白蝦類專一性免疫作用分子 Dscam 之蛋白質特性分析

Abstract

Invertebrate Down syndrome cell adhesion molecule (Dscam) seems to play key roles in neuron guidance and “innate immunity with specificity”. The extracellular region of Dscam is involved in heterophilic binding specificity and pathogen recognition while the cytoplasmic tail is involved in subsequent signal transduction. In 2009, Litopenaeus vannamei Dscam (LvDscam) with extraordinary molecular diversity was identified and shown a unique tail-less secreted form. In particular, at different WSSV-infected situations, the specific population of LvDscam isoforms was appeared. These isoforms might play antibody-like function. The membrane-bound shrimp Dscam was first identified from Penaeus monodon with the cytoplasmic tail with extra hypervariable ability. Because of this uniqueness of shrimp Dscam, we investigated the characteristics of shrimp Dscam protein using the recombinant protein expression system in alternative cell line. This observation may help us to clarify the interaction between shrimp Dscam and cell. Four tail-less secreted shrimp Dscam isoforms were selected to analyze the protein characteristics. Isoform 7 is the highest expressed isoform in heamocyte of WSSV-free shrimp, while isoforms 18, 19, and 20 are the redundant isoforms isolated from WSSV-persistent shrimp. Because shrimp cell line has not yet been established, we set to the expression platform of recombinant shrimp Dscam (rLvDscam) in Spodoptera frugiperda (Sf9) cells using baculovirus expression system and the transfection system. The cellular localization patterns of recombinant tail-less Dscam (rLvDscam) in insect cells show that rLvDscam may localize both in vesicles in cytoplasm and cell membrane. Some vesicles containing rLvDscam may involve in the processes of endocytosis or exocytosis. Moreover, tail-less rLvDscam can be secreted. Based on these information, we propose a model of how the tail-less LvDscam return cell. For membrane-bound shrimp Dscam, using the alignment of the cytoplasmic tail elements of PmDscam and LvDscam, the high identity was observed (97.78%). We’ll further analysis the localization and secreted-ability of the membrane-bound rLvDscam. The study provides the first information about shrimp Dscam protein. In future, we hope to find the evidence supporting the Dscam involvement in the invertebrate innate immunity with specificity.