Subcellular distribution of DsRed-NT5E fusion proteins.

<p>COS-7 cells were transiently transfected with expression vectors for DsRed monomer, wild type (WT) and mutant (F1–3) DsRed-hNT5E proteins and labeled with GFP-based stains specific for subcellular compartments: plasma membrane (<i>A, GFP-PM</i>), cytoplasm (<i>B, GFP-CYTO</i>) and endoplasmic reticulum (<i>C, GFP-ER</i>). A. While punctate fluorescence signals for all DsRed-hNT5E fusion proteins are observed in areas surrounding nuclei, only DsRed-hNT5E wild type form is also detected at the plasma membrane level and co-localizes with the plasma membrane-specific GFP stain (arrows). Fluorescence signal for control DsRed monomer was more uniformly distributed within cells relatively to DsRed-hNT5E fusion proteins and absent from the plasma membrane compartment. <i>Magnification 1000X</i>. B. All DsRed-hNT5E fusion proteins decorate intracellular punctate-shaped structures, with the exception DsRed-hNT5E wild type form being also present at the plasma membrane level (arrows). Also, fluorescence signal for DsRed-hNT5E F3 fusion protein appears to partially co-localize with the cytoplasm-specific GFP stain. Fluorescence signal for control DsRed monomer completely overlaps with that of the cytoplasm-specific GFP stain. <i>Magnification 1000X</i>. C. Fluorescence signals for all three mutant DsRed-hNT5E fusion proteins significantly co-localize with that of endoplasmic reticulum-specific GFP stain, although DsRed-hNT5E F2 fusion protein to a lesser extent than its F1 and F3 counterparts. Fluorescence signal for DsRed-hNT5E wild type form only partially co-localizes with that of endoplasmic reticulum-specific GFP stain, as it is also noted for control DsRed monomer. DsRed-hNT5E wild type protein is the only fusion protein associated with the plasma membrane (arrows) <i>Magnification 1000X</i>.