The tumor homing peptide F3 that binds both endothelial and tumor cells is internalized via surface nucleolin, while endostatin that inhibits angiogenesis binds nucleolin on the surface of endothelial cells before translocation to the nucleus [31], [32].
The tumor homing peptide F3 that binds both endothelial and tumor cells is internalized via surface nucleolin, while endostatin that inhibits angiogenesis binds nucleolin on the surface of endothelial cells before translocation to the nucleus [ 11, 13].
In cells without dynamin, the internalized to surface ratio of TfR was reduced ∼50% compared to control cells (Figure 4B).
Quantification of viral internalization showed that in RhoA CA transfected cells the virus uptake was clearly decreased (ratio of internalized vs. surface baculovirus; Figure 5B).
Cumulatively, these results indicate that although MVB-12 is not essential for the degradation of internalized cell surface proteins, it accelerates the kinetics of this process, consistent with a role for MVB-12 in modulating ESCRT-I activity.
We conclude that the C. elegans ESCRT-I complex has a conserved role in the degradation of internalized cell surface proteins that can be conveniently visualized by monitoring the fate of proteins normally targeted for degradation following fertilization.
Since depletion of the other three ESCRT-I subunits inhibits the degradation of internalized cell surface proteins, we wanted to determine if depletion of MVB-12 also affects this process.
HeLa cells grown in 12-well dishes were treated with 125I-EGF at 37°C and the ratio of internalized and surface radioactivity was determined during linear 3-min to calculate the specific internalization rate constant ke as described previously [7].
More recently, compartment-specific endosomal signalling mechanisms of internalized cell surface receptors have been identified [41], suggesting a role of endosomal signalling in development and progression of cancer [42].
In order to characterize the endocytic pathway followed by PrPC, we developed an indirect immunofluorescence assay in cells grown in polarized conditions, which allowed us to distinguish between internalized and surface protein (Figure 1A, Materials and Methods).
