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Basolateral expression of GRP94 in parietal cells of gastric mucosa

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Abstract

GRP94 is a member of the heat shock protein family normally confined to the endoplasmic reticulum that sometimes escapes the KDEL-mediated retention system. It is overexpressed in some gastric and other gastrointestinal carcinomas, but little is known about the physiological role of GRP94 in gastric mucosa. We investigated the membrane presence of GRP94 in parietal cells, which secrete acid into the gastric lumen, using subcellular fractionation, selective solubilization of membrane proteins, Western blotting, and radio-ligand binding and provided evidence of functional GRP94 expression at the surface of gastric mucosa parietal cells anchored to the basolateral domain. Our results show that GRP94 is not an integral membrane protein since 50 mM Na2CO3 treatment dissociates part of it from the membrane. However, 100 mM Na2CO3 treatment did not extract all GRP94 from the membrane, which indicates that it is strongly associated with it. The presence of GRP94 in isolated plasma membrane was demonstrated by Western blotting and its functionality by radio-lig- and binding experiments. Both the K D value obtained in saturation experiments with N-ethylcarboxamido-[3H]adenosine at 4°C, at the nanomolar range, and the inhibition constant of its binding by radicicol, the most specific GRP94 inhibitor, indicate that active receptor regions are exposed at the membrane surface. Western blotting of plasma membrane subfractions showed that GRP94 is mainly expressed in the basolateral membrane of gastric parietal cells, while its presence in the apical domain is negligible, thereby inferring a role for GRP94 in processes operating in this membrane domain.

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Abbreviations

ER:

endoplasmic reticulum

GRP94:

glucose-regulated protein of 94 kDa

Hsp:

heat shock protein

NECA:

N-ethylcarboxamidoadenosine

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Authors and Affiliations

  1. Department of Physiology, Faculty of Medicine and Dentistry, University of the Basque Country UPV/EHU, Barrio Sarriena s/n, 48940, Leioa, Spain

    R. M. Arin, Y. Rueda, A. I. Vallejo & B. Ochoa

  2. Department of Physiology, Faculty of Pharmacy, University of the Basque Country UPV/EHU, Paseo de la Universidad 7, 01006, Vitoria-Gasteiz, Spain

    O. Casis & M. Gallego

Authors
  1. R. M. Arin
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  2. Y. Rueda
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  3. O. Casis
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  4. M. Gallego
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  5. A. I. Vallejo
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  6. B. Ochoa
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Correspondence to R. M. Arin.

Additional information

Published in Russian in Biokhimiya, 2014, Vol. 79, No. 1, pp. 13–21.

Originally published in Biochemistry (Moscow) On-Line Papers in Press, as Manuscript BM13-177, November 17, 2013.

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Arin, R.M., Rueda, Y., Casis, O. et al. Basolateral expression of GRP94 in parietal cells of gastric mucosa. Biochemistry Moscow 79, 8–15 (2014). https://doi.org/10.1134/S0006297914010027

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  • DOI: https://doi.org/10.1134/S0006297914010027

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