The voltage dependence of the TMEM16B/anoctamin2 calciumactivated chloride channel is modified by mutations in the first putative intracellular loop

Valentina Cenedese, Giulia Betto, Fulvio Celsi, O. Lijo Cherian, Simone Pifferi, Anna Menini

Research output: Contribution to journalArticle

Abstract

Ca 2+-activated Cl - channels (CaCCs) are involved in several physiological processes. Recently, TMEM16A/anoctamin1 and TMEM16B/anoctamin2 have been shown to function as CaCCs, but very little information is available on the structure-function relations of these channels. TMEM16B is expressed in the cilia of olfactory sensory neurons, in microvilli of vomeronasal sensory neurons, and in the synaptic terminals of retinal photoreceptors. Here, we have performed the first site-directed mutagenesis study on TMEM16B to understand the molecular mechanisms of voltage and Ca 2+ dependence. We have mutated amino acids in the first putative intracellular loop and measured the properties of the wild-type and mutant TMEM16B channels expressed in HEK 293T cells using the whole cell voltage-clamp technique in the presence of various intracellular Ca 2+ concentrations. We mutated E367 into glutamine or deleted the five consecutive glutamates 386EEEEE 390 and 399EYE 401. The EYE deletion did not significantly modify the apparent Ca 2+ dependence nor the voltage dependence of channel activation. E367Q and deletion of the five glutamates did not greatly affect the apparent Ca 2+ affinity but modified the voltage dependence, shifting the conductance-voltage relations toward more positive voltages. These findings indicate that glutamates E367 and 386EEEEE 390 in the first intracellular putative loop play an important role in the voltage dependence of TMEM16B, thus providing an initial structure-function study for this channel.

Original languageEnglish
Pages (from-to)285-294
Number of pages10
JournalJournal of General Physiology
Volume139
Issue number4
DOIs
Publication statusPublished - Apr 2012

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Glutamates
Chloride Channels
Mutation
Olfactory Receptor Neurons
Physiological Phenomena
Vertebrate Photoreceptor Cells
Cilia
HEK293 Cells
Presynaptic Terminals
Patch-Clamp Techniques
Sensory Receptor Cells
Microvilli
Site-Directed Mutagenesis
Glutamine
Amino Acids

ASJC Scopus subject areas

  • Physiology

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The voltage dependence of the TMEM16B/anoctamin2 calciumactivated chloride channel is modified by mutations in the first putative intracellular loop. / Cenedese, Valentina; Betto, Giulia; Celsi, Fulvio; Cherian, O. Lijo; Pifferi, Simone; Menini, Anna.

In: Journal of General Physiology, Vol. 139, No. 4, 04.2012, p. 285-294.

Research output: Contribution to journalArticle

Cenedese, Valentina ; Betto, Giulia ; Celsi, Fulvio ; Cherian, O. Lijo ; Pifferi, Simone ; Menini, Anna. / The voltage dependence of the TMEM16B/anoctamin2 calciumactivated chloride channel is modified by mutations in the first putative intracellular loop. In: Journal of General Physiology. 2012 ; Vol. 139, No. 4. pp. 285-294.
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