The circadian clock modulates enamel development

Rodrigo Lacruz, Joseph G. Hacia, Timothy Bromage, Alan Boyde, Yaping Lei, Yucheng Xu, Joseph D. Miller, Michael L. Paine, Malcolm L. Snead

Research output: Contribution to journalArticle

Abstract

Fully mature enamel is about 98% mineral by weight. While mineral crystals appear very early during its formative phase, the newly secreted enamel is a soft gel-like matrix containing several enamel matrix proteins of which the most abundant is amelogenin (Amelx). Histological analysis of mineralized dental enamel reveals markings called cross-striations associated with daily increments of enamel formation, as evidenced by injections of labeling dyes at known time intervals. The daily incremental growth of enamel has led to the hypothesis that the circadian clock might be involved in the regulation of enamel development. To identify daily rhythms of clock genes and Amelx, we subjected murine ameloblast cells to serum synchronization to analyze the expression of the circadian transcription factors Per2 and Bmal1 by real-time PCR. Results indicate that these key genetic regulators of the circadian clock are expressed in synchronized murine ameloblast cell cultures and that their expression profile follows a circadian pattern with acrophase and bathyphase for both gene transcripts in antiphase. Immunohistological analysis confirms the protein expression of Bmal and Cry in enamel cells. Amelx expression in 2-day postnatal mouse molars dissected every 4 hours for a duration of 48 hours oscillated with an approximately 24-hour period, with a significant approximately 2-fold decrease in expression during the dark period compared to the light period. The expression of genes involved in bicarbonate production (Car2) and transport (Slc4a4), as well as in enamel matrix endocytosis (Lamp1), was greater during the dark period, indicating that ameloblasts express these proteins when Amelx expression is at the nadir. The human and mouse Amelx genes each contain a single nonconserved E-box element within 10 kb upstream of their respective transcription start sites. We also found that within 2 kb of the transcription start site of the human NFYA gene, which encodes a positive regulator of amelogenin, there is an E-box element that is conserved in rodents and other mammals. Moreover, we found that Nfya expression in serum-synchronized murine ameloblasts oscillated with a strong 24-hour rhythm. Taken together, our data support the hypothesis that the circadian clock temporally regulates enamel development.

Original languageEnglish (US)
Pages (from-to)237-245
Number of pages9
JournalJournal of Biological Rhythms
Volume27
Issue number3
DOIs
StatePublished - Jun 2012

Fingerprint

Circadian Clocks
Dental Enamel
Ameloblasts
E-Box Elements
Amelogenin
Transcription Initiation Site
Genes
Minerals
Bicarbonates
Endocytosis
Serum
Real-Time Polymerase Chain Reaction
Mammals
Rodentia
Proteins
Transcription Factors
Coloring Agents
Cell Culture Techniques
Gels
Gene Expression

Keywords

  • ameloblast cells
  • amelogenin
  • circadian rhythms
  • enamel development

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

Cite this

The circadian clock modulates enamel development. / Lacruz, Rodrigo; Hacia, Joseph G.; Bromage, Timothy; Boyde, Alan; Lei, Yaping; Xu, Yucheng; Miller, Joseph D.; Paine, Michael L.; Snead, Malcolm L.

In: Journal of Biological Rhythms, Vol. 27, No. 3, 06.2012, p. 237-245.

Research output: Contribution to journalArticle

Lacruz, R, Hacia, JG, Bromage, T, Boyde, A, Lei, Y, Xu, Y, Miller, JD, Paine, ML & Snead, ML 2012, 'The circadian clock modulates enamel development', Journal of Biological Rhythms, vol. 27, no. 3, pp. 237-245. https://doi.org/10.1177/0748730412442830
Lacruz, Rodrigo ; Hacia, Joseph G. ; Bromage, Timothy ; Boyde, Alan ; Lei, Yaping ; Xu, Yucheng ; Miller, Joseph D. ; Paine, Michael L. ; Snead, Malcolm L. / The circadian clock modulates enamel development. In: Journal of Biological Rhythms. 2012 ; Vol. 27, No. 3. pp. 237-245.
@article{5d829766f55940eeb383ef03d548d07b,
title = "The circadian clock modulates enamel development",
abstract = "Fully mature enamel is about 98{\%} mineral by weight. While mineral crystals appear very early during its formative phase, the newly secreted enamel is a soft gel-like matrix containing several enamel matrix proteins of which the most abundant is amelogenin (Amelx). Histological analysis of mineralized dental enamel reveals markings called cross-striations associated with daily increments of enamel formation, as evidenced by injections of labeling dyes at known time intervals. The daily incremental growth of enamel has led to the hypothesis that the circadian clock might be involved in the regulation of enamel development. To identify daily rhythms of clock genes and Amelx, we subjected murine ameloblast cells to serum synchronization to analyze the expression of the circadian transcription factors Per2 and Bmal1 by real-time PCR. Results indicate that these key genetic regulators of the circadian clock are expressed in synchronized murine ameloblast cell cultures and that their expression profile follows a circadian pattern with acrophase and bathyphase for both gene transcripts in antiphase. Immunohistological analysis confirms the protein expression of Bmal and Cry in enamel cells. Amelx expression in 2-day postnatal mouse molars dissected every 4 hours for a duration of 48 hours oscillated with an approximately 24-hour period, with a significant approximately 2-fold decrease in expression during the dark period compared to the light period. The expression of genes involved in bicarbonate production (Car2) and transport (Slc4a4), as well as in enamel matrix endocytosis (Lamp1), was greater during the dark period, indicating that ameloblasts express these proteins when Amelx expression is at the nadir. The human and mouse Amelx genes each contain a single nonconserved E-box element within 10 kb upstream of their respective transcription start sites. We also found that within 2 kb of the transcription start site of the human NFYA gene, which encodes a positive regulator of amelogenin, there is an E-box element that is conserved in rodents and other mammals. Moreover, we found that Nfya expression in serum-synchronized murine ameloblasts oscillated with a strong 24-hour rhythm. Taken together, our data support the hypothesis that the circadian clock temporally regulates enamel development.",
keywords = "ameloblast cells, amelogenin, circadian rhythms, enamel development",
author = "Rodrigo Lacruz and Hacia, {Joseph G.} and Timothy Bromage and Alan Boyde and Yaping Lei and Yucheng Xu and Miller, {Joseph D.} and Paine, {Michael L.} and Snead, {Malcolm L.}",
year = "2012",
month = "6",
doi = "10.1177/0748730412442830",
language = "English (US)",
volume = "27",
pages = "237--245",
journal = "Journal of Biological Rhythms",
issn = "0748-7304",
publisher = "SAGE Publications Inc.",
number = "3",

