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Updated: Aug 6, 2026

Combined DNA-RNA Fluorescent In situ Hybridization (FISH) to Study X Chromosome Inactivation in Differentiated Female Mouse Embryonic Stem Cells
Published on: June 14, 2014
Antisense transcription through the Xist locus mediates Tsix function in embryonic stem cells
S Luikenhuis1, A Wutz, R Jaenisch
1Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
Antisense Tsix transcript is crucial for random X inactivation in mammals. Disrupting Tsix transcription leads to nonrandom X chromosome inactivation, establishing its regulatory role.
Area of Science:
- Genetics
- Epigenetics
- Developmental Biology
Background:
- Xist gene expression regulates mammalian X inactivation.
- The antisense Tsix transcript is proposed to control Xist expression.
- Previous studies involving Tsix promoter deletion showed nonrandom X inactivation, but the cause was unclear.
Purpose of the Study:
- To determine the role of Tsix transcription in random X inactivation.
- To investigate if Tsix transcription itself, not just DNA elements, regulates X inactivation.
Main Methods:
- Utilized mouse embryonic stem (ES) cells for modeling X inactivation.
- Modulated Tsix transcription using two distinct approaches: transcriptional inhibition and inducible expression.
- Introduced a transcriptional stop signal to inhibit Tsix transcription.
- Created an inducible system for controlled Tsix expression.
Main Results:
- Truncation of the Tsix transcript resulted in complete nonrandom inactivation of the targeted X chromosome.
- Induction of Tsix expression during ES cell differentiation led to the targeted chromosome being consistently chosen as the active X chromosome.
- Demonstrated a direct correlation between Tsix transcription levels and X inactivation patterns.
Conclusions:
- Established the functional role of antisense transcription in regulating X inactivation.
- Confirmed that Tsix transcription is essential for the random choice of the X chromosome during inactivation.
- Provided definitive evidence for Tsix's regulatory function in X inactivation.
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