FIGURE 3.

Intact heparan sulfate chains are required for shed syndecan-1 to translocate to the nucleus of cells. A, sSDC1-containing medium was added to ST2 cells without or with prior treatment of medium with hep III for 2 h, or the conditioned medium was added in the presence of excess heparan sulfate (10 μg/ml) or heparin (10 μg/ml). The amount of sSDC1 in the nucleus of ST2 cells was quantified by ELISA. Data are mean ± S.E. of three independent experiments. *, p < 0.05 versus untreated controls. B, following incubation with medium containing sSDC1, ST2 cells were immunostained for syndecan-1 (red). Results confirm ELISA data shown in A. Incubation of ST2 cells with conditioned medium collected from CAG cells expressing sSDC1 without glycosaminoglycan chains (sSDC1 ΔGAG) confirms heparan sulfate chains are required for sSDC1 translocation to the nucleus. The nuclei are counterstained with DAPI (blue). Scale bar, 10 μm. C, CAG cells engineered to express sSDC1 were grown in the presence of NaClO₃ or NaClO₃ + Na₂SO₄ for 48 h, and conditioned media were collected and syndecan-1 levels measured. Equal amounts of syndecan-1 were added to ST2 cells for 2 h; nuclear lysates were collected, and the level of sSDC1 in the nucleus was quantified by ELISA. Data are mean ± S.E. of three independent experiments. *, p < 0.05 versus untreated controls.