Luxmi Raj, Blaby-Haas Crysten, Kumar Dhivya, Rauniyar Navin, King Stephen M, Mains Richard E, Eipper Betty A
Department of Neuroscience, University of Connecticut Health Center, Farmington, CT 06030-3401, USA.
Department of Biology, Brookhaven National Laboratory, Upton, NY 11973-5000, USA.
Proteomes. 2018 Sep 23;6(4):36. doi: 10.3390/proteomes6040036.
The recent identification of catalytically active peptidylglycine α-amidating monooxygenase (PAM) in Chlamydomonas reinhardtii, a unicellular green alga, suggested the presence of a PAM-like gene and peptidergic signaling in the last eukaryotic common ancestor (LECA). We identified prototypical neuropeptide precursors and essential peptide processing enzymes (subtilisin-like prohormone convertases and carboxypeptidase B-like enzymes) in the genome. Reasoning that sexual reproduction by C. reinhardtii requires extensive communication between cells, we used mass spectrometry to identify proteins recovered from the soluble secretome of mating gametes, and searched for evidence that the putative peptidergic processing enzymes were functional. After fractionation by SDS-PAGE, signal peptide-containing proteins that remained intact, and those that had been subjected to cleavage, were identified. The C. reinhardtii mating secretome contained multiple matrix metalloproteinases, cysteine endopeptidases, and serine carboxypeptidases, along with one subtilisin-like proteinase. Published transcriptomic studies support a role for these proteases in sexual reproduction. Multiple extracellular matrix proteins (ECM) were identified in the secretome. Several pherophorins, ECM glycoproteins homologous to the sex-inducing pheromone, were present; most contained typical peptide processing sites, and many had been cleaved, generating stable N- or C-terminal fragments. Our data suggest that subtilisin endoproteases and matrix metalloproteinases similar to those important in vertebrate peptidergic and growth factor signaling play an important role in stage transitions during the life cycle of .
最近在单细胞绿藻莱茵衣藻中发现了具有催化活性的肽基甘氨酸α-酰胺化单加氧酶（PAM），这表明在最后的真核生物共同祖先（LECA）中存在类似PAM的基因和肽能信号传导。我们在基因组中鉴定出了典型的神经肽前体和必需的肽加工酶（枯草杆菌蛋白酶样激素原转化酶和羧肽酶B样酶）。鉴于莱茵衣藻的有性生殖需要细胞间广泛的通讯，我们使用质谱法鉴定从交配配子的可溶性分泌组中回收的蛋白质，并寻找证据证明推定的肽能加工酶具有功能。通过SDS-PAGE分级分离后，鉴定出了保持完整的含信号肽的蛋白质以及经过切割的蛋白质。莱茵衣藻的交配分泌组包含多种基质金属蛋白酶、半胱氨酸内肽酶和丝氨酸羧肽酶，以及一种枯草杆菌蛋白酶样蛋白酶。已发表的转录组学研究支持这些蛋白酶在有性生殖中的作用。在分泌组中鉴定出了多种细胞外基质蛋白（ECM）。存在几种与性诱导信息素同源的ECM糖蛋白——性信息素结合蛋白；大多数含有典型的肽加工位点，许多已经被切割，产生了稳定的N端或C端片段。我们的数据表明，类似于在脊椎动物肽能和生长因子信号传导中起重要作用的枯草杆菌蛋白酶内切酶和基质金属蛋白酶，在莱茵衣藻生命周期的阶段转变中发挥着重要作用。
