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PIP₃-dependent macropinocytosis is incompatible with chemotaxis.

Veltman DM, Lemieux MG, Knecht DA, Insall RH - J. Cell Biol. (2014)

Bottom Line: In eukaryotic chemotaxis, the mechanisms connecting external signals to the motile apparatus remain unclear.Wild-type cells, unlike the widely used axenic mutants, show little macropinocytosis and few large PIP₃ patches, but migrate more efficiently toward folate.Tellingly, folate chemotaxis in axenic cells is rescued by knocking out phosphatidylinositide 3-kinases (PI 3-kinases).

View Article: PubMed Central - HTML - PubMed

Affiliation: Beatson Institute for Cancer Research, Glasgow G61 1BD, Scotland, UK.

ABSTRACT
In eukaryotic chemotaxis, the mechanisms connecting external signals to the motile apparatus remain unclear. The role of the lipid phosphatidylinositol 3,4,5-trisphosphate (PIP₃) has been particularly controversial. PIP₃ has many cellular roles, notably in growth control and macropinocytosis as well as cell motility. Here we show that PIP₃ is not only unnecessary for Dictyostelium discoideum to migrate toward folate, but actively inhibits chemotaxis. We find that macropinosomes, but not pseudopods, in growing cells are dependent on PIP₃. PIP₃ patches in these cells show no directional bias, and overall only PIP₃-free pseudopods orient up-gradient. The pseudopod driver suppressor of cAR mutations (SCAR)/WASP and verprolin homologue (WAVE) is not recruited to the center of PIP₃ patches, just the edges, where it causes macropinosome formation. Wild-type cells, unlike the widely used axenic mutants, show little macropinocytosis and few large PIP₃ patches, but migrate more efficiently toward folate. Tellingly, folate chemotaxis in axenic cells is rescued by knocking out phosphatidylinositide 3-kinases (PI 3-kinases). Thus PIP₃ promotes macropinocytosis and interferes with pseudopod orientation during chemotaxis of growing cells.

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PIP3-induced macropinocytosis is incompatible with chemotaxis. (A) Negative correlation of macropinocytosis and chemotaxis. The rate of macropinosome formation under different conditions was measured using FITC-dextran and fluorescence microscopy. Each data point represents a single cell that was imaged for at least 5 min. Data were taken from at least two independent experiments. Chemotaxis is represented qualitatively to avoid quantitative comparisons made under noncomparable conditions. (B) Deletion of PI 3-kinases from axenically growing cells restores chemotaxis to folate. (B, right) Insall chamber chemotaxis of axenic AX2 cells quintuply deleted for all p110 PI 3-kinases. (B, left) Chemotaxis of axenic wild-type cells (see data from Fig. 1 C for comparison).
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fig5: PIP3-induced macropinocytosis is incompatible with chemotaxis. (A) Negative correlation of macropinocytosis and chemotaxis. The rate of macropinosome formation under different conditions was measured using FITC-dextran and fluorescence microscopy. Each data point represents a single cell that was imaged for at least 5 min. Data were taken from at least two independent experiments. Chemotaxis is represented qualitatively to avoid quantitative comparisons made under noncomparable conditions. (B) Deletion of PI 3-kinases from axenically growing cells restores chemotaxis to folate. (B, right) Insall chamber chemotaxis of axenic AX2 cells quintuply deleted for all p110 PI 3-kinases. (B, left) Chemotaxis of axenic wild-type cells (see data from Fig. 1 C for comparison).

Mentions: We assessed the relationship between macropinocytosis and chemotaxis by altering the macropinocytic rate. Axenically grown AX2 cells form FITC-labeled vesicles at a rate of 0.44/min (Fig. 5 A). However, AX2 cells that have been cultivated on bacteria have a diminished rate of macropinocytosis. Macropinosomes in NC4 cells are even rarer, with a mean rate of 0.05/min. These changes correlate with the efficiency of chemotaxis toward folate (Fig. 5 A).


