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Harmonics of circadian gene transcription in mammals.

Hughes ME, DiTacchio L, Hayes KR, Vollmers C, Pulivarthy S, Baggs JE, Panda S, Hogenesch JB - PLoS Genet. (2009)

Bottom Line: We recently developed a method to look at transcriptional oscillations with unprecedented precision and accuracy using high-density time sampling.Strikingly, we found two clusters of genes that cycle at the second and third harmonic of circadian rhythmicity in liver, but not cultured cells.Validation experiments show that 12-hour oscillatory transcripts occur in several other peripheral tissues as well including heart, kidney, and lungs.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Institute for Translational Medicine and Therapeutics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
The circadian clock is a molecular and cellular oscillator found in most mammalian tissues that regulates rhythmic physiology and behavior. Numerous investigations have addressed the contribution of circadian rhythmicity to cellular, organ, and organismal physiology. We recently developed a method to look at transcriptional oscillations with unprecedented precision and accuracy using high-density time sampling. Here, we report a comparison of oscillating transcription from mouse liver, NIH3T3, and U2OS cells. Several surprising observations resulted from this study, including a 100-fold difference in the number of cycling transcripts in autonomous cellular models of the oscillator versus tissues harvested from intact mice. Strikingly, we found two clusters of genes that cycle at the second and third harmonic of circadian rhythmicity in liver, but not cultured cells. Validation experiments show that 12-hour oscillatory transcripts occur in several other peripheral tissues as well including heart, kidney, and lungs. These harmonics are lost ex vivo, as well as under restricted feeding conditions. Taken in sum, these studies illustrate the importance of time sampling with respect to multiple testing, suggest caution in use of autonomous cellular models to study clock output, and demonstrate the existence of harmonics of circadian gene expression in the mouse.

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Related in: MedlinePlus

High resolution profiling in the liver identified circadian and sub-ciradian rhythms.Liver samples were collected every hour for 48 h and analyzed with Affymetrix expression arrays. Rhythmic genes were identified using both COSOPT and Fisher's G-test at a false-discovery rate of <0.05. The period length of every rhythmic transcript was plotted as a histogram; clusters of rhythmic genes with period lengths of approximately 24 (>20 and <30 hours), 12 (>10 and <14 hours) and 8-hours (>7 and <9) were observed (A). In panels B–D, the microarray intensity from three examples was plotted against CT time. Bmal1 (B), Hspa5 (C), and Zfp560 (D) expression profiles demonstrate 24, 12, and 8 h period lengths, respectively.
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pgen-1000442-g001: High resolution profiling in the liver identified circadian and sub-ciradian rhythms.Liver samples were collected every hour for 48 h and analyzed with Affymetrix expression arrays. Rhythmic genes were identified using both COSOPT and Fisher's G-test at a false-discovery rate of <0.05. The period length of every rhythmic transcript was plotted as a histogram; clusters of rhythmic genes with period lengths of approximately 24 (>20 and <30 hours), 12 (>10 and <14 hours) and 8-hours (>7 and <9) were observed (A). In panels B–D, the microarray intensity from three examples was plotted against CT time. Bmal1 (B), Hspa5 (C), and Zfp560 (D) expression profiles demonstrate 24, 12, and 8 h period lengths, respectively.

Mentions: These tests were corrected for multiple comparisons post hoc using the method described by Storey and colleagues[27],[28]. Briefly, by examining the distribution of p-values from a given data set, an estimate of the proportion that are truly non-rhythmic can be derived. Using this approach to model the rate of false-discoveries, the p-value for each transcript, which estimates the frequency that a truly observation will be labeled as significant, can be converted to a more stringent q-value which instead estimates the frequency that significant observations are truly non-rhythmic. At a false discovery rate [27] of <0.05, over 3000 transcripts were found to oscillate by both statistical tests in liver, while fewer than a dozen were found in NIH3T3 and U2OS cells. As expected, the majority of cycling transcripts from liver (and all from NIH3T3 and U2OS cells) had period lengths of approximately 24 h (Figure 1A, Figure 2A and B, Table 1).


