Phorbol Myristate Acetate Selectively Stimulates Apical Endocytosis via Protein Kinase C in Polarized MDCK Cells

Holm, Pernille; Eker, Per; Sandvig, Kirsten; Deurs, Bo van

doi:10.1006/excr.1995.1075

Cited by 48 publications

(46 citation statements)

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“…This phenomenon does not only apply to protein endocytosis and was well documented in MDCK cells when activators (such as derivatives of phorbol esters) and/or inhibitors of protein kinase C were employed (21,22). This fact was evident also in the present study, with marked inhibition of CaOx internalization after exposure to staurosporine.…”

Section: Discussionsupporting

confidence: 78%

Mechanisms involved in calcium oxalate endocytosis by Madin-Darby canine kidney cells

Campos

Schor

2000

Braz J Med Biol Res

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Calcium oxalate (CaOx) crystals adhere to and are internalized by tubular renal cells and it seems that this interaction is related (positively or negatively) to the appearance of urinary calculi. The present study analyzes a series of mechanisms possibly involved in CaOx uptake by Madin-Darby canine kidney (MDCK) cells. CaOx crystals were added to MDCK cell cultures and endocytosis was evaluated by polarized light microscopy. This process was inhibited by an increase in intracellular calcium by means of ionomycin (100 nM; N = 6; 43.9% inhibition; P<0.001) or thapsigargin (1 µM; N = 6; 33.3% inhibition; P<0.005) administration, and via blockade of cytoskeleton assembly by the addition of colchicine (10 µM; N = 8; 46.1% inhibition; P<0.001) or cytochalasin B (10 µM; N = 8; 34.2% inhibition; P<0.001). Furthermore, CaOx uptake was reduced when the activity of protein kinase C was inhibited by staurosporine (10 nM; N = 6; 44% inhibition; P<0.01), or that of cyclo-oxygenase by indomethacin (3 µM; N = 12; 17.2% inhibition; P<0.05); however, the uptake was unaffected by modulation of potassium channel activity with glibenclamide (3 µM; N = 6), tetraethylammonium (1 mM; N = 6) or cromakalim (1 µM; N = 6). Taken together, these data indicate that the process of CaOx internalization by renal tubular cells is similar to the endocytosis reported for other systems. These findings may be relevant to cellular phenomena involved in early stages of the formation of renal stones.

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Section: Discussionsupporting

confidence: 78%

Mechanisms involved in calcium oxalate endocytosis by Madin-Darby canine kidney cells

Campos

Schor

2000

Braz J Med Biol Res

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“…To examine potential cross-talk between K270 ubiquitylation and AQP2-S269 phosphorylation, we generated cell lines expressing N-terminal FLAG-tagged AQP2-WT, and AQP2-K270R and AQP2-S269A and -S269D mutants, and assessed the AQP2 ubiquitylation levels under control, forskolin, forskolin washout and forskolin plus TPA conditions. TPA is an activator of PKC that enhances apical endocytosis in MDCK cells (Holm et al, 1995) and enhances AQP2 ubiquitylation in the presence of forskolin (Kamsteeg et al, 2006). In MDCK cells, AQP2 was ubiquitylated at K270 with one to three moieties ( Fig.…”

Section: Aqp2 Ubiquitylation Occurs Independently Of Phosphorylation mentioning

confidence: 96%

Phosphorylation and ubiquitylation are opposing players in regulating endocytosis of the water channel Aquaporin-2

Moeller

Aroankins

Slengerik-Hansen

et al. 2014

Journal of Cell Science

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The post-translational modifications (PTMs) phosphorylation and ubiquitylation regulate plasma membrane protein function. Here, we examine the interplay between phosphorylation and ubiquitylation of the membrane protein aquaporin-2 (AQP2) and demonstrate that phosphorylation can override the previously suggested dominant endocytic signal of K63-linked polyubiquitylation. In polarized epithelial cells, although S256 is an important phosphorylation site for AQP2 membrane localization, the rate of AQP2 endocytosis was reduced by prolonging phosphorylation specifically at S269. Despite their close proximity, AQP2 phosphorylation at S269 and ubiquitylation at K270 can occur in parallel, with increased S269 phosphorylation and decreased AQP2 endocytosis occurring when K270 polyubiquitylation levels are maximal. In vivo studies support this data, with maximal levels of AQP2 ubiquitylation occurring in parallel to maximal S269 phosphorylation and enhanced AQP2 plasma membrane localization. In conclusion, we demonstrate for the first time that although K63-linked polyubiquitylation marks AQP2 for endocytosis, site-specific phosphorylation can counteract polyubiquitylation to determine its final localization. Similar mechanisms might exist for other plasma membrane proteins.

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“…Interestingly, this process turns out to be highly regulated, and also transcytosis is as a consequence changed. As indicated in Figure 3, endocytosis at the apical side is upregulated by activation of heterotrimeric G proteins 77 and activation of protein kinase C, 78 by protein kinase A, 77 and by addition of brefeldin A. 75 Also, it is regulated by the cyclooxygenase pathway 79 and by calmodulin.…”

Section: Transcytosis In Polarized Epithelial Cellsmentioning

confidence: 98%

Delivery into cells: lessons learned from plant and bacterial toxins

2005

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Phorbol Myristate Acetate Selectively Stimulates Apical Endocytosis via Protein Kinase C in Polarized MDCK Cells

Cited by 48 publications

References 0 publications

Mechanisms involved in calcium oxalate endocytosis by Madin-Darby canine kidney cells

Mechanisms involved in calcium oxalate endocytosis by Madin-Darby canine kidney cells

Phosphorylation and ubiquitylation are opposing players in regulating endocytosis of the water channel Aquaporin-2

Delivery into cells: lessons learned from plant and bacterial toxins

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