dational in demonstrating that the positive arm
serves the crucial function of integrating light
cues into the clock.
While the WCC is the point protein for
clock entrainment to light cues, the activity of
another protein, Vivid (VVD), is also crucial for
proper clock phase timing based on light in the
environment (Fig. 4.2a). A small protein, only
containing an N-terminal cap and a LOV
domain, VVD plays an important role in light
integration into the clock by altering the activity of the WCC (Zoltowski et al. 2007; Hunt
et al. 2010; Chen et al. 2010). The WCC induces
vvd expression in response to light, after which
VVD interacts with WC-1’s LOV domain and
blocks the transcriptional capabilities of WC-1
(Zoltowski et al. 2007; Hunt et al. 2010; Malzahn
et al. 2010; Chen et al. 2010; Dasgupta et al.
2015). The action of VVD on the WCC is an
important example of photoadaptation, ensuring that light-induced genes are only transiently
upregulated (Hunt et al. 2010; Chen et al. 2010).
VVD’s photoadaptive activities are not only
important for keeping the clock from resetting
inappropriately, such as during nights with
bright moonlight, but also in supporting the
optimal timing of light responses by the WCC
(Heintzen et al. 2001; Malzahn et al. 2010).
B. The Effect of Temperature on the Negative
Arm of the Clock
Though it is logical to think of light as the
primary environmental input for the clock, surprisingly many other factors, such as temperature, can strongly impact the circadian
mechanism. For example, temperature changes
can reset the phase of the clock, such as an
increase from a low to a high temperature.
While light input is chiefly transduced via the
positive arm (see Sects. II.A and III.A), evidence from N. crassa shows that temperature
input is primarily conveyed by changes in the
molecular dynamics of the negative arm of the
clock (Liu et al. 1997; Pregueiro et al. 2005,
2006; Diernfellner et al. 2005), particularly the
dynamics of the levels of the core negative arm
protein frequency (FRQ) (see Sect. III.B.1 and
Fig. 4.2a). The current understanding of the
response to temperature shifts is that while
the level of frq mRNA remains stable at different temperatures, FRQ protein levels oscillate
around a higher midline at higher temperatures, with the peak and trough of FRQ levels
falling above the peak level of FRQ in a strain
grown at a low enough temperature (Liu et al.
1997). Due to the difference in FRQ levels at
different temperatures, the comparatively low
levels of FRQ at a lower temperature will create
an artificial “trough” from which FRQ levels
rise when the organism is shifted to a higher
temperature, thereby resetting the phase of the
clock (Liu et al. 1998). A decrease in temperature can have the same resetting effect, as FRQ
levels decrease from an artificial “peak” time in
higher temperatures to start oscillating around
a lower midline of protein levels (Liu et al.
1998). Extreme temperature changes (Æ10
C)
have a strong effect on setting the phase of the
clock, so much so that it supersedes the effects
of light, re-entraining the molecular clock even
in a conflicting light-dark regime (Liu et al.
1998). This research paved the way for similar
discoveries in higher eukaryotes. Temperature
has subsequently been shown to be an important phase resetting cue in mammal peripheral
tissues, such that daily temperature changes
within physiological ranges (36–37.5
C) can
entrain mouse peripheral tissues (Buhr et al.
2010). Moreover, within Drosophila, the mutation of all detected phosphorylation sites to
alanine on the positive arm clock protein
dCLOCK led to compromised temperature
resetting, but unaffected light resetting, suggesting that temperature is more important
than light as a timing cue for Drosophila tissues
as well (Lee et al. 2014).
Though temperature changes can shift the
phase of the clock, one of the key tenets of a
circadian rhythm is that its period is compensated and regulated at different growing temperatures. However, the underlying mechanism
for temperature regulation in the clock is
poorly understood. What is known is that temperature impacts the choice of FRQ start
codons during translation (Colot et al. 2005;
Diernfellner et al. 2007; Liu et al. 1997). A
temperature-sensitive alternate splicing mechanism occurs in frq mRNA, leading to different
82
M. S. Jankowski et al.
serves the crucial function of integrating light
cues into the clock.
