Subcellular localization and preliminary function study of dynein regulation protein robl/LC7 from Nocardia seriolae

Fish nocardiosis is a chronic systemic granulomatous disease, and Nocardia seriolae is its main pathogen. A dynein regulation protein robl/LC7, which is likely to be secreted and to target host cell mitochondria, was found by the bioinformatics analysis on the whole genome sequence of N. seriolae. In order to determine the subcellular localization and study the preliminary function of robl/LC7 from N. seriolae, the gene cloning, eukaryotic expression plasmid construction, secreted protein identification, subcellular localization and apoptosis detection of robl/LC7 were carried out in this study. robl/LC7 was successfully cloned from N. seriolae and the eukaryotic expression plasmid of pEGFP- robl/LC7 and pcDNA- robl/LC7 were constructed. Mass spectrometry analysis of extracellular products from N. seriolae confirmed that robl/LC7 was a secreted protein. Subcellular localization of robl/LC7-GFP fusion protein discovered that the green fluorescence was distributed in the whole cell and did not co-localize with mitochondria. In addition, apoptosis assay revealed that apoptosis was induced by the over-expression of robl/LC7 in FHM cell. robl/LC7 from N. seriolae is a secreted protein which does not co-localize with mitochondria. It may help the host cell survival and immune escape of N. seriolae by participating the cell apoptosis regulation and may play an important role in the pathogenesis of N. seriolae.