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Class II phosphoinositide 3-kinases contribute to endothelial cells morphogenesis.

Tibolla G, Piñeiro R, Chiozzotto D, Mavrommati I, Wheeler AP, Norata GD, Catapano AL, Maffucci T, Falasca M - PLoS ONE (2013)

Bottom Line: In this study we investigated the relative contribution of distinct PI3K isoforms to endothelial cells (EC) functions specifically regulated by the sphingolipid sphingosine-1-phosphate (S1P) and by high density lipoproteins (HDL), the major carrier of S1P in human plasma.Here we show that a co-ordinated action of different PI3Ks is required to tightly regulate remodelling of EC on Matrigel, a process dependent on cell proliferation, apoptosis and migration.The contribution of each isoform to this process appears to be distinct, with the class II enzyme PI3K-C2β and the class IB isoform p110γ mainly regulating the S1P- and HDL-dependent EC migration and PI3K-C2α primarily controlling EC survival.

View Article: PubMed Central - PubMed

Affiliation: Queen Mary University of London, Barts and The London School of Medicine and Dentistry, Blizard Institute, Centre for Diabetes, Inositide Signalling Group, London, United Kingdom.

ABSTRACT
The question of whether the distinct isoforms of the family of enzymes phosphoinositide 3-kinases (PI3Ks) play redundant roles within a cell or whether they control distinct cellular processes or distinct steps within the same cellular process has gained considerable importance in the recent years due to the development of inhibitors able to selectively target individual isoforms. It is important to understand whether inhibition of one PI3K can result in compensatory effect from other isoform(s) and therefore whether strategies aimed at simultaneously blocking more than one PI3K may be needed. In this study we investigated the relative contribution of distinct PI3K isoforms to endothelial cells (EC) functions specifically regulated by the sphingolipid sphingosine-1-phosphate (S1P) and by high density lipoproteins (HDL), the major carrier of S1P in human plasma. Here we show that a co-ordinated action of different PI3Ks is required to tightly regulate remodelling of EC on Matrigel, a process dependent on cell proliferation, apoptosis and migration. The contribution of each isoform to this process appears to be distinct, with the class II enzyme PI3K-C2β and the class IB isoform p110γ mainly regulating the S1P- and HDL-dependent EC migration and PI3K-C2α primarily controlling EC survival. Data further indicate that PI3K-C2β and p110γ control distinct steps involved in cell migration supporting the hypothesis that different PI3Ks regulate distinct cellular processes.

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PI3K-C2β and p110γ play distinct role in regulation of S1P-dependent cell migration.Random motility of HUVEC transfected with the indicated siRNAs was monitored as described in the Materials and Methods section. (A) Representative track paths throughout time of 6 cells are shown (8 h). (B) Data indicate the mean velocity/min and are means ± SEM from 3 independent experiments. **p<0.01 vs cells transfected with scrambled siRNA. (C) Results from Transwell assays performed in HUVEC transfected with the indicated siRNAs. Data are expressed as percentage of migration of cells transfected with scrambled siRNA and unstimulated (control) and are means ± SEM from 3 independent experiments. *p<0.01. Downregulation of PI3K-C2β and p110γ was confirmed by Western blotting.
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pone-0053808-g005: PI3K-C2β and p110γ play distinct role in regulation of S1P-dependent cell migration.Random motility of HUVEC transfected with the indicated siRNAs was monitored as described in the Materials and Methods section. (A) Representative track paths throughout time of 6 cells are shown (8 h). (B) Data indicate the mean velocity/min and are means ± SEM from 3 independent experiments. **p<0.01 vs cells transfected with scrambled siRNA. (C) Results from Transwell assays performed in HUVEC transfected with the indicated siRNAs. Data are expressed as percentage of migration of cells transfected with scrambled siRNA and unstimulated (control) and are means ± SEM from 3 independent experiments. *p<0.01. Downregulation of PI3K-C2β and p110γ was confirmed by Western blotting.

Mentions: Our data so far indicate that three different PI3K isoforms participate in EC morphogenesis. Remodelling of EC on Matrigel is the result of several cellular functions, including migration, proliferation and survival. We therefore decided to investigate in more details the specific contribution of each enzyme to this process. Our data have shown that p110γ and PI3K-C2β are mainly involved in both S1P- and HDL3-induced cell migration. To gain further insight into the specific role of each isofom in regulation of cell migration we performed random motility assays. Specifically, we measured the mean velocity of cells in serum by tracking individual HUVEC transfected with siRNAs targeting each distinct PI3K or with the control scrambled siRNA. Representative track paths are shown in Figure 5A. A significant reduction of the mean velocity was detected in cells upon p110γ but not PI3K-C2β downregulation (Figure 5B), suggesting that the two PI3Ks may control different steps of cell migration.


