Supplemental Material Items in Supplemental Materials and their relationship to main text and main figures Figure S1 relates to Figure 1 and shows the phase relationship of different circadian transcripts. Figure S2 relates to the Result section Low-amplitude temperature fluctuations can synchronize circadian gene expression over a wide temperature range and shows that within the examined range of temperatures (30°C and 40°C) circadian clocks can be phase-entrained to low amplitude temperature oscillations. Figure S3 relates to Figure 5 and shows the phase relationship between the bioluminescence cycles produced by different reporter genes in cells exposed to T-cycles with different period lengths. Figure S4 relates to Figures 5 and S3 and shows the behavior of luminescence cycles entrained to a 12-hour T-cycle after release into a constant temperature Figure S5 relates to Figure 5 and shows the phase relationship of various rhythmically expressed transcripts in cells entrained to a very short T-cycle of only 10 hours. Figure S6 relates to Figure 6 and shows the downregulation of HSF1 and HFS2 expression by RNA interference using shRNAs. Table 1 lists the DNA primers and Taqman probes used in quantitative RT-PCR experiments
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Supplemental Material
Items in Supplemental Materials and their relationship to main text and main figures
Figure S1 relates to Figure 1 and shows the phase relationship of different circadian
transcripts.
Figure S2 relates to the Result section Low-amplitude temperature fluctuations can
synchronize circadian gene expression over a wide temperature range and shows that within
the examined range of temperatures (30°C and 40°C) circadian clocks can be phase-entrained
to low amplitude temperature oscillations.
Figure S3 relates to Figure 5 and shows the phase relationship between the bioluminescence
cycles produced by different reporter genes in cells exposed to T-cycles with different period
lengths.
Figure S4 relates to Figures 5 and S3 and shows the behavior of luminescence cycles
entrained to a 12-hour T-cycle after release into a constant temperature
Figure S5 relates to Figure 5 and shows the phase relationship of various rhythmically
expressed transcripts in cells entrained to a very short T-cycle of only 10 hours.
Figure S6 relates to Figure 6 and shows the downregulation of HSF1 and HFS2 expression
by RNA interference using shRNAs.
Table 1 lists the DNA primers and Taqman probes used in quantitative RT-PCR experiments
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Legends to Supplemental Figures
Figure S1. The expression of various mRNAs in NIH3T3 cells exposed to simulated
body temperature rhythms.
(A) Real-time RT-qPCR (TaqMan) was used to quantify mRNAs in whole cell RNAs
prepared from NIH3T3 cells at four-hour intervals during the sixth day of temperature
entrainment (orange) or during the sixth day at constant temperature (blue). The primer- and
probe-sequences used for measuring Bmal1, Rev-erbα, Per2, Per3, Cry1 mRNAs and Hsp90
pre-mRNA levels are given in Supplemental Table 1. Two different biological samples per
time point were analyzed. For graphical purposes, the same values are plotted twice.
Bioluminescence profiles of NIH3T3/Bmal1-luc cells subjected to the same temperature
conditions (B) and corresponding temperature recordings (C) are shown for comparison.
Figure S2. Temperature cycles can synchronize circadian clocks within a broad
temperature range.
NIH3T3/Bmal1-luc cells whose circadian oscillators were transiently synchronized by a
serum shock are efficiently re-synchronized by temperature cycles with different magnitudes
but with the same relative amplitudes. Note the different phase relationships between the
peaks of Bmal1-luciferase expression and the peak of temperature (blue curves), which likely
reflects an over-compensation of period length at reduced temperatures. Indeed, in cells
exposed to the indicated constant temperatures, free-running periods shortened when the
incubation temperatures were lowered (grey curves). Thus, the shorter period length at lower
temperatures elicits phase advances under phase-entrained conditions. All bioluminescence
data were filtered by moving average transformation.
Figure S3. The phase relationship of circadian gene expression in cells exposed to T-