PIP₃-dependent macropinocytosis is incompatible with chemotaxis.

Veltman DM, Lemieux MG, Knecht DA, Insall RH - J. Cell Biol. (2014)

PIP3-induced macropinocytosis is incompatible with chemotaxis. (A) Negative correlation of macropinocytosis and chemotaxis. The rate of macropinosome formation under different conditions was measured using FITC-dextran and fluorescence microscopy. Each data point represents a single cell that was imaged for at least 5 min. Data were taken from at least two independent experiments. Chemotaxis is represented qualitatively to avoid quantitative comparisons made under noncomparable conditions. (B) Deletion of PI 3-kinases from axenically growing cells restores chemotaxis to folate. (B, right) Insall chamber chemotaxis of axenic AX2 cells quintuply deleted for all p110 PI 3-kinases. (B, left) Chemotaxis of axenic wild-type cells (see data from Fig. 1 C for comparison).
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3926956&req=5

fig5: PIP3-induced macropinocytosis is incompatible with chemotaxis. (A) Negative correlation of macropinocytosis and chemotaxis. The rate of macropinosome formation under different conditions was measured using FITC-dextran and fluorescence microscopy. Each data point represents a single cell that was imaged for at least 5 min. Data were taken from at least two independent experiments. Chemotaxis is represented qualitatively to avoid quantitative comparisons made under noncomparable conditions. (B) Deletion of PI 3-kinases from axenically growing cells restores chemotaxis to folate. (B, right) Insall chamber chemotaxis of axenic AX2 cells quintuply deleted for all p110 PI 3-kinases. (B, left) Chemotaxis of axenic wild-type cells (see data from Fig. 1 C for comparison).
Mentions: We assessed the relationship between macropinocytosis and chemotaxis by altering the macropinocytic rate. Axenically grown AX2 cells form FITC-labeled vesicles at a rate of 0.44/min (Fig. 5 A). However, AX2 cells that have been cultivated on bacteria have a diminished rate of macropinocytosis. Macropinosomes in NC4 cells are even rarer, with a mean rate of 0.05/min. These changes correlate with the efficiency of chemotaxis toward folate (Fig. 5 A).

Bottom Line: In eukaryotic chemotaxis, the mechanisms connecting external signals to the motile apparatus remain unclear.Wild-type cells, unlike the widely used axenic mutants, show little macropinocytosis and few large PIP₃ patches, but migrate more efficiently toward folate.Tellingly, folate chemotaxis in axenic cells is rescued by knocking out phosphatidylinositide 3-kinases (PI 3-kinases).

View Article: PubMed Central - HTML - PubMed

Affiliation: Beatson Institute for Cancer Research, Glasgow G61 1BD, Scotland, UK.

ABSTRACT
In eukaryotic chemotaxis, the mechanisms connecting external signals to the motile apparatus remain unclear. The role of the lipid phosphatidylinositol 3,4,5-trisphosphate (PIP₃) has been particularly controversial. PIP₃ has many cellular roles, notably in growth control and macropinocytosis as well as cell motility. Here we show that PIP₃ is not only unnecessary for Dictyostelium discoideum to migrate toward folate, but actively inhibits chemotaxis. We find that macropinosomes, but not pseudopods, in growing cells are dependent on PIP₃. PIP₃ patches in these cells show no directional bias, and overall only PIP₃-free pseudopods orient up-gradient. The pseudopod driver suppressor of cAR mutations (SCAR)/WASP and verprolin homologue (WAVE) is not recruited to the center of PIP₃ patches, just the edges, where it causes macropinosome formation. Wild-type cells, unlike the widely used axenic mutants, show little macropinocytosis and few large PIP₃ patches, but migrate more efficiently toward folate. Tellingly, folate chemotaxis in axenic cells is rescued by knocking out phosphatidylinositide 3-kinases (PI 3-kinases). Thus PIP₃ promotes macropinocytosis and interferes with pseudopod orientation during chemotaxis of growing cells.

Show MeSH
Related in: MedlinePlus