Class II phosphoinositide 3-kinases contribute to endothelial cells morphogenesis.

Tibolla G, Piñeiro R, Chiozzotto D, Mavrommati I, Wheeler AP, Norata GD, Catapano AL, Maffucci T, Falasca M - PLoS ONE (2013)

PI3K-C2β and p110γ play distinct role in regulation of S1P-dependent cell migration.Random motility of HUVEC transfected with the indicated siRNAs was monitored as described in the Materials and Methods section. (A) Representative track paths throughout time of 6 cells are shown (8 h). (B) Data indicate the mean velocity/min and are means ± SEM from 3 independent experiments. **p<0.01 vs cells transfected with scrambled siRNA. (C) Results from Transwell assays performed in HUVEC transfected with the indicated siRNAs. Data are expressed as percentage of migration of cells transfected with scrambled siRNA and unstimulated (control) and are means ± SEM from 3 independent experiments. *p<0.01. Downregulation of PI3K-C2β and p110γ was confirmed by Western blotting.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3539993&req=5

pone-0053808-g005: PI3K-C2β and p110γ play distinct role in regulation of S1P-dependent cell migration.Random motility of HUVEC transfected with the indicated siRNAs was monitored as described in the Materials and Methods section. (A) Representative track paths throughout time of 6 cells are shown (8 h). (B) Data indicate the mean velocity/min and are means ± SEM from 3 independent experiments. **p<0.01 vs cells transfected with scrambled siRNA. (C) Results from Transwell assays performed in HUVEC transfected with the indicated siRNAs. Data are expressed as percentage of migration of cells transfected with scrambled siRNA and unstimulated (control) and are means ± SEM from 3 independent experiments. *p<0.01. Downregulation of PI3K-C2β and p110γ was confirmed by Western blotting.
Mentions: Our data so far indicate that three different PI3K isoforms participate in EC morphogenesis. Remodelling of EC on Matrigel is the result of several cellular functions, including migration, proliferation and survival. We therefore decided to investigate in more details the specific contribution of each enzyme to this process. Our data have shown that p110γ and PI3K-C2β are mainly involved in both S1P- and HDL3-induced cell migration. To gain further insight into the specific role of each isofom in regulation of cell migration we performed random motility assays. Specifically, we measured the mean velocity of cells in serum by tracking individual HUVEC transfected with siRNAs targeting each distinct PI3K or with the control scrambled siRNA. Representative track paths are shown in Figure 5A. A significant reduction of the mean velocity was detected in cells upon p110γ but not PI3K-C2β downregulation (Figure 5B), suggesting that the two PI3Ks may control different steps of cell migration.

Bottom Line: In this study we investigated the relative contribution of distinct PI3K isoforms to endothelial cells (EC) functions specifically regulated by the sphingolipid sphingosine-1-phosphate (S1P) and by high density lipoproteins (HDL), the major carrier of S1P in human plasma.Here we show that a co-ordinated action of different PI3Ks is required to tightly regulate remodelling of EC on Matrigel, a process dependent on cell proliferation, apoptosis and migration.The contribution of each isoform to this process appears to be distinct, with the class II enzyme PI3K-C2β and the class IB isoform p110γ mainly regulating the S1P- and HDL-dependent EC migration and PI3K-C2α primarily controlling EC survival.

View Article: PubMed Central - PubMed

Affiliation: Queen Mary University of London, Barts and The London School of Medicine and Dentistry, Blizard Institute, Centre for Diabetes, Inositide Signalling Group, London, United Kingdom.

ABSTRACT
The question of whether the distinct isoforms of the family of enzymes phosphoinositide 3-kinases (PI3Ks) play redundant roles within a cell or whether they control distinct cellular processes or distinct steps within the same cellular process has gained considerable importance in the recent years due to the development of inhibitors able to selectively target individual isoforms. It is important to understand whether inhibition of one PI3K can result in compensatory effect from other isoform(s) and therefore whether strategies aimed at simultaneously blocking more than one PI3K may be needed. In this study we investigated the relative contribution of distinct PI3K isoforms to endothelial cells (EC) functions specifically regulated by the sphingolipid sphingosine-1-phosphate (S1P) and by high density lipoproteins (HDL), the major carrier of S1P in human plasma. Here we show that a co-ordinated action of different PI3Ks is required to tightly regulate remodelling of EC on Matrigel, a process dependent on cell proliferation, apoptosis and migration. The contribution of each isoform to this process appears to be distinct, with the class II enzyme PI3K-C2β and the class IB isoform p110γ mainly regulating the S1P- and HDL-dependent EC migration and PI3K-C2α primarily controlling EC survival. Data further indicate that PI3K-C2β and p110γ control distinct steps involved in cell migration supporting the hypothesis that different PI3Ks regulate distinct cellular processes.

Show MeSH
Related in: MedlinePlus