Quantitative Transcription Factor Analysis of Undifferentiated Single Human Embryonic Stem Cells

Forskningsoutput: TidskriftsbidragArtikel i vetenskaplig tidskrift

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 >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

Detaljer

Författare
  • Anders Stahlberg
  • Martin Bengtsson
  • Martin Hemberg
  • Henrik Semb
Enheter & grupper
Forskningsområden

Ämnesklassifikation (UKÄ) – OBLIGATORISK

  • Biokemi och molekylärbiologi
Originalspråkengelska
Sidor (från-till)2162-2170
TidskriftClinical Chemistry
Volym55
Utgåva nummer12
StatusPublished - 2009
PublikationskategoriForskning
Peer review utfördJa