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Professor of Biological Sciences and of Genetics
Genetics And Molecular Genetics Of Plant Circadian Rhythms
The ability of an organism to measure time is the product of a cellular
biological clock. Many phenomena controlled by the biological clock cycle
on a daily basis and are called circadian rhythms. My goal is to
understand the genetic and biochemical mechanisms by which an organism measures
time and uses that temporal information to regulate gene expression and
cellular physiology. The circadian clock is an endogenous oscillator that
drives rhythms with periods of approximately 24 hours. By definition,
these circadian (from the Latin, circa, approximately; dies,
day) rhythms persist in constant conditions and reflect the activity of an
endogenous biological clock. Plants are richly rhythmic and the circadian
clock regulates a number of key metabolic pathways and stress responses.
In addition, the circadian clock plays a critical role in the photoperiodic
regulation of the transition to flowering in many species. Circadian
rhythms in plants have been the subject of a number of recent reviews,
including several from my lab (McClung, 2001; McClung et al., 2002; Salomé and
McClung, 2004, 2005b; McClung, 2006).
Ongoing Research Projects
1. Mutational Analysis of the Arabidopsis Circadian
Clock
We have identified a number of loci which, when mutated, alter circadian
rhythmicity (period or phase is altered, or the plants are arrhythmic).
These studies include novel genes identified in forward genetic screens as well
as defined genes identified through a candidate gene (reverse genetics)
approach. The genetic and molecular biological analysis of these
mutations is ongoing (Salomé et al., 2002; Michael et al., 2003b; Tseng et al.,
2004; Lidder et al., 2005; Salomé and McClung, 2005a, b).
2. Temperature Sensitivity of the Circadian Oscillator
Regulating CAT3
In most studies, one synchronizes the clocks of a population of seedlings by
exposing them to a series of light-dark cycles, but temperature cycles are also
effective in the entrainment of the Arabidopsis clock. Little is known of
the mechanisms by which temperature signals are perceived and transmitted into
the clock (Salomé and McClung, 2005a). Calcium signaling, temperature
sensitive changes in RNA secondary structure, altered splicing patterns, and
membrane fluidity changes have all been implicated in temperature
signaling. We have shown that the clock regulating CAT3
transcription is more sensitive to temperature signals than the clock
regulating CAB2 transcription (Michael et al., 2003a). We wish
to characterize temperature signaling—for example, is the potency of a
temperature cycle determined by the temperature differential between warm and
cold or, instead, by the absolute value of the cold temperature, as is
suggested by the results of preliminary studies. We already know that the
prr7 prr9 double mutant is defective in clock responses to temperature
cues (Salomé and McClung, 2005a). PRR7 and PRR9 are two members of a
five-gene family of Pseudo-Response Regulators. All members of the family
are important for clock function, although their biochemical functions remain
uncertain. We have initiated genetic screens to identify other loci
required for appropriate input of temperature signals to the clock and a number
of putative mutants have been identified. These mutants need to be
verified and cloned by positional cloning.
3. Temperature Compensation of the Circadian Clock
One of the hallmarks of circadian clocks is that they run at approximately
the same pace at different temperatures throughout the physiological
range. This makes sense, as a clock that slowed down on a cloudy day or
intermittently as clouds passed overhead would be unreliable. However,
clocks are not independent of temperature. Rather, organisms have
developed mechanisms to compensate for the increased pace at elevated
temperature, and for the decreased pace at reduced temperature. It is not
know how this is accomplished in Arabidopsis, although the prr7 prr9
double mutant (Salomé, unpublished data) and other prr single and
double mutant combinations (Xie, unpublished data) are defective in clock
responses to temperature cues.
4. Evolutionary and Quantitative Analysis of the Arabidopsis
Circadian Clock
PSEUDO-RESPONSE REGULATOR 7 (PRR7) is a gene that plays a
role in the Arabidopsis thaliana circadian clock, because eliminating
function of the PRR7 gene results in a lengthening of the circadian
period. Also, we had mapped a number of loci (Quantitative Trait Loci; QTL)
that contribute incrementally to the definition of period length and one of the
QTL’s mapped near PRR7 (Michael et al., 2003b). We wish to determine
whether PRR7 encodes a period QTL. If it does, we would predict
there to be different PRR7 alleles among multiple accessions (natural
populations). Sequence analysis confirms this prediction; most simply these can
be divided into two clades that show distinct geographic distributions, with
one clade in southwestern Europe and North Africa and the second clade in
central and northeastern Europe. Excitingly, the distribution of
mutations among these PRR7 alleles suggests that PRR7 is
undergoing strong diversifying selection, consistent with the hypothesis that
selection is acting to enrich multiple PRR7 forms, each of which is
suited to a particular geographic habitat. Flanking genes fail to show
similar distributions of mutations, consistent with natural selection acting on
PRR7. We are extending this analysis to other “clock genes,”
including PRR9 and PRR1 (TOC1).
