A role for SNX5 in the regulation of macropinocytosis.
BACKGROUND
The mechanisms and components that regulate macropinocytosis are poorly understood. Here we have investigated the role of sorting nexin 5 (SNX5) in the regulation of macropinocytic activity.


RESULTS
SNX5 is abundantly expressed in macrophages, cells very active in macropinocytosis, and is recruited onto newly-formed macropinosomes. LPS treatment of bone marrow-derived macrophages resulted in a 2.5 fold decrease in macropinosome formation that correlates with a reduction in the levels of SNX5. To investigate the relationship between SNX5 levels and macropinocytic activity we examined the formation of macropinosomes in HEK-FlpIn cells stably expressing GFP-SNX5. Constitutive macropinocytosis was increased approximately 2 fold in HEK-GFP-SNX5 cells compared with parental HEK-FlpIn cells. Furthermore, EGF stimulation resulted in a significant increase in macropinocytosis and there was also a 2.0 fold increase in the generation of macropinosomes in HEK-GFP-SNX5 cells compared with parental HEK-FlpIn cells. SNX5, which interacts specifically with PtdIns(3)P and PtdIns(3,4)P2 through its PX domain, was recruited to regions on the plasma membrane containing EGF receptor or positive for PtdIns(3,4)P2 as detected with the PH domain of TAPP1. Treatment with AG1478, an EGF receptor specific tyrosine kinase inhibitor, prevented the recruitment of SNX5 to the cytosolic face of the plasma membrane and inhibited the formation of macropinosomes in response to EGF treatment.


CONCLUSIONS
Based on these data, we propose that SNX5 requires the generation of phosphoinositides for recruitment to the plasma membrane and, moreover, influences the level of macropinocytic activity.
Department of Biochemistry and Molecular Biology, The University of Melbourne, Melbourne, Victoria 3010, Australia. j.lim20@pgrad.unimelb.edu.au
Jet Phey
JP
Lim
Jack T H
JT
Wang
Markus C
MC
Kerr
Rohan D
RD
Teasdale
Paul A
PA
Gleeson
D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000037:antagonists & inhibitors; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism; D008262:Macrophage Activation; D008264:Macrophages / Q000378:metabolism*; D008563:Membrane Lipids / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010873:Pinocytosis / Q000187:drug effects / Q000502:physiology*; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology*; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors; D011799:Quinazolines; D058305:Sorting Nexins; D019076:Transgenes / Q000235:genetics; D020032:Tyrphostins / Q000494:pharmacology; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*
D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides; D008563:Membrane Lipids; D008565:Membrane Proteins; D018129:Phosphatidylinositol Phosphates; C461495:Plekha1 protein, mouse; D011799:Quinazolines; C533712:Snx5 protein, mouse; D058305:Sorting Nexins; D020032:Tyrphostins; D033921:Vesicular Transport Proteins; C060980:phosphatidylinositol 3,4-diphosphate; C055525:phosphatidylinositol 3-phosphate; C101044:RTKI cpd; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D017027:Protein Tyrosine Phosphatases