“Presence of vitamin D3 receptor (VDR)-like proteins in Solanum glaucophyllum”.

LORENA MILANESI; RICARDO BOLAND

PHYSIOLOGIA PLANTARUM

Elsevier

Lugar: Irlanda; Año: 2006 vol. 128 p. 341 - 350

0031-9317

ABSTRACT     The detection of 1a,25(OH)2-vitamin D3 [1a,25(OH)2D3] in Solanum glaucophyllum and other plant species has been reported. Little is known about the mode of action of 1a,25(OH)2D3 in plants. The steroid acts in animal systems through binding to a specific receptor (VDR). In the present work, competition assays allowed the detection of binding sites for [3H]-1a,25(OH)2D3 in cultured S. glaucophyllum cells. The binding of [3H]-1a,25(OH)2D3 to total protein from calli was a saturable process with respect to the ligand concentration (Kd=  3.49 ± 0.75 nM; Bmax= 255.7 ± 18.6 fmol/mg protein). Western blots employing antibodies directed against the avian VDR revealed three immunoreactive bands of ~72, ~58 and ~43 kDa in different plant organs, cells and callus cultures. The immunoreactive proteins were tyrosine phosphorylated and the signal detected was increased using hypertonic buffers for protein extraction suggesting that these proteins are associated to subcellular structures. When the callus cells were subjected to homogenization and differential centrifugation, a substantial proportion of the immunolabeling was localized in the nuclear and mitochondrial fractions. Ligand blot assays revealed the ability of the reactive bands to bind [3H]-1a,25(OH)2D3. Immunocytochemistry detection using Oregon Green-conjugated secondary antibodies showed immunofluorescent staining mainly in nucleus and in nucleus, cytoplasmic corpuscles and outer cell membrane for the monoclonal and polyclonal antibodies, respectively. [3H]thymidine incorporation assays and immunoblot analysis of mitogen-activated protein kinase (MAPK) indicated that 1a,25(OH)2D3 and cell lipid extracts stimulate DNA synthesis but do not activate MAPKs with TEY domains in S. glaucophyllum cells. These data demonstrate that S. glaucophyllum cells, which produce 1a,25(OH)2D3, also express a cognate binding protein (s) and are able to respond to 1a,25(OH)2D3. The exact functional role of the VDR-like protein (s) remains to be established.