Hastings Margaret H, Gong Katrina, Freibauer Alexander, Courchesne Caitlin, Fan Xiaotang, Sossin Wayne S
Department of Psychology Montreal Neurological Institute McGill University, Montreal, Quebec, Canada.
Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.
PLoS One. 2017 Oct 20;12(10):e0186646. doi: 10.1371/journal.pone.0186646. eCollection 2017.
Calpains are a family of intracellular proteases defined by a conserved protease domain. In the marine mollusk Aplysia californica, calpains are important for the induction of long-term synaptic plasticity and memory, at least in part by cleaving protein kinase Cs (PKCs) into constitutively active kinases, termed protein kinase Ms (PKMs). We identify 14 genes encoding calpains in Aplysia using bioinformatics, including at least one member of each of the four major calpain families into which metazoan calpains are generally classified, as well as additional truncated and atypical calpains. Six classical calpains containing a penta-EF-hand (PEF) domain are present in Aplysia. Phylogenetic analysis determined that these six calpains come from three separate classical calpain families. One of the classical calpains in Aplysia, AplCCal1, has been implicated in plasticity. We identify three splice cassettes and an alternative transcriptional start site in AplCCal1. We characterize several of the possible isoforms of AplCCal1 in vitro, and demonstrate that AplCCal1 can cleave PKCs into PKMs in a calcium-dependent manner in vitro. We also find that AplCCal1 has a novel mechanism of auto-inactivation through N-terminal cleavage that is modulated through its alternative transcriptional start site.
钙蛋白酶是一类由保守蛋白酶结构域定义的细胞内蛋白酶。在海洋软体动物加州海兔中，钙蛋白酶对于长期突触可塑性和记忆的诱导很重要，至少部分是通过将蛋白激酶C（PKC）切割成组成型活性激酶，即蛋白激酶M（PKM）来实现的。我们利用生物信息学在海兔中鉴定出14个编码钙蛋白酶的基因，包括后生动物钙蛋白酶通常分类的四个主要钙蛋白酶家族中每个家族的至少一个成员，以及额外的截短型和非典型钙蛋白酶。海兔中存在六种含有五聚EF手（PEF）结构域的经典钙蛋白酶。系统发育分析确定这六种钙蛋白酶来自三个不同的经典钙蛋白酶家族。海兔中的一种经典钙蛋白酶AplCCal1与可塑性有关。我们在AplCCal1中鉴定出三个剪接盒和一个可变转录起始位点。我们在体外对AplCCal1的几种可能的同工型进行了表征，并证明AplCCal1在体外可以以钙依赖的方式将PKC切割成PKM。我们还发现AplCCal1具有一种通过N端切割实现自动失活的新机制，这种机制通过其可变转录起始位点进行调节。
