RNAi knockdown of human kinetochore protein CENP-H |
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Authors: | Orthaus Sandra Ohndorf Sabine Diekmann Stephan |
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Institution: | Department of Molecular Biology, FLI e.V., Beutenbergstrasse 11, D-07745 Jena, Germany. |
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Abstract: | The inner kinetochore protein complex binds to centromeres during the whole cell cycle. It serves as the basis for the binding of further kinetochore proteins during mitosis. CENP-H is one of the inner kinetochore proteins which is conserved amongst many eukaryotes. By specific RNAi knockdown, we reduced the CENP-H protein level in human HEp-2 cells down to less than 5% of its normal value. In these CENP-H knocked-down cells, we observed severe mitotic phenotypes like misaligned chromosomes and multipolar spindles, however, no mitotic arrest. Strong reduction of CENP-H resulted in a slightly reduced CENP-C level at the kinetochores and normal localisation of hBubR1, indicating a functional mitotic checkpoint at the hBubR1 protein level. In CENP-H knocked-down human cells, the misaligned chromosomes contained only reduced levels of CENP-E. Our data clearly indicate that CENP-H has an important impact on the architecture and function of the human kinetochore complex. |
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Keywords: | Mitosis Human kinetochore CENP-H CENP-C hBubR1 CENP-E Mitotic checkpoint RNAi knockdown |
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