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Differential Response of the Urothelial V-ATPase Activity to the Lipid Environment

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Abstract

The vesicle population beneath the apical plasma membrane of the most superficial urothelial cells is heterogeneous and their traffic and activity seems to be dependent on their membrane composition and inversely related to their development stage. Although the uroplakins, the major proteins of the highly differentiated urinary bladder umbrella cells, can maintain the bladder permeability barrier, the role of the membrane lipid composition still remains elusive. We have recently reported the lipid induced leakage of the vesicular content as a path of diversion in the degradative pathway. To extend the knowledge on how the lipid environment can affect vesicular acidification and membrane traffic through the regulation of the V-ATPase (vacuolar ATPase), we studied the proton translocation and ATP hydrolytic capacity of endocytic vesicles having different lipid composition obtained from rats fed with 18:1n-9 and 18:2n-6 fatty acid enriched diets. The proton translocation rate decreases while the enzymatic activity increases in oleic acid-rich vesicles (OAV), revealing an uncoupled state of V-ATPase complex which was further demonstrated by Western Blotting. A decrease of the very long fatty acyl chains length (C20–C24) and increase of the C16–C18 chains length in OAV membranes was observed, concomitant with increased hydrolytic activity of the V-ATPase. This response of the urothelial V-ATPase was similar to that of the Na–K ATPase when the activity of the latter was probed in reconstituted systems with lipids bearing different lengths of fatty acid chains. The studies describe for the first time a lipid composition-dependent activity of the urothelial V-ATPase, identified by immunofluorescence microscopy which is related to an effective coupling between the channel proton flux and ATP hydrolysis.

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Abbreviations

V-ATPase:

Vacuolar ATPase

AUM:

Asymmetric membrane unit

ATP:

Adenosine 5′-triphosphate

FITC:

Fluorescein isothiocyanate

PK/LDH:

Piruvate kinase/lactate dehydrogenase

NADH:

Reduced nicotinamide-adenine dinucleotide

PEP:

Phospho-enol-piruvate

CV:

Control vesicles

LAV:

Linoleic acid derived vesicles

OAV:

Oleic acid derived vesicles

VLCFAs:

Very long chain fatty acids

VLCPUFAs:

Very long chain unsaturated fatty acids

LCFAs:

Long chain fatty acids

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Acknowledgments

This study was supported by grants from SECYT-UNC and CONICET, Argentina. E.J. Grasso is a doctoral fellow of CONICET, Argentina. We are grateful to Mariana Piegari and Gina Mazzudulli for their technical assistance and to Dr. Pietro Ciancaglini for the kind gift of P-ATPase inhibitors.

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Authors and Affiliations

  1. Primera Cátedra de Biología Celular, H. y E, Instituto de Biología Celular-Facultad de Ciencias Médicas, Universidad Nacional de Córdoba, CC220, Ciudad Universitaria, 5000, Córdoba, Argentina

    E. J. Grasso, M. B. Scalambro & R. O. Calderón

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  1. E. J. Grasso
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  2. M. B. Scalambro
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  3. R. O. Calderón
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Correspondence to R. O. Calderón.

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Grasso, E.J., Scalambro, M.B. & Calderón, R.O. Differential Response of the Urothelial V-ATPase Activity to the Lipid Environment. Cell Biochem Biophys 61, 157–168 (2011). https://doi.org/10.1007/s12013-011-9172-x

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