Effects of cyclic nucleotides on the function of prestin

Levente Deák, Jing Zheng, Alex Orem, Guo Guang Du, Salvador Aguiñaga, Keiji Matsuda, Peter Dallos*

*Corresponding author for this work

Research output: Contribution to journalArticle

66 Scopus citations

Abstract

Outer hair cells (OHCs) in the mammalian organ of Corti display electromotility, which is thought to provide the local active mechanical amplification of the cochlear response. Prestin is the key molecule responsible for OHC electromotility. Several compounds, including cGMP, have been shown to influence OHC electromotility. There are two potential cAMP/cGMP-dependent protein kinase phosphorylation sites on prestin. Whether these sites are involved in cGMP-dependent reactions is as yet unknown. In this study, prestin cDNA was transiently transfected into TSA 201 cells. Cells that expressed prestin were selected to measure non-linear capacitance (NLC), a signature of outer hair cell motility. We applied cGMP and cAMP analogues and a protein kinase G (PKG) antagonist to the cells. Furthermore, nine mutations at putative phosphorylation sites of prestin were produced. The neutral amino acid alanine replaced serine/threonine at phosphorylation sites to change the conserved phosphorylation motif in order to mimic the dephosphorylated state of prestin, whereas replacement with the negatively charged aspartic acid mimicked the phosphorylated state. The properties of such modified prestin-expressing cells were examined, through measurement of NLC and with confocal microscopy. Our data demonstrate that cGMP is significantly more influential than cAMP in modifying the non-linear, voltage-dependent charge displacement in prestin-transfected cells. The electrical properties of the single and double mutations further indicate a possible interaction between the two PKG target sites. One of these sites may influence the membrane targeting process of prestin. Finally, a new topology map of prestin is proposed.

Original languageEnglish (US)
Pages (from-to)483-496
Number of pages14
JournalJournal of Physiology
Volume563
Issue number2
DOIs
StatePublished - Mar 1 2005

ASJC Scopus subject areas

  • Physiology

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    Deák, L., Zheng, J., Orem, A., Du, G. G., Aguiñaga, S., Matsuda, K., & Dallos, P. (2005). Effects of cyclic nucleotides on the function of prestin. Journal of Physiology, 563(2), 483-496. https://doi.org/10.1113/jphysiol.2004.078857