ABSTRACT
The differentiation of embryonic mesenchymal cells into chondrocytes and the subsequent formation of a cartilaginous scaffold that enables the formation of long bones are hallmarks of endochondral ossification. During this process, chondrocytes undergo a remarkable sequence of events involving proliferation, differentiation, hypertrophy and eventually apoptosis. Forkhead Box O (FoxO) transcription factors (TFs) are well-known regulators of such cellular processes. Although FoxO3a was previously shown to be regulated by 1,25-dihydroxyvitamin D3 in osteoblasts, a possible role for this family of TFs in chondrocytes during endochondral ossification remains largely unstudied. By crossing Collagen2-Cre mice with FoxO1lox/lox;FoxO3alox/lox;FoxO4lox/lox mice, we generated mice in which the three main FoxO isoforms were deleted in growth plate chondrocytes (chondrocyte triple knock-out; CTKO). Intriguingly, CTKO neonates showed a distinct elongation of the hypertrophic zone of the growth plate. CTKO mice had increased overall body and tail length at eight weeks of age and suffered from severe skeletal deformities at older ages. CTKO chondrocytes displayed decreased expression of genes involved in redox homeostasis. These observations illustrate the importance of FoxO signaling in chondrocytes during endochondral ossification.
Subject(s)
Bone and Bones/metabolism , Chondrocytes/metabolism , Forkhead Box Protein O1/genetics , Forkhead Box Protein O3/genetics , Forkhead Transcription Factors/genetics , Osteogenesis/genetics , Animals , Bone and Bones/cytology , Cell Cycle Proteins , Cell Differentiation , Cell Proliferation , Chondrocytes/cytology , Collagen Type II/genetics , Collagen Type II/metabolism , Crosses, Genetic , Female , Forkhead Box Protein O1/deficiency , Forkhead Box Protein O3/deficiency , Forkhead Transcription Factors/deficiency , Gene Expression Profiling , Gene Expression Regulation, Developmental , Integrases/genetics , Integrases/metabolism , Male , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Mice , Mice, Transgenic , Oxidoreductases/genetics , Oxidoreductases/metabolism , Primary Cell Culture , Signal TransductionABSTRACT
1Alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3], the biologically active form of vitamin D3, is a pleiotropic hormone that exerts its effects on a wide range of tissues, resulting in different biological responses such as anticancer activity. It is the ligand of the vitamin D receptor (VDR), a nuclear receptor with transactivating capacity. We demonstrated in this study that 1,25(OH)2D3 induces PDZ-LIM domain-containing protein 2 (PDLIM2) expression. PDLIM2 is an adaptor molecule that links different components of the cytoskeleton, and was recently shown to be repressed in human breast cancer cells by hypermethylation of regulatory promoter regions, leading to enhanced tumorigenicity. We demonstrated that PDLIM2 was a direct target gene of 1,25(OH)2D3; its upregulation was VDR-dependent and a functional VDRE in the promoter was identified. Moreover, 1,25(OH)2D3 induced demethylation of the PDLIM2 promoter, leading to enhanced transcription. Finally, PDLIM2 was found to be crucial for 1,25(OH)2D3-induced cell adhesion and for mediating the ability of 1,25(OH)2D3 to suppress cancer cell migration and invasion. This study provides mechanistic insights into the anticancer activities of 1,25(OH)2D3 in human breast cancer cells.
