Abstract
Cells respond to their environment via an intricate cellular signaling network, directing cell fate. Changes in cell fate are characterized by changes in gene transcription, dictated by (master) transcription factor activity. SOX9 is the master transcription factor for chondrocyte development. Its impaired function is implicated in osteoarthritis and growth disorders, such as dwarfism. However, the factors regulating SOX9 transcriptional activity are not yet fully mapped. Current methods to study transcription factor activity are indirect and largely limited to quantification of SOX9 target gene and protein expression levels after several hours or days of stimulation, leading to poor temporal resolution. We used Fluorescence Recovery After Photobleaching (FRAP) to study the mobility of SOX9 and correlated the changes in mobility to changes in its transcriptional activity by cross-validating with chromatin immunoprecipitation and qPCR. We show that using FRAP, we can quantify the changes in SOX9 mobility on short time scales as an indication of transcriptional activity, which correlated to changes of SOX9 DNA-binding and long-term target gene expression.
| Original language | English |
|---|---|
| Pages (from-to) | 107-117 |
| Number of pages | 11 |
| Journal | Biochimica et Biophysica Acta - Gene Regulatory Mechanisms |
| Volume | 1862 |
| Issue number | 1 |
| Early online date | 20 Nov 2018 |
| DOIs | |
| Publication status | Published - Jan 2019 |
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Keywords
- FRAP
- Osteoarthritis
- SOX9
- Transcription factor dynamics
- Transcriptional activity
- Cartilage
Cite this
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Changes in Fluorescence Recovery After Photobleaching (FRAP) as an indicator of SOX9 transcription factor activity. / Govindaraj, Kannan; Hendriks, Jan; Lidke, Diane S.; Karperien, Marcel; Post, Janine N. (Corresponding Author).
In: Biochimica et Biophysica Acta - Gene Regulatory Mechanisms, Vol. 1862, No. 1, 01.2019, p. 107-117.Research output: Contribution to journal › Article › Academic › peer-review
TY - JOUR
T1 - Changes in Fluorescence Recovery After Photobleaching (FRAP) as an indicator of SOX9 transcription factor activity
AU - Govindaraj, Kannan
AU - Hendriks, Jan
AU - Lidke, Diane S.
AU - Karperien, Marcel
AU - Post, Janine N.
PY - 2019/1
Y1 - 2019/1
N2 - Cells respond to their environment via an intricate cellular signaling network, directing cell fate. Changes in cell fate are characterized by changes in gene transcription, dictated by (master) transcription factor activity. SOX9 is the master transcription factor for chondrocyte development. Its impaired function is implicated in osteoarthritis and growth disorders, such as dwarfism. However, the factors regulating SOX9 transcriptional activity are not yet fully mapped. Current methods to study transcription factor activity are indirect and largely limited to quantification of SOX9 target gene and protein expression levels after several hours or days of stimulation, leading to poor temporal resolution. We used Fluorescence Recovery After Photobleaching (FRAP) to study the mobility of SOX9 and correlated the changes in mobility to changes in its transcriptional activity by cross-validating with chromatin immunoprecipitation and qPCR. We show that using FRAP, we can quantify the changes in SOX9 mobility on short time scales as an indication of transcriptional activity, which correlated to changes of SOX9 DNA-binding and long-term target gene expression.
AB - Cells respond to their environment via an intricate cellular signaling network, directing cell fate. Changes in cell fate are characterized by changes in gene transcription, dictated by (master) transcription factor activity. SOX9 is the master transcription factor for chondrocyte development. Its impaired function is implicated in osteoarthritis and growth disorders, such as dwarfism. However, the factors regulating SOX9 transcriptional activity are not yet fully mapped. Current methods to study transcription factor activity are indirect and largely limited to quantification of SOX9 target gene and protein expression levels after several hours or days of stimulation, leading to poor temporal resolution. We used Fluorescence Recovery After Photobleaching (FRAP) to study the mobility of SOX9 and correlated the changes in mobility to changes in its transcriptional activity by cross-validating with chromatin immunoprecipitation and qPCR. We show that using FRAP, we can quantify the changes in SOX9 mobility on short time scales as an indication of transcriptional activity, which correlated to changes of SOX9 DNA-binding and long-term target gene expression.
KW - FRAP
KW - Osteoarthritis
KW - SOX9
KW - Transcription factor dynamics
KW - Transcriptional activity
KW - Cartilage
UR - http://www.scopus.com/inward/record.url?scp=85058506070&partnerID=8YFLogxK
U2 - 10.1016/j.bbagrm.2018.11.001
DO - 10.1016/j.bbagrm.2018.11.001
M3 - Article
C2 - 30465885
AN - SCOPUS:85058506070
VL - 1862
SP - 107
EP - 117
JO - Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
JF - Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
SN - 1874-9399
IS - 1
ER -