Growth hormone attenuates the transcriptional activity of Runx2 by facilitating its physical association with Stat3β

Abstract

We document that GH controls osteoblast function by modulating the biological activity of the osteospecific transcription factor Runx2. Evidence is provided for a physical interaction between Runx2 and Stat3β, which is enhanced by GH and downregulates the transcriptional properties of this key osteogenic regulator. Introduction: Growth hormone (GH) signals to bone either through insulin-like growth factor-1 or directly by influencing the function of osteoblasts, the bone-forming cells. This study aimed at exploring the molecular events that underlie the direct biological action of GH on osteoblastic cells, and specifically, the effects that it might exert on the function of the bone-specific transcriptional regulator Runx2. Materials and Methods: The GH-responsive human osteoblastic cell line Saos-2 was used as our experimental system. Western blot analyses were used to monitor the presence of several parameters known to be affected by GH in these cells (i.e., downregulation of GH receptor, induction of STATs, and extracellular signal-regulated kinase [ERK] mitogen-activated protein kinase [MAPK] pathways). Electrophoretic mobility shift assays were used to assess Runx2 and Stat3 binding activity on an osteoblast-specific element (OSE2) after GH treatment. A combination of yeast two-hybrid and co-immunoprecipitation assays were performed to test for the existence of a physical Runx2-Stat3β association. Finally, co-transfection experiments were used to investigate the interplay of the two transcription factors on the activity of a p6OSE2-Luc promoter after GH stimulation. Results: We show that GH signaling through Stat3/ERK MAPK potentiates the DNA binding activity of Runx2 but, at the same time, restrains its transcriptional potential. Moreover, a novel physical interaction of Runx2 with transcription factor Stat3β, which is enhanced by GH stimulation, was documented both in vitro and in vivo. Importantly, this interaction impairs the transcriptional activity of Runx2 without affecting its DNA binding capacity. Conclusion: Our data provide the first evidence that GH modulates the transcriptional function of Runx2 in osteoblastic cells by promoting its inhibitory interaction with Stat3β. Shedding light on such mechanisms will contribute to a better understanding of GH effects on skeletal homeostasis that may impact on decisions at the clinical level, especially in diseases affecting bone quantity and quality (e.g., osteoporosis). © 2004 American Society for Bone and Mineral Research.