed by about 6 to 11 h. The cikAprkE double mutant resembled a cikA null mutant in showing a
non-resetting phenotype. The ircA and cdpA genes could not be inactivated completely as revealed
by existence of wild-type copies of these genes by PCR analysis. Overexpression mutants of these
genes against wild-type and cikA background have been isolated and their analysis revealed that
the corresponding proteins are involved in either circadian period regulation, phase resetting and
cell division. Mackey et al. (2008) are of the opinion that these four proteins function through protein
phosphorylation, iron-cluster biosynthesis and redox regulation.
SasA ( Synechococcus adaptive sensor A) is a histidine kinase (Iwasaki et al. , 2000). This is a member
of the bacterial two-component regulatory system. It senses specifi c external stimuli or ligands and
activates its autophosphorylation at a conserved His-residue and then transfers the phosphoryl
group to an Asp residue in the cognate response regulator. SasA binds to KaiC to form a complex.
This regulator drives physiological responses to the stimulus or signal by activating downstream
genes or enzyme activities. The sensory domain of SasA has an amino acid sequence similar to that
of full length KaiB and is suffi ciently similar to bind to KaiC (Iwasaki et al . 2000). In sasA -inactivated
strains, kaiBC expression is dramatically lowered with a concomitant lowering of the amplitude
in rhythm and shortening of the period. SasA overexpression also strongly interfered with the
rhythmicity. Although SasA is not required to drive a basic oscillation, it forms a secondary loop
to stabilize Kai-based timing loop by coupling with KaiC. SasA levels essentially remain constant
throughout the circadian cycle. SasA is important for growth in LD cycle conditions. The mutants
devoid of SasA did not show any growth in LL. Iwasaki and Kondo (2004) concluded that SasA is
interesting as a multifunctional regulator affecting the clock system, photoresponses and metabolism.
Recently, Takai et al. (2006) identifi ed a DNA-binding response regulator, RpaA (Regulator of
phycobilisome associated) as a part of SasA. Mutants of S . elongatus PCC 7942 defective in SasA and
RpaA showed a severe inhibition in circadian transcription. This two-component regulatory system
is responsible for receiving the circadian time keeping signal from the oscillator to the transcription
machinery. Moreover, autophosphorylation of SasA is enhanced in response to KaiC binding (Smith
and Williams, 2006; Takai et al ., 2006) and this phosphate group is then transferred to RpaA (Takai
et al ., 2006). According to Takai et al. (2006) the KaiC-SasA-RpaA pathway is likely to be the major
pathway for circadian gene expression. This has further been confi rmed by Taniguchi et al. (2007)
who reported a novel gene labA (low amplitude and bright) that plays a role in negative feedback
regulation of KaiC. The overexpression of labA signifi cantly lowered circadian gene expression
whereas its disruption abolished transcriptional repression caused by overexpression of KaiC.