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First published online 23 June 2009
doi: 10.1242/jcs.046979

Journal of Cell Science 122, 2491-2503 (2009)
Published by The Company of Biologists 2009
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Research Article

Rescue of atypical protein kinase C in epithelia by the cytoskeleton and Hsp70 family chaperones

Anastasia Mashukova1, Andrea S. Oriolo1, Flavia A. Wald1, M. Llanos Casanova2, Cornelia Kröger3, Thomas M. Magin3, M. Bishr Omary4 and Pedro J. I. Salas1,*

1 Department of Cell Biology and Anatomy, University of Miami Miller School of Medicine, Miami, FL 33136, USA
2 CIEMAT, 28040 Madrid, Spain
3 University of Bonn Institute of Physiological Chemistry, 53115 Bonn, Germany
4 Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, MI 48109, USA

* Author for correspondence (e-mail: psalas{at}miami.edu)

Accepted 14 April 2009

Atypical PKC (PKC{iota}) is a key organizer of cellular asymmetry. Sequential extractions of intestinal cells showed a pool of enzymatically active PKC{iota} and the chaperone Hsp70.1 attached to the apical cytoskeleton. Pull-down experiments using purified and recombinant proteins showed a complex of Hsp70 and atypical PKC on filamentous keratins. Transgenic animals overexpressing keratin 8 displayed delocalization of Hsp70 and atypical PKC. Two different keratin-null mouse models, as well as keratin-8 knockdown cells in tissue culture, also showed redistribution of Hsp70 and a sharp decrease in the active form of atypical PKC, which was also reduced by Hsp70 knockdown. An in-vitro turn motif rephosphorylation assay indicated that PKC{iota} is dephosphorylated by prolonged activity. The Triton-soluble fraction could rephosphorylate PKC{iota} only when supplemented with the cytoskeletal pellet or filamentous highly purified keratins, a function abolished by immunodepletion of Hsp70 but rescued by recombinant Hsp70. We conclude that both filamentous keratins and Hsp70 are required for the rescue rephosphorylation of mature atypical PKC, regulating the subcellular distribution and steady-state levels of active PKC{iota}.

Key words: PKC{iota}, Keratins, Epithelial polarity, Protein chaperones, Apical kinases


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Atypical PKC: keratin to the rescue

JCS 2009 122: 1403. [Full Text]  





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