}

TY - JOUR

T1 - The circadian clock modulates enamel development

AU - Lacruz, Rodrigo

AU - Hacia, Joseph G.

AU - Bromage, Timothy

AU - Boyde, Alan

AU - Lei, Yaping

AU - Xu, Yucheng

AU - Miller, Joseph D.

AU - Paine, Michael L.

AU - Snead, Malcolm L.

PY - 2012/6

Y1 - 2012/6

N2 - Fully mature enamel is about 98% mineral by weight. While mineral crystals appear very early during its formative phase, the newly secreted enamel is a soft gel-like matrix containing several enamel matrix proteins of which the most abundant is amelogenin (Amelx). Histological analysis of mineralized dental enamel reveals markings called cross-striations associated with daily increments of enamel formation, as evidenced by injections of labeling dyes at known time intervals. The daily incremental growth of enamel has led to the hypothesis that the circadian clock might be involved in the regulation of enamel development. To identify daily rhythms of clock genes and Amelx, we subjected murine ameloblast cells to serum synchronization to analyze the expression of the circadian transcription factors Per2 and Bmal1 by real-time PCR. Results indicate that these key genetic regulators of the circadian clock are expressed in synchronized murine ameloblast cell cultures and that their expression profile follows a circadian pattern with acrophase and bathyphase for both gene transcripts in antiphase. Immunohistological analysis confirms the protein expression of Bmal and Cry in enamel cells. Amelx expression in 2-day postnatal mouse molars dissected every 4 hours for a duration of 48 hours oscillated with an approximately 24-hour period, with a significant approximately 2-fold decrease in expression during the dark period compared to the light period. The expression of genes involved in bicarbonate production (Car2) and transport (Slc4a4), as well as in enamel matrix endocytosis (Lamp1), was greater during the dark period, indicating that ameloblasts express these proteins when Amelx expression is at the nadir. The human and mouse Amelx genes each contain a single nonconserved E-box element within 10 kb upstream of their respective transcription start sites. We also found that within 2 kb of the transcription start site of the human NFYA gene, which encodes a positive regulator of amelogenin, there is an E-box element that is conserved in rodents and other mammals. Moreover, we found that Nfya expression in serum-synchronized murine ameloblasts oscillated with a strong 24-hour rhythm. Taken together, our data support the hypothesis that the circadian clock temporally regulates enamel development.

AB - Fully mature enamel is about 98% mineral by weight. While mineral crystals appear very early during its formative phase, the newly secreted enamel is a soft gel-like matrix containing several enamel matrix proteins of which the most abundant is amelogenin (Amelx). Histological analysis of mineralized dental enamel reveals markings called cross-striations associated with daily increments of enamel formation, as evidenced by injections of labeling dyes at known time intervals. The daily incremental growth of enamel has led to the hypothesis that the circadian clock might be involved in the regulation of enamel development. To identify daily rhythms of clock genes and Amelx, we subjected murine ameloblast cells to serum synchronization to analyze the expression of the circadian transcription factors Per2 and Bmal1 by real-time PCR. Results indicate that these key genetic regulators of the circadian clock are expressed in synchronized murine ameloblast cell cultures and that their expression profile follows a circadian pattern with acrophase and bathyphase for both gene transcripts in antiphase. Immunohistological analysis confirms the protein expression of Bmal and Cry in enamel cells. Amelx expression in 2-day postnatal mouse molars dissected every 4 hours for a duration of 48 hours oscillated with an approximately 24-hour period, with a significant approximately 2-fold decrease in expression during the dark period compared to the light period. The expression of genes involved in bicarbonate production (Car2) and transport (Slc4a4), as well as in enamel matrix endocytosis (Lamp1), was greater during the dark period, indicating that ameloblasts express these proteins when Amelx expression is at the nadir. The human and mouse Amelx genes each contain a single nonconserved E-box element within 10 kb upstream of their respective transcription start sites. We also found that within 2 kb of the transcription start site of the human NFYA gene, which encodes a positive regulator of amelogenin, there is an E-box element that is conserved in rodents and other mammals. Moreover, we found that Nfya expression in serum-synchronized murine ameloblasts oscillated with a strong 24-hour rhythm. Taken together, our data support the hypothesis that the circadian clock temporally regulates enamel development.

KW - ameloblast cells

KW - amelogenin

KW - circadian rhythms

KW - enamel development

UR - http://www.scopus.com/inward/record.url?scp=84861818163&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84861818163&partnerID=8YFLogxK

U2 - 10.1177/0748730412442830

DO - 10.1177/0748730412442830

M3 - Article

VL - 27

SP - 237

EP - 245

JO - Journal of Biological Rhythms

JF - Journal of Biological Rhythms

SN - 0748-7304

IS - 3

ER -