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. 2011 Dec 7;40(7):3006–3017. doi: 10.1093/nar/gkr1197

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© The Author(s) 2011. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 4. — Manipulation of the p180 level affects membrane-associated polysomes. The amounts of ER-associated polysomes were analyzed using stable HeLa transfectants overexpressing p180 (HeLa/p180) and a control cell line (HeLa/puro). (A) The membrane fractions were obtained by sequential digitonin extractions from cont-si-treated HeLa/puro cells, cont-si-treated HeLa/p180 cells and p180-si-treated HeLa/p180 cells, and subjected to polysome analyses using a 15–50% sucrose gradient. (B) The RNA contents of the polysome fractions were estimated by densitometric scanning of the gels, and the relative ratios of polysome:80S are shown. (C) The distributions of p180 and other translocon-related proteins in the collected fractions were analyzed by western blotting. Data for cont-si-treated HeLa/puro cells (left), cont-si-treated HeLa/p180 cells (middle) and p180-si-treated HeLa/p180 cells (right) are presented. (D) The cytosolic fractions were subjected to polysome analyses on a 15–50% sucrose gradient, and the rRNAs were analyzed.