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Suppression of the PI3K subunit p85α delays embryoid body development and inhibits cell adhesion
Author(s) -
Gurney Susan M.R.,
Forster Peter,
Just Ursula,
Schwanbeck Ralf
Publication year - 2011
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.23285
Subject(s) - embryoid body , microbiology and biotechnology , protein subunit , chemistry , pi3k/akt/mtor pathway , cell growth , cell adhesion , integrin , cell , biology , signal transduction , cellular differentiation , biochemistry , adult stem cell , gene
Phosphatidylinositol‐3‐kinases (PI3Ks) exert a variety of signaling functions in eukaryotes. We suppressed the PI3K regulatory subunit p85α using a small interfering RNA ( Pik3r1 siRNA) and examined the effects on embryoid body (EB) development in hanging drop culture. We observed a 150% increase in the volume of the treated EBs within 24 h, compared to the negative controls. Fluorescence Activated Cell Sorting (FACS) assays showed that this increase in volume is not due to increased cellular proliferation. Instead, the increase in volume appears to be due to reduced cellular aggregation and adherence. This is further shown by our observation that 40% of treated EBs form twin instead of single EBs, and that they have a significantly reduced ability to adhere to culture dishes when plated. A time course over the first 96 h reveals that the impaired adherence is transient and explained by an initial 12‐hour delay in EB development. Quantitative PCR expression analysis suggests that the adhesion molecule integrin‐β1 (ITGB1) is transiently downregulated by the p85α suppression. In conclusion we found that suppressing p85α leads to a delay in forming compact EBs, accompanied by a transient inability of the EBs to undergo normal cell–cell and cell–substrate adhesion. J. Cell. Biochem. 112: 3573–3581, 2011. © 2011 Wiley Periodicals, Inc.