Harmonics of circadian gene transcription in mammals.

Hughes ME, DiTacchio L, Hayes KR, Vollmers C, Pulivarthy S, Baggs JE, Panda S, Hogenesch JB - PLoS Genet. (2009)

High resolution profiling in the liver identified circadian and sub-ciradian rhythms.Liver samples were collected every hour for 48 h and analyzed with Affymetrix expression arrays. Rhythmic genes were identified using both COSOPT and Fisher's G-test at a false-discovery rate of <0.05. The period length of every rhythmic transcript was plotted as a histogram; clusters of rhythmic genes with period lengths of approximately 24 (>20 and <30 hours), 12 (>10 and <14 hours) and 8-hours (>7 and <9) were observed (A). In panels B–D, the microarray intensity from three examples was plotted against CT time. Bmal1 (B), Hspa5 (C), and Zfp560 (D) expression profiles demonstrate 24, 12, and 8 h period lengths, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2654964&req=5

pgen-1000442-g001: High resolution profiling in the liver identified circadian and sub-ciradian rhythms.Liver samples were collected every hour for 48 h and analyzed with Affymetrix expression arrays. Rhythmic genes were identified using both COSOPT and Fisher's G-test at a false-discovery rate of <0.05. The period length of every rhythmic transcript was plotted as a histogram; clusters of rhythmic genes with period lengths of approximately 24 (>20 and <30 hours), 12 (>10 and <14 hours) and 8-hours (>7 and <9) were observed (A). In panels B–D, the microarray intensity from three examples was plotted against CT time. Bmal1 (B), Hspa5 (C), and Zfp560 (D) expression profiles demonstrate 24, 12, and 8 h period lengths, respectively.
Mentions: These tests were corrected for multiple comparisons post hoc using the method described by Storey and colleagues[27],[28]. Briefly, by examining the distribution of p-values from a given data set, an estimate of the proportion that are truly non-rhythmic can be derived. Using this approach to model the rate of false-discoveries, the p-value for each transcript, which estimates the frequency that a truly observation will be labeled as significant, can be converted to a more stringent q-value which instead estimates the frequency that significant observations are truly non-rhythmic. At a false discovery rate [27] of <0.05, over 3000 transcripts were found to oscillate by both statistical tests in liver, while fewer than a dozen were found in NIH3T3 and U2OS cells. As expected, the majority of cycling transcripts from liver (and all from NIH3T3 and U2OS cells) had period lengths of approximately 24 h (Figure 1A, Figure 2A and B, Table 1).

Bottom Line: We recently developed a method to look at transcriptional oscillations with unprecedented precision and accuracy using high-density time sampling.Strikingly, we found two clusters of genes that cycle at the second and third harmonic of circadian rhythmicity in liver, but not cultured cells.Validation experiments show that 12-hour oscillatory transcripts occur in several other peripheral tissues as well including heart, kidney, and lungs.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Institute for Translational Medicine and Therapeutics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
The circadian clock is a molecular and cellular oscillator found in most mammalian tissues that regulates rhythmic physiology and behavior. Numerous investigations have addressed the contribution of circadian rhythmicity to cellular, organ, and organismal physiology. We recently developed a method to look at transcriptional oscillations with unprecedented precision and accuracy using high-density time sampling. Here, we report a comparison of oscillating transcription from mouse liver, NIH3T3, and U2OS cells. Several surprising observations resulted from this study, including a 100-fold difference in the number of cycling transcripts in autonomous cellular models of the oscillator versus tissues harvested from intact mice. Strikingly, we found two clusters of genes that cycle at the second and third harmonic of circadian rhythmicity in liver, but not cultured cells. Validation experiments show that 12-hour oscillatory transcripts occur in several other peripheral tissues as well including heart, kidney, and lungs. These harmonics are lost ex vivo, as well as under restricted feeding conditions. Taken in sum, these studies illustrate the importance of time sampling with respect to multiple testing, suggest caution in use of autonomous cellular models to study clock output, and demonstrate the existence of harmonics of circadian gene expression in the mouse.

Show MeSH
Related in: MedlinePlus