While the WCC is the point protein for
clock entrainment to light cues, the activity of
another protein, Vivid (VVD), is also crucial for
proper clock phase timing based on light in the
environment (Fig. 4.2a). A small protein, only
containing an N-terminal cap and a LOV
domain, VVD plays an important role in light
integration into the clock by altering the activity of the WCC (Zoltowski et al. 2007; Hunt
et al. 2010; Chen et al. 2010). The WCC induces
vvd expression in response to light, after which
VVD interacts with WC-1’s LOV domain and
blocks the transcriptional capabilities of WC-1
(Zoltowski et al. 2007; Hunt et al. 2010; Malzahn
et al. 2010; Chen et al. 2010; Dasgupta et al.
2015). The action of VVD on the WCC is an
important example of photoadaptation, ensuring that light-induced genes are only transiently
upregulated (Hunt et al. 2010; Chen et al. 2010).
VVD’s photoadaptive activities are not only
important for keeping the clock from resetting
inappropriately, such as during nights with
bright moonlight, but also in supporting the
optimal timing of light responses by the WCC
(Heintzen et al. 2001; Malzahn et al. 2010).
B. The Effect of Temperature on the Negative
Arm of the Clock
Though it is logical to think of light as the
primary environmental input for the clock, surprisingly many other factors, such as temperature, can strongly impact the circadian
mechanism. For example, temperature changes
can reset the phase of the clock, such as an
increase from a low to a high temperature.
While light input is chiefly transduced via the
positive arm (see Sects. II.A and III.A), evidence from N. crassa shows that temperature
input is primarily conveyed by changes in the
molecular dynamics of the negative arm of the
clock (Liu et al. 1997; Pregueiro et al. 2005,
2006; Diernfellner et al. 2005), particularly the
dynamics of the levels of the core negative arm
protein frequency (FRQ) (see Sect. III.B.1 and
Fig. 4.2a). The current understanding of the
response to temperature shifts is that while
the level of frq mRNA remains stable at different temperatures, FRQ protein levels oscillate
around a higher midline at higher temperatures, with the peak and trough of FRQ levels
falling above the peak level of FRQ in a strain
grown at a low enough temperature (Liu et al.
1997). Due to the difference in FRQ levels at
different temperatures, the comparatively low
levels of FRQ at a lower temperature will create
an artificial “trough” from which FRQ levels
rise when the organism is shifted to a higher
temperature, thereby resetting the phase of the
clock (Liu et al. 1998). A decrease in temperature can have the same resetting effect, as FRQ
levels decrease from an artificial “peak” time in
higher temperatures to start oscillating around
a lower midline of protein levels (Liu et al.
1998). Extreme temperature changes (Æ10
C)
have a strong effect on setting the phase of the
clock, so much so that it supersedes the effects
of light, re-entraining the molecular clock even
in a conflicting light-dark regime (Liu et al.
1998). This research paved the way for similar
discoveries in higher eukaryotes. Temperature
has subsequently been shown to be an important phase resetting cue in mammal peripheral
tissues, such that daily temperature changes
within physiological ranges (36–37.5
C) can
entrain mouse peripheral tissues (Buhr et al.
2010). Moreover, within Drosophila, the mutation of all detected phosphorylation sites to
alanine on the positive arm clock protein
dCLOCK led to compromised temperature
resetting, but unaffected light resetting, suggesting that temperature is more important
than light as a timing cue for Drosophila tissues
as well (Lee et al. 2014).
Though temperature changes can shift the
phase of the clock, one of the key tenets of a
circadian rhythm is that its period is compensated and regulated at different growing temperatures. However, the underlying mechanism
for temperature regulation in the clock is
poorly understood. What is known is that temperature impacts the choice of FRQ start
codons during translation (Colot et al. 2005;
Diernfellner et al. 2007; Liu et al. 1997). A
temperature-sensitive alternate splicing mechanism occurs in frq mRNA, leading to different
82
M. S. Jankowski et al.