Integrating rapid responses to 1,25-dihydroxyvitamin D3 with transcriptional changes in osteoblasts: Ca2+ regulated pathways to the nucleus.
It is well established that 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) treatment of target cells including osteoblasts activates both membrane-initiated rapid Ca2+ responses linked to influx through voltage sensitive Ca2+ channels (VSCCs) and longer term nuclear receptor-mediated changes in gene expression. We recently reported use of a cDNA microarray strategy to identify transcriptional changes after 3 and 24h of treatment with 1,25(OH)2D3 and with an analog of 1,25(OH)2D3 (25(OH)-16ene-23yne-D3 [AT]) that activates Ca2+ influx without binding to the nuclear receptor. Among 5000 different clones on the array filters, we identified families of genes in osteoblasts that were altered two-fold or greater following treatment with 1,25(OH)2D3 or analog AT for 3h. Cluster analysis further revealed complex patterns of changes in gene expression, indicative of multiple pathways to the nucleus. Evidenced by changes in target gene expression, activation of a Ca2+/CaMK/CREB/CRE pathway clearly occurs and modulates expression of a variety of genes associated with changes in protein secretion including those involved in paracrine regulation of bone resorption, RANKL and osteoprotegerin (OPG). The changes in gene expression can be inhibited by L-type VSCC channel blockers, confirming the role of Ca2+ entry in pathway activation. These findings provide clear evidence of rapid changes in gene expression associated with Ca2+ influx after treatment with 1,25(OH)2D3, and open the door to novel nuclear receptor-independent signaling pathways that affect gene transcription.
Department of Biological Sciences, University of Delaware, 326 Wolf Hall, Newark, DE 19716, USA. farachca@udel.edu
Mary C
MC
Farach-Carson
Joel J
JJ
Bergh
Yihuan
Y
Xu
D000818:Animals; D001862:Bone Resorption / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006023:Glycoproteins / Q000096:biosynthesis / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D010006:Osteoblasts / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D053244:Osteoprotegerin; D019899:Paracrine Communication; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear / Q000096:biosynthesis / Q000378:metabolism; D018124:Receptors, Tumor Necrosis Factor; D014158:Transcription, Genetic / Q000187:drug effects; D014807:Vitamin D / Q000031:analogs & derivatives* / Q000494:pharmacology*
D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D053244:Osteoprotegerin; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear; D018124:Receptors, Tumor Necrosis Factor; C505687:Tnfrsf11a protein, mouse; C506031:Tnfrsf11b protein, mouse; C505689:Tnfsf11 protein, mouse; C118756:dihydroxy-vitamin D3; D014807:Vitamin D; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium