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. 2002 Nov 11;159(3):465–476. doi: 10.1083/jcb.200205115

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Copyright © 2002, The Rockefeller University Press

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Figure 8. — p120 expression stabilizes E-cadherin by a post-transcriptional mechanism. (A) SW48 cells were infected with LZRS-p120-IRES-neo, and stable cell lines isolated. Levels of E-cadherin were assayed by Western blotting RIPA cell lysates. Parental (lane 1) or cell lines expressing neo only (lanes 2 and 3) were compared with two separately derived p120 expressing cell lines (lanes 4 and 5) and to HCT116 cells (which express normal levels of p120 and E-cadherin). GM130 blotting (bottom) was used as a loading control. p120 expression increased E-cadherin levels by more than fivefold in SW48 cells. (B) E-cadherin mRNA levels are unchanged by p120 expression. Total RNA was isolated and compared by Northern analysis using a human E-cadherin cDNA probe. To control for loading, the blot was stripped and reprobed with GAPDH cDNA. MDA-MB-231 cells (lane 1) do not express E-cadherin mRNA. SW48 cells stably expressing p120–4A (lane 2), p120–3A (lane 3), or neo alone (lane 4), are compared with SW48 parental (lane 5), HCA7 (lane 6), and HCT116 cells (lane 7). (C) Analysis of E-cadherin stability in p120-expressing cells. Clonal (p120 3A/neo-15) or polyclonal (p120–3A/neo) cell lines were generated by infection. E-cadherin half life was ascertained by pulse-chase analysis and compared with control clonal (neo-18) and polyclonal (neo) cell lines. Chase times are indicated across the top. Densitometric analyses are located below each lane and represent a normalized value where the 4 h chase lane (top) or the 2 h chase lane (bottom) represent the value 1.0. p120 expression essentially doubles the half life of E-cadherin.