Detection of cell type and marker specificity of nuclear binding sites for anionic carbohydrate ligands.
The emerging functionality of glycosaminoglycan chains engenders interest in localizing specific binding sites using cytochemical tools. We investigated nuclear binding of labeled heparin, heparan sulfate, a sulfated fucan, chondroitin sulfate, and hyaluronic acid in epidermal keratinocytes, bone marrow stromal cells, 3T3 fibroblasts and glioma cells using chemically prepared biotinylated probes. Binding of the markers was cell-type specific and influenced by extraction of histones, but was not markedly affected by degree of proliferation, differentiation or malignancy. Cell uptake of labeled heparin and other selected probes and their transport into the nucleus also was monitored. Differences between keratinocytes and bone marrow stromal cells were found. Preincubation of permeabilized bone marrow stromal cells with label-free heparin reduced the binding of carrier-immobilized hydrocortisone to its nuclear receptors. Thus, these tools enabled binding sites for glycosaminoglycans to be monitored in routine assays.
Chovanec
M
M
Institute of Anatomy, Charles University, 1st Faculty of Medicine, U nemocnice 3, 128 000 Prague 2, Czech Republic.
Smetana
K
K
Purkrábková
T
T
Holíková
Z
Z
Dvoránková
B
B
André
S
S
Pytlík
R
R
Hozák
P
P
Plzák
J
J
Sedo
A
A
Vacík
J
J
Gabius
H
H
D016475:3T3 Cells; D000818:Animals; D000838:Anions; D001665:Binding Sites; D019921:Biotinylation / Q000379:methods*; D001854:Bone Marrow Cells / Q000378:metabolism*; D050260:Carbohydrate Metabolism; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D005910:Glioma / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000378:metabolism*; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism*; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D000838:Anions; D005456:Fluorescent Dyes; D006025:Glycosaminoglycans; D018160:Receptors, Cytoplasmic and Nuclear