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KMID : 0578319960060030316
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Molecules and Cells
1996 Volume.6 No. 3 p.316 ~ p.324
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Lysosomes Are in subpopulations in Amoeba proteus
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Yoo, So Yeun
Choi, Ji Young/Kim, Hyeonjung/Ahn, Tae In
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Abstract
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We obtained two clones of cells producing monoclonal antibodies (mAbs), LCA45 and LYA64, against Iysosomal proteins of Amoeba proteus. The specificity of these mAbs was determined by indirect immunofluorescence microscopy and immunoblotting following SDS-polyacrylamide gel electrophoresis of Iysosomal fractions. LCA45 reacted with 45 kDa proteins shown as the granules in the Iysosomes, while LYA64 recognized 64 and 45 kDa proteins of Iysosomal membranes. LCA45-stained Iysosomes began to fuse with newly formed phagososomes containing Tetrahymena from 3 h after their formation, and the immunofluorescence of the phagolysosomes was the most intense at 8 h after phagocytosis. Hereafter LCA45-stained antigens were dispersed in sporadic small Iysosomes all over the cytoplasm even in the presence of tunicamycin. On the other hand, LYA64-stained Iysosomes began to fuse with phagosomes from 10 min, and the intensity of immunofluorescence on the membranes of phagolysosomes reached maximum in 2 h. Prior to fusing with phagosomes, Iysosomes identified by both mAbs fused among themselves. When the amoebae were stained with both antibodies, some Iysosomes showed double immunofluorescence and more Iysosomes got stained with LYA64 only. Thus, LCA45-stained late-fusing Iysosomes are apparently a subpopulation of Iysosomes containing LYA64 antigens on the membrane. This implies that the early fusing Iysosomes are another subpopulation of Iysosomes sharing the same membrane antigens. With these observations we conclude that Iysosomes in amoeba are composed of more than two subpopulations that are different in behavior of fusing with the newly formed phagosomes.
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