Down regulation of the expression of the p110, p85 and p55 subunits of phosphatidylinositol 3-kinase during colon cancer cell anchorage-independent growth

Laura Roden, A. Gallegos, A.P. Kozikowski, G. Powis

Research output: Contribution to journalArticle

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Abstract

We have found that analogues of phosphatidylinositol modified at the 3'-myo-inositol position prevent phosphorylation by phosphatidylinositol 3-kinase (PI3-kinase) inhibit the anchorage-independent growth of HT-29 human colon adenocarcinoma cells as colonies in soft agarose but have no effect on monolayer growth on a plastic surface. Examination of an incomplete differential display library revealed 3 genes whose expression was increased and 11 genes whose expression was decreased or absent in HT-29 cells growing as colonies compared to monolayer growth. One of the cDNAs corresponding to an mRNA that was expressed only in cells growing as a monolayer was a 126 bp fragment that had 98% identity with a fragment of mRNA for human p55PIK PI3-kinase regulatory subunit. The down regulation of p55PIK gene expression in HT-29 cells growing as colonies was confirmed by RT-PCR using p55PIK-specific oligonucleotide primers. RT-PCR and Northern hybridisation were used to show that the expression of the p110 catalytic subunit and the p85 regulatory subunit of PI3-kinase were also down regulated in HT-29 cells growing as colonies. We measured a decrease of approximately 25% in total PI3-kinase activity of the HT-29 cells grown in soft agarose compared to HT-29 cells grown as monolayer. The results suggest that p110 PI3-kinase activity may be limiting in cells showing anchorage independent-growth which could explain their increased sensitivity, compared to cells in monolayer culture, to inhibitors of PI3-kinase signalling.
Original languageEnglish
Pages (from-to)4171-4176
Number of pages6
JournalAnticancer Research
Volume19
Publication statusPublished - 1999

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Phosphatidylinositol 3-Kinase
HT29 Cells
Colonic Neoplasms
Down-Regulation
Growth
Sepharose
Gene Expression
Polymerase Chain Reaction
Messenger RNA
DNA Primers
Gene Expression Regulation
Inositol
Phosphatidylinositols
Plastics
Libraries
Catalytic Domain
Colon
Adenocarcinoma
Complementary DNA
Phosphorylation

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Down regulation of the expression of the p110, p85 and p55 subunits of phosphatidylinositol 3-kinase during colon cancer cell anchorage-independent growth. / Roden, Laura; Gallegos, A.; Kozikowski, A.P.; Powis, G.

In: Anticancer Research, Vol. 19, 1999, p. 4171-4176.

Research output: Contribution to journalArticle

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abstract = "We have found that analogues of phosphatidylinositol modified at the 3'-myo-inositol position prevent phosphorylation by phosphatidylinositol 3-kinase (PI3-kinase) inhibit the anchorage-independent growth of HT-29 human colon adenocarcinoma cells as colonies in soft agarose but have no effect on monolayer growth on a plastic surface. Examination of an incomplete differential display library revealed 3 genes whose expression was increased and 11 genes whose expression was decreased or absent in HT-29 cells growing as colonies compared to monolayer growth. One of the cDNAs corresponding to an mRNA that was expressed only in cells growing as a monolayer was a 126 bp fragment that had 98{\%} identity with a fragment of mRNA for human p55PIK PI3-kinase regulatory subunit. The down regulation of p55PIK gene expression in HT-29 cells growing as colonies was confirmed by RT-PCR using p55PIK-specific oligonucleotide primers. RT-PCR and Northern hybridisation were used to show that the expression of the p110 catalytic subunit and the p85 regulatory subunit of PI3-kinase were also down regulated in HT-29 cells growing as colonies. We measured a decrease of approximately 25{\%} in total PI3-kinase activity of the HT-29 cells grown in soft agarose compared to HT-29 cells grown as monolayer. The results suggest that p110 PI3-kinase activity may be limiting in cells showing anchorage independent-growth which could explain their increased sensitivity, compared to cells in monolayer culture, to inhibitors of PI3-kinase signalling.",
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