Cens Thierry, Rousset Matthieu, Collet Claude, Charreton Mercedes, Garnery Lionel, Le Conte Yves, Chahine Mohamed, Sandoz Jean-Christophe, Charnet Pierre
Institut des Biomolécules Max Mousseron (IBMM), CNRS UMR 5247, Place Eugène Bataillon, 34095 Montpellier cedex 5, France; Centre de Recherche de Biochimie Macromoléculaire (CRBM), CNRS UMR 5237, 1919 Route de Mende, 34293 Montpellier cedex 5, France; Université de Montpellier, Place Eugène Bataillon, 34095 Montpellier cedex 5, France.
INRA UR 406 Abeilles et Environnement, 228 Route de l'aérodrome, Domaine Saint Paul, Site Agroparc, CS40509, 84914 Avignon cedex 9, France.
Insect Biochem Mol Biol. 2015 Mar;58:12-27. doi: 10.1016/j.ibmb.2015.01.005. Epub 2015 Jan 17.
Voltage-gated Ca(2+) channels allow the influx of Ca(2+) ions from the extracellular space upon membrane depolarization and thus serve as a transducer between membrane potential and cellular events initiated by Ca(2+) transients. Most insects are predicted to possess three genes encoding Cavα, the main subunit of Ca(2+) channels, and several genes encoding the two auxiliary subunits, Cavβ and Cavα2δ; however very few of these genes have been cloned so far. Here, we cloned three full-length cDNAs encoding the three Cavα subunits (AmelCav1a, AmelCav2a and AmelCav3a), a cDNA encoding a novel variant of the Cavβ subunit (AmelCavβc), and three full-length cDNAs encoding three Cavα2δ subunits (AmelCavα2δ1 to 3) of the honeybee Apis mellifera. We identified several alternative or mutually exclusive exons in the sequence of the AmelCav2 and AmelCav3 genes. Moreover, we detected a stretch of glutamine residues in the C-terminus of the AmelCav1 subunit that is reminiscent of the motif found in the human Cav2.1 subunit of patients with Spinocerebellar Ataxia type 6. All these subunits contain structural domains that have been identified as functionally important in their mammalian homologues. For the first time, we could express three insect Cavα subunits in Xenopus oocytes and we show that AmelCav1a, 2a and 3a form Ca(2+) channels with distinctive properties. Notably, the co-expression of AmelCav1a or AmelCav2a with AmelCavβc and AmCavα2δ1 produces High Voltage-Activated Ca(2+) channels. On the other hand, expression of AmelCav3a alone leads to Low Voltage-Activated Ca(2+) channels.
电压门控性Ca(2+)通道可在细胞膜去极化时使Ca(2+)离子从细胞外间隙内流，因此充当膜电位与由Ca(2+)瞬变引发的细胞事件之间的转换器。据预测，大多数昆虫拥有三个编码Ca(2+)通道主要亚基Cavα的基因，以及几个编码两个辅助亚基Cavβ和Cavα2δ的基因；然而，到目前为止这些基因中只有极少数被克隆出来。在此，我们克隆了三个编码三个Cavα亚基（AmelCav1a、AmelCav2a和AmelCav3a）的全长cDNA、一个编码Cavβ亚基新变体（AmelCavβc）的cDNA，以及三个编码蜜蜂意大利蜜蜂三个Cavα2δ亚基（AmelCavα2δ1至3）的全长cDNA。我们在AmelCav2和AmelCav3基因序列中鉴定出了几个可变或互斥外显子。此外，我们在AmelCav1亚基的C末端检测到一段谷氨酰胺残基，这让人联想到6型脊髓小脑共济失调患者的人类Cav2.1亚基中发现的基序。所有这些亚基都包含在其哺乳动物同源物中已被确定为功能重要的结构域。我们首次能够在非洲爪蟾卵母细胞中表达三种昆虫Cavα亚基，并且我们表明AmelCav1a、2a和3a形成具有独特特性的Ca(2+)通道。值得注意的是，AmelCav1a或AmelCav2a与AmelCavβc和AmCavα2δ1共表达会产生高电压激活的Ca(2+)通道。另一方面，单独表达AmelCav3a会导致低电压激活的Ca(2+)通道。
