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Quantitative Transcription Factor Analysis of Undifferentiated Single Human Embryonic Stem Cells

Stahlberg, Anders; Bengtsson, Martin LU ; Hemberg, Martin and Semb, Henrik LU (2009) In Clinical Chemistry 55(12). p.2162-2170
Abstract
BACKGROUND: Human embryonic stem cells (hESCs) require expression of transcription factor genes POU5F1 (POU class 5 homeobox 1), NANOG (Nanog homeobox), and SOX2 [SRY (sex determining region Y)-box 2] to maintain their capacity for self-renewal and pluripotency. Because of the heterogeneous nature of cell populations, it is desirable to study the gene regulation in single cells. Large and potentially important fluctuations in a few cells cannot be detected at the population scale with microarrays or sequencing technologies. We used single-cell gene expression profiling to study cell heterogeneity in hESCs. METHODS: We collected 47 single hESCs from cell line SA121 manually by glass capillaries and 57 single hESCs from cell line HUES3 by... (More)
BACKGROUND: Human embryonic stem cells (hESCs) require expression of transcription factor genes POU5F1 (POU class 5 homeobox 1), NANOG (Nanog homeobox), and SOX2 [SRY (sex determining region Y)-box 2] to maintain their capacity for self-renewal and pluripotency. Because of the heterogeneous nature of cell populations, it is desirable to study the gene regulation in single cells. Large and potentially important fluctuations in a few cells cannot be detected at the population scale with microarrays or sequencing technologies. We used single-cell gene expression profiling to study cell heterogeneity in hESCs. METHODS: We collected 47 single hESCs from cell line SA121 manually by glass capillaries and 57 single hESCs from cell line HUES3 by flow cytometry. Single hESCs were lysed and reverse-transcribed. Reverse-transcription quantitative real-time PCR was then used to measure the expression POU5F1, NANOG, SOX2, and the inhibitor of DNA binding genes ID1, ID2, and ID3. A quantitative noise model was used to remove measurement noise when pairwise correlations were estimated. RESULTS: The numbers of transcripts per cell varied >100-fold between cells and showed lognormal features. POU5F1 expression positively correlated with ID1 and ID3 expression (P < 0.05) but not with NANOG or SOX2 expression. When we accounted for measurement noise, SOX2 expression was also correlated with ID1, ID2, and NANOG expression (P < 0.05). CONCLUSIONS: We demonstrate an accurate method for transcription profiling of individual hESCs. Cell-to-cell variability is large and is at least partly nonrandom because we observed correlations between core transcription factors. High fluctuations in gene expression may explain why individual cells in a seemingly undifferentiated cell population have different susceptibilities for inductive cues. (C) 2009 American Association for Clinical Chemistry (Less)
Please use this url to cite or link to this publication:
author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Clinical Chemistry
volume
55
issue
12
pages
2162 - 2170
publisher
American Association for Clinical Chemistry
external identifiers
  • wos:000272518500017
  • scopus:71749084088
ISSN
0009-9147
DOI
10.1373/clinchem.2009.131433
language
English
LU publication?
yes
id
00c36c41-a7ef-47b8-80f0-3eda18bac2e6 (old id 1532739)
date added to LUP
2010-01-28 17:13:21
date last changed
2017-08-20 03:43:18
@article{00c36c41-a7ef-47b8-80f0-3eda18bac2e6,
  abstract     = {BACKGROUND: Human embryonic stem cells (hESCs) require expression of transcription factor genes POU5F1 (POU class 5 homeobox 1), NANOG (Nanog homeobox), and SOX2 [SRY (sex determining region Y)-box 2] to maintain their capacity for self-renewal and pluripotency. Because of the heterogeneous nature of cell populations, it is desirable to study the gene regulation in single cells. Large and potentially important fluctuations in a few cells cannot be detected at the population scale with microarrays or sequencing technologies. We used single-cell gene expression profiling to study cell heterogeneity in hESCs. METHODS: We collected 47 single hESCs from cell line SA121 manually by glass capillaries and 57 single hESCs from cell line HUES3 by flow cytometry. Single hESCs were lysed and reverse-transcribed. Reverse-transcription quantitative real-time PCR was then used to measure the expression POU5F1, NANOG, SOX2, and the inhibitor of DNA binding genes ID1, ID2, and ID3. A quantitative noise model was used to remove measurement noise when pairwise correlations were estimated. RESULTS: The numbers of transcripts per cell varied &gt;100-fold between cells and showed lognormal features. POU5F1 expression positively correlated with ID1 and ID3 expression (P &lt; 0.05) but not with NANOG or SOX2 expression. When we accounted for measurement noise, SOX2 expression was also correlated with ID1, ID2, and NANOG expression (P &lt; 0.05). CONCLUSIONS: We demonstrate an accurate method for transcription profiling of individual hESCs. Cell-to-cell variability is large and is at least partly nonrandom because we observed correlations between core transcription factors. High fluctuations in gene expression may explain why individual cells in a seemingly undifferentiated cell population have different susceptibilities for inductive cues. (C) 2009 American Association for Clinical Chemistry},
  author       = {Stahlberg, Anders and Bengtsson, Martin and Hemberg, Martin and Semb, Henrik},
  issn         = {0009-9147},
  language     = {eng},
  number       = {12},
  pages        = {2162--2170},
  publisher    = {American Association for Clinical Chemistry},
  series       = {Clinical Chemistry},
  title        = {Quantitative Transcription Factor Analysis of Undifferentiated Single Human Embryonic Stem Cells},
  url          = {http://dx.doi.org/10.1373/clinchem.2009.131433},
  volume       = {55},
  year         = {2009},
}