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. 2009 Aug 13;28(17):2554–2567. doi: 10.1038/emboj.2009.199

Figure 5.

Figure 5

GLFG repeat domains also form a highly selective permeability barrier. Panels show influx of mobile species into a hydrogel made of 200 mg/ml of a fusion between the GLFG repeat domains of Nup49p and Nup57p. Quantification was carried out analogous to Figure 3. (A) Influx of 3 μM of MBP-mCherry alone. (B) Simultaneous influx of 3 μM of MBP-mCherry and 1 μM of IBB-MBP-mEGFP·scImpβ complex. (C) As panel B, however, the gel was pre-incubated for 180 min with 10 μM of scImpβ. This pre-incubation tightened the barrier against passive influx, but also diminished receptor-mediated cargo entry as well as intra-gel diffusion of the NTR·cargo complex, suggesting that the GLFG gel is more sensitive to competition by scImpβ than the FG/FxFG gel (see Figure 3C). (D) As panel B, however, the gel was pre-incubated with the dominant-negative human Impβ45-462 fragment. This pre-incubation lead to a virtually complete block of passive influx and to a 100-fold reduction in receptor-mediated gel entry. Intra-gel movement of the scImpβ·cargo complex was essentially blocked (also see Supplementary Figure S4). The effect of the dominant-negative mutant on the GLFG gel was much stronger than on an FG/FxFG gel (see Supplementary Figure S4).