We intend to take two additional approaches concerning the identity of
PRR7 as the QTL we identified. The first is a genetic approach
in which we generate two sets of Near Isogenic Lines (NILs), in which the
PRR7 allele from one Arabidopsis accession, Col, is introduced into a
second accession, Ler and vice versa. One would predict that the Col and
Ler PRR7 alleles would result in distinct periods. The second
approach is more molecular biological. We would introduce the
PRR7 alleles from several different accessions into a loss-of-function
prr7 prr9 double mutant and ask whether each of the different alleles
can restore PRR7 function equally well.
5. Post-transcriptional Regulation by the Arabidopsis
Circadian Clock.
A hallmark of circadian control is persistence in the absence of external
time signals. CAT3 transcription is robustly rhythmic in either
continuous light or continuous dark. Curiously, a number of years ago we
showed that the abundance of the CAT3 transcript does not continue to
oscillate in the dark (Zhong et al., 1997), despite rhythmic
transcription. Apparently clock modulation of CAT3 mRNA
stability plays a role in circadian oscillations in transcript abundance. We
wish to study this with CAT3 mRNA half-life determinations and the
identification of CAT3 mRNA half-life determinants. This then
will allow us to use genetic and molecular biological approaches to identify
other genetic loci required for CAT3 mRNA half-life determination. See
Lidder et al. (2005)
6. Evolutionary and Quantitative Analysis of the Brassica
rapa Circadian Clock
We have started a new collaboration to look at clocks in a second crop
species, Brassica rapa (Chinese cabbage, turnip). This study
will parallel our QTL analyses in Arabidopsis, taking advantage of a new set of
RILs developed by colleagues at U Wisconsin-Madison and U. Missouri. We
are measuring clock parameters (again, by leaf movement) in these lines in
order to map QTLs that contribute to period, phase and amplitude. The
study is ongoing, but we already have identified two QTLs, one of which maps
near a B. rapa PRR7 ortholog. In addition, we will map genes
that contribute to temperature entrainment and temperature compensation.
Besides the QTL mapping, we will identify B. rapa orthologs of
Arabidopsis clock genes and test whether they play similar roles in the B.
rapa clock. Our collaborators will be conducting parallel studies on
flowering timing and inflorescence architecture as well as on fitness in field
experiments.
Visit the McClung
Lab website
Publications
Lidder, P., Gutiérrez, R.A., Salomé, P.A., McClung, C.R., and Green, P.J.
(2005). Circadian control of mRNA stability: association with DST-mediated mRNA
decay. Plant Physiol. 138, 2374-2385.
McClung, C.R. (2001). Circadian rhythms in plants. Annu. Rev. Plant Physiol.
Plant Mol. Biol. 52, 139-162.
McClung, C.R. (2006). Plant circadian rhythms. Plant Cell 18, 792-803.
McClung, C.R., Salomé, P.A., and Michael, T.P. (2002). The Arabidopsis
circadian system. In The Arabidopsis Book, C.R. Somerville and E.M. Meyerowitz,
eds (Rockville MD: American Society of Plant Biologists), pp. DOI
10.1199/tab.0044 http://www.aspb.org/publications/arabidopsis/.
Michael, T.P., Salomé, P.A., and McClung, C.R. (2003a). Two Arabidopsis
circadian oscillators can be distinguished by differential temperature
sensitivity. Proc. Natl. Acad. Sci. USA 100, 6878-6883.
Michael, T.P., Salomé, P.A., Yu, H.J., Spencer, T.R., Sharp, E.L., Alonso,
J.M., Ecker, J.R., and McClung, C.R. (2003b). Enhanced fitness conferred by
naturally occurring variation in the circadian clock. Science 302,
1049-1053.
Salomé, P.A., and McClung, C.R. (2004). The Arabidopsis thaliana
clock. J. Biol. Rhythms 19, 425-435.
Salomé, P.A., and McClung, C.R. (2005a). PRR7 and PRR9 are
partially redundant genes essential for the temperature responsiveness of the
Arabidopsis circadian clock. Plant Cell 17, 791-803.
Salomé, P.A., and McClung, C.R. (2005b). What makes Arabidopsis tick: Light
and temperature entrainment of the circadian clock. Plant Cell Environ. 28,
21-38.
Salomé, P.A., Michael, T.P., Kearns, E.V., Fett-Neto, A.G., Sharrock, R.A.,
and McClung, C.R. (2002). The out of phase 1 mutant defines a role for
PHYB in circadian phase control in Arabidopsis. Plant Physiol. 129,
1674-1685.
Tseng, T.-S., Salomé, P.A., McClung, C.R., and Olszewski, N.E. (2004).
SPINDLY and GIGANTEA interact and act in Arabidopsis thaliana pathways
involved in light responses, flowering and rhythms in leaf movements. Plant
Cell 16, 1550-1563.
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