FIG. 1.

Deficient FN expression and responsiveness in Mgat5^−/− cells. (A) Cell spreading of Mgat5^+/+ and Mgat5^−/− cells plated for 24 h was measured on an FN substrate at the indicated concentrations. (B) Total cell lysates (40 μg protein) of Mgat5^+/+ and Mgat5^−/− PyMT mammary tumor cell lines and Mgat5^−/− cells rescued by retroviral expression of Mgat5 (rescued) were blotted for FN, laminin, and β-actin, as indicated. Molecular mass markers (in kDa) are indicated, and the graph shows the densitometric quantification of band intensity (±SEM; n = 3; *, P < 0.01 relative to Mgat5^+/+ cells). (C) Mgat5^+/+, Mgat5^−/−, and rescued cells grown in serum (plus fetal calf serum [+FCS]) or in the absence of serum (−FCS) on coverslips or on coverslips coated with 10 μg/ml of FN (+FN sub) for 48 h. Cells were fixed and labeled with mouse anti-FN monoclonal antibody and Alexa 568-conjugated anti-mouse secondary antibody. Bar, 20 μm.