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Neurofibromin controls macropinocytosis and phagocytosis in Dictyostelium.

Bloomfield G, Traynor D, Sander SP, Veltman DM, Pachebat JA, Kay RR - Elife (2015)

Bottom Line: Mutants form outsized macropinosomes which are promoted by greater Ras and PI3K activity at sites of endocytosis.An NF1 reporter is recruited to nascent macropinosomes, suggesting that NF1 limits their size by locally inhibiting Ras signalling.Our results link NF1 with macropinocytosis and phagocytosis for the first time, and we propose that NF1 evolved in early phagotrophs to spatially modulate Ras activity, thereby constraining and shaping their feeding structures.

View Article: PubMed Central - PubMed

Affiliation: MRC Laboratory of Molecular Biology, Cambridge, United Kingdom.

ABSTRACT
Cells use phagocytosis and macropinocytosis to internalise bulk material, which in phagotrophic organisms supplies the nutrients necessary for growth. Wildtype Dictyostelium amoebae feed on bacteria, but for decades laboratory work has relied on axenic mutants that can also grow on liquid media. We used forward genetics to identify the causative gene underlying this phenotype. This gene encodes the RasGAP Neurofibromin (NF1). Loss of NF1 enables axenic growth by increasing fluid uptake. Mutants form outsized macropinosomes which are promoted by greater Ras and PI3K activity at sites of endocytosis. Relatedly, NF1 mutants can ingest larger-than-normal particles using phagocytosis. An NF1 reporter is recruited to nascent macropinosomes, suggesting that NF1 limits their size by locally inhibiting Ras signalling. Our results link NF1 with macropinocytosis and phagocytosis for the first time, and we propose that NF1 evolved in early phagotrophs to spatially modulate Ras activity, thereby constraining and shaping their feeding structures.

No MeSH data available.


Related in: MedlinePlus

NF1 mutants grow normally when shaken in suspensions of dead bacteria.To quantify cell growth without complicating factors such as cell motility and susceptibility to harmful bacterial metabolites, cells were grown on heat-killed Escherichia coli strain B/r in shaking suspension. The established axenic strain Ax2 grows consistently more slowly than DdB and HM1591, with a doubling time on average approximately 5% greater. Data are means plus and minus standard error for three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.04940.029
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fig7s2: NF1 mutants grow normally when shaken in suspensions of dead bacteria.To quantify cell growth without complicating factors such as cell motility and susceptibility to harmful bacterial metabolites, cells were grown on heat-killed Escherichia coli strain B/r in shaking suspension. The established axenic strain Ax2 grows consistently more slowly than DdB and HM1591, with a doubling time on average approximately 5% greater. Data are means plus and minus standard error for three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.04940.029


Neurofibromin controls macropinocytosis and phagocytosis in Dictyostelium.

Bloomfield G, Traynor D, Sander SP, Veltman DM, Pachebat JA, Kay RR - Elife (2015)

NF1 mutants grow normally when shaken in suspensions of dead bacteria.To quantify cell growth without complicating factors such as cell motility and susceptibility to harmful bacterial metabolites, cells were grown on heat-killed Escherichia coli strain B/r in shaking suspension. The established axenic strain Ax2 grows consistently more slowly than DdB and HM1591, with a doubling time on average approximately 5% greater. Data are means plus and minus standard error for three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.04940.029
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4374526&req=5

fig7s2: NF1 mutants grow normally when shaken in suspensions of dead bacteria.To quantify cell growth without complicating factors such as cell motility and susceptibility to harmful bacterial metabolites, cells were grown on heat-killed Escherichia coli strain B/r in shaking suspension. The established axenic strain Ax2 grows consistently more slowly than DdB and HM1591, with a doubling time on average approximately 5% greater. Data are means plus and minus standard error for three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.04940.029
Bottom Line: Mutants form outsized macropinosomes which are promoted by greater Ras and PI3K activity at sites of endocytosis.An NF1 reporter is recruited to nascent macropinosomes, suggesting that NF1 limits their size by locally inhibiting Ras signalling.Our results link NF1 with macropinocytosis and phagocytosis for the first time, and we propose that NF1 evolved in early phagotrophs to spatially modulate Ras activity, thereby constraining and shaping their feeding structures.

View Article: PubMed Central - PubMed

Affiliation: MRC Laboratory of Molecular Biology, Cambridge, United Kingdom.

ABSTRACT
Cells use phagocytosis and macropinocytosis to internalise bulk material, which in phagotrophic organisms supplies the nutrients necessary for growth. Wildtype Dictyostelium amoebae feed on bacteria, but for decades laboratory work has relied on axenic mutants that can also grow on liquid media. We used forward genetics to identify the causative gene underlying this phenotype. This gene encodes the RasGAP Neurofibromin (NF1). Loss of NF1 enables axenic growth by increasing fluid uptake. Mutants form outsized macropinosomes which are promoted by greater Ras and PI3K activity at sites of endocytosis. Relatedly, NF1 mutants can ingest larger-than-normal particles using phagocytosis. An NF1 reporter is recruited to nascent macropinosomes, suggesting that NF1 limits their size by locally inhibiting Ras signalling. Our results link NF1 with macropinocytosis and phagocytosis for the first time, and we propose that NF1 evolved in early phagotrophs to spatially modulate Ras activity, thereby constraining and shaping their feeding structures.

No MeSH data available.


Related in: MedlinePlus