Martin Samuel A M, Mohanty Bimal P, Cash Phillip, Houlihan Dominic F, Secombes Christopher J
Scottish Fish Immunology Research Centre, School of Biological Sciences, University of Aberdeen, Aberdeen, UK.
Proteomics. 2007 Jun;7(13):2275-86. doi: 10.1002/pmic.200700020.
Type II IFN exists as a single molecule (IFN-gamma) in contrast to type I IFN, of which there are a number of different forms. IFN-gamma is involved both directly and indirectly in antiviral activity, stimulation of bactericidal activity, antigen presentation and activation of macrophages. Recently IFN-gamma was cloned from a salmonid fish, the rainbow trout and a functional recombinant protein produced exhibited IFN-gamma activity. This recombinant IFN-gamma was used to stimulate an Atlantic salmon cell line, SHK-1, to monitor the changes in protein expression by proteomic analysis 24 h after stimulation compared to unstimulated control cells. An SHK-1 cell proteome map was developed and proteins altered in abundance by the IFN-gamma stimulation were identified. Under the analytical conditions used, 22 proteins were found to be altered in abundance, 15 increased and 7 decreased. Several proteins were excised from the gel and identified, following trypsin digestion and MALDI-MS/MS/LC-MS and database interrogation. Transcriptional analysis of five mRNAs encoding proteins increased in abundance by IFN-gamma in the proteome analysis was determined by real-time PCR. We assessed the correlation between gene expression and change in abundance of proteins for these genes.
与I型干扰素不同，II型干扰素以单一分子（干扰素-γ）的形式存在，I型干扰素有多种不同形式。干扰素-γ直接或间接参与抗病毒活性、刺激杀菌活性、抗原呈递以及巨噬细胞的激活。最近，从一种鲑科鱼类虹鳟鱼中克隆出了干扰素-γ，并产生了具有干扰素-γ活性的功能性重组蛋白。该重组干扰素-γ用于刺激大西洋鲑鱼细胞系SHK-1，通过蛋白质组分析监测刺激后24小时与未刺激对照细胞相比蛋白质表达的变化。绘制了SHK-1细胞蛋白质组图谱，并鉴定了因干扰素-γ刺激而丰度改变的蛋白质。在所使用的分析条件下，发现22种蛋白质的丰度发生了改变，15种增加，7种减少。从凝胶中切下几种蛋白质，经胰蛋白酶消化、基质辅助激光解吸电离串联质谱/液相色谱-质谱分析和数据库查询后进行鉴定。通过实时PCR对蛋白质组分析中因干扰素-γ而丰度增加的5种编码蛋白质的mRNA进行转录分析。我们评估了这些基因的基因表达与蛋白质丰度变化之间的相关性。
