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Oct4 distribution and level in mouse clones: consequences for pluripotency
Michele Boiani1, Sigrid Eckardt, Hans R Schöler
1Germline Development Group, Center for Animal Transgenesis and Germ Cell Research, The School of Veterinary Medicine, University of Pennsylvania, New Bolton Center, Kennett Square, Pennsylvania 19348, USA.
Abstract:
Somatic cell clones often fail at a developmental stage coincident with commencement of differentiation. The transcription factor Oct4 is expressed during cleavage stages and is essential for the differentiation of the blastocyst. Oct4 expression becomes restricted to the inner cell mass and epiblast. After gastrulation Oct4 is active only in germ cells and is silent in somatic cells. Here, Oct4 and an Oct4-GFP transgene were used as markers for which gene reprogramming could be directly related to the developmental potential of somatic cell clones. Cumulus cell clones initiated Oct4 expression at the correct stage but showed an incorrect spatial expression in the majority of blastocysts. The ability of clones to form outgrowths was reduced, and the outgrowths had low or even undetectable levels of Oct4 RNA or GFP. The quality of GFP signals in blastocysts correlated with the ability to generate outgrowths that maintain GFP expression and the frequency of embryonic stem (ES) cell derivation. Abnormal Oct4 expression in clones is either directly or indirectly caused by reprogramming errors and is indicative of a general failure to reset the genetic program. The abnormal Oct4 expression may be associated with aberrant expression of other crucial developmental genes, leading to abnormalities at various embryonic stages. Regardless of other genes, the variations observed in Oct4 levels alone account for the majority of failures currently observed for somatic cell cloning.
Insights
Somatic cell cloning failures are linked to abnormal Oct4 gene expression during early development. Correcting Oct4 reprogramming is crucial for successful cloning and developmental potential.
Area of Science:
- Developmental Biology
- Stem Cell Biology
- Reproductive Biology
Background:
- Somatic cell cloning frequently fails during differentiation initiation.
- Oct4 (octamer-binding transcription factor 4) is critical for early embryonic development and pluripotency.
- Oct4 expression patterns change dynamically during normal embryonic development.
Purpose of the Study:
- To investigate the role of Oct4 expression in somatic cell cloning success.
- To correlate Oct4 reprogramming with the developmental potential of somatic cell clones.
- To identify Oct4 expression as a marker for cloning errors.
Main Methods:
- Utilized Oct4 and an Oct4-GFP transgene as markers in cloned cells.
- Analyzed Oct4 expression patterns in blastocysts derived from cumulus cell clones.
- Assessed the ability of clones to form outgrowths and derive embryonic stem (ES) cells.
Main Results:
- Cumulus cell clones initiated Oct4 expression but often showed incorrect spatial patterns in blastocysts.
- Reduced outgrowth formation and low/undetectable Oct4 RNA or GFP levels were observed.
- GFP signal quality correlated with outgrowth maintenance and ES cell derivation frequency.
Conclusions:
- Abnormal Oct4 expression indicates reprogramming errors and failure to reset the genetic program.
- Aberrant Oct4 levels are a primary cause of somatic cell cloning failures.
- Oct4 expression serves as a key indicator of developmental potential in cloned embryos.