Subject(s)
Antineoplastic Agents/pharmacology , Cell Movement , Gene Expression Regulation, Neoplastic/drug effects , LIM Domain Proteins/biosynthesis , Microfilament Proteins/biosynthesis , Vitamin D/analogs & derivatives , Blotting, Western , Cell Adhesion/drug effects , Cell Adhesion/genetics , Cell Movement/drug effects , Cell Movement/genetics , Chromatin Immunoprecipitation , Gene Expression Regulation, Neoplastic/genetics , Humans , LIM Domain Proteins/genetics , MCF-7 Cells , Microfilament Proteins/genetics , Neoplasm Invasiveness/genetics , Promoter Regions, Genetic/genetics , Real-Time Polymerase Chain Reaction , Receptors, Calcitriol/genetics , Receptors, Calcitriol/metabolism , Transcriptional Activation/drug effects , Vitamin D/pharmacologyABSTRACT
Various epidemiological studies have shown an aetiological link between vitamin D deficiency and cancer incidence. The active metabolite of vitamin D, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], has potent anti-cancer activities both in vitro and in vivo. These anti-cancer effects are attained by regulating the transcription of numerous genes that are involved in different pathways to reduce tumorigenesis and are dependent on the cancer cell type. Besides reducing cell growth and inducing apoptosis, 1,25(OH)2D3 also inhibits angiogenesis and metastasis. Moreover, its potency to inhibit inflammation also contributes to its anti-tumoral activity. Here, we report the different ways in which 1,25(OH)2D3 interferes with the malignant processes that are activated in cancer cells.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Anticarcinogenic Agents/therapeutic use , Antineoplastic Agents/therapeutic use , Calcitriol/therapeutic use , Animals , Anti-Inflammatory Agents, Non-Steroidal/metabolism , Anticarcinogenic Agents/metabolism , Antineoplastic Agents/metabolism , Calcitriol/metabolism , Humans , Neoplasms/etiology , Neoplasms/prevention & control , Neoplasms/therapy , Vitamin D Deficiency/physiopathologyABSTRACT
Mutations in the BRCA1-interacting DEAH helicase Brip1 confer an increased risk of breast cancer. In the present study we aimed to unravel the transcriptional control of Brip1 and to determine its expression levels in a set of 101 primary invasive breast carcinomas. Transcription of Brip1 was found to be cell growth-related and controlled by the E2F/retinoblastoma (Rb) pathway through a conserved E2F-responsive site. Repression of Brip1 expression by the cell growth-inhibiting compound 1alpha,25-dihydroxyvitamin D3 depended on this same E2F-responsive site. In spite of its role as a tumor suppressor, both quantitative reverse transcriptase-PCR analyses and immunohistochemical stainings showed significantly elevated Brip1 expression levels in grade 3 tumors as compared to grade 1 or 2 carcinomas. Furthermore, increased Brip1 transcript levels were found in tumors with an estrogen receptor-negative, progesterone receptor-negative or HER-2-positive status. In conclusion, these data show that Brip1 is a genuine target gene for the E2F/Rb pathway and that elevated expression levels of Brip1 are detected in primary invasive breast carcinomas with unfavorable characteristics.
Subject(s)
Breast Neoplasms/genetics , Carcinoma/genetics , DNA-Binding Proteins/genetics , E2F Transcription Factors/physiology , Gene Expression Regulation, Neoplastic , RNA Helicases/genetics , Animals , Base Sequence , Basic-Leucine Zipper Transcription Factors/genetics , Binding Sites , Breast Neoplasms/pathology , Carcinoma/pathology , Conserved Sequence , E2F Transcription Factors/metabolism , Fanconi Anemia Complementation Group Proteins/genetics , Female , Humans , Mice , Molecular Sequence Data , Neoplasm Invasiveness , Sequence Homology, Nucleic Acid , Transcription, Genetic , Tumor Cells, CulturedABSTRACT
1alpha,25-Dihydroxyvitamin D(3) [1,25-(OH) (2)D(3)] can exert its biological actions through binding with the nuclear vitamin D receptor (VDR), a ligand-activated transcription factor. Next to control of bone and mineral homeostasis, these actions include an immunomodulatory effect and a potent growth-inhibitory, antiproliferative or prodifferentiating action on a wide variety of cell types. The molecular mechanisms underlying this antiproliferative action form an intriguing research topic and they remain, although thoroughly studied, not completely understood. Important cell cycle regulators are involved such as cyclins, cyclin dependent kinases and their corresponding inhibitors as well as E2F transcription factors and accompanying pocket proteins. Whether 1,25-(OH)(2)D(3) influences the expression of all these proteins directly through the nuclear VDR or rather in an indirect manner is not always clear. The antiproliferative action makes 1,25-(OH) (2)D(3) a possible therapeutic tool to treat hyperproliferative disorders, among which different types of cancer. Clinical application, however, is severely hampered by calcemic effects such as hypercalcemia, hypercalciuria and increased bone resorption. Rational design of chemically modified 1,25-(OH) (2)D(3)-analogs tries to overcome this problem. As such, several thousands of analogs have been synthesized and evaluated, some of which display the desired dissociation between beneficial antiproliferative and unwanted calcemic effects. A number of those analogs are 'superagonistic' and have a several-fold stronger antiproliferative action than the parent compound. This review focuses on recent findings about the complex mechanisms behind the antiproliferative and prodifferentiating effect of 1,25-(OH) (2)D(3). Furthermore, the mode of action and possible clinical application of chemically modified 1,25-(OH) (2)D(3)-analogs will be discussed.
