Phospholipase D2 is required for efficient endocytic recycling of transferrin receptors.
RNA interference-mediated depletion of phospholipase D2 (PLD2), but not PLD1, inhibited recycling of transferrin receptors in HeLa cells, whereas the internalization rate was unaffected by depletion of either PLD. Although reduction of both PLD isoforms inhibits PLD activity stimulated by phorbol 12-myristic 13-acetate, only depletion of PLD2 decreased nonstimulated activity. Cells with reduced PLD2 accumulated a greater fraction of transferrin receptors in a perinuclear compartment that was positive for Rab11, a marker of recycling endosomes. EFA6, an exchange factor for Arf6, has been proposed to stimulate the recycling of transferrin receptors. Thus, one consequence of EFA6 overexpression would be a reduction of the internal pool of receptors. We confirmed this observation in control HeLa cells; however, overexpression of EFA6 failed to decrease the internal pool of transferrin receptors that accumulate in cells previously depleted of PLD2. These observations suggest that either PLD2 is required for a constitutive Arf6-mediated recycling pathway or in the absence of PLD2 transferrin receptors accumulate in recycling endosomes that are not responsive to overexpression of EFA6.
Padrón
David
D
Department of Biochemistry, The University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390-9038.
Tall
Renee D
RD
Roth
Michael G
MG
D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D020662:Guanine Nucleotide Exchange Factors; D006367:HeLa Cells; D006801:Humans; D009419:Nerve Tissue Proteins / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000096:biosynthesis; D010739:Phospholipase D / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D034622:RNA Interference; D011990:Receptors, Transferrin / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000099193:rab11 GTP-Binding Proteins
D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors; D020662:Guanine Nucleotide Exchange Factors; D009419:Nerve Tissue Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D010739:Phospholipase D; D017853:Phosphotransferases (Alcohol Group Acceptor); D011990:Receptors, Transferrin; D013755:Tetradecanoylphorbol Acetate; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C109970:PSD protein, human; C105606:phospholipase D2; C106801:phospholipase D1; C000715348:ARF6 protein, human