Interaction Between Endosymbiont Holospora and the Cell

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1. We found that nuclear envelope-recognition sensor substances of H.obtusa and H.undulata were lipo-polysaccharides (LPSs) on their outer membranes. Indirect immunofluorescence microscopy and immuno-electron microscopy with monoclonal antibodies for these LPSs showed that the antibodies reacted with their target nuclear envelopes.2. Holospora penetrates the target nuclear envelope with its specific tip. When this special tip was cut off from about 3,400 infectious forrns with a micro-dissection system and loaded on SDS-PAGE, 2 bands appeared with silver staining. These area of SDS-PAGE gels were cut and inside materials were electroeluted. Using these eluted substances as antigens, we succeeded to develop a monoclonal antibody specific for the special substance with indirect immunofluorescence microscopy.3. Eighteen spots were cut off from 2D-SDS-PAGE of H.obtusa and the micro-sequences of the purified proteins from their N-terminals were clarified and homology checks with known proteins were carried out. Also, their ORFs were searched from H.obtusa genomic DNA sequences so far clarified. As a the results, only 3 ORFs were found among 18 proteins, and 8 proteins showed high homologies with known proteins. The fact that only 3 ORF were detected shows that majority of the H.obtusa proteins may derived from the host cells.4. We found 12 host genes that were depressed and 3 host genes by the infection of H.obtusa by differential display (DD) method. Partial DNA sequences of 4 genes of the former ones were clarified. One of the 4 genes showed was an immobilization antigen A homologous gene of Paramecium tetraurelia.