Abstract
Sex steroids exert anti-apoptotic effects on osteoblasts/osteocytes but exert pro-apoptotic effects on osteoclasts, in both cases requiring activation of the extracellular signal-regulated kinases (ERKs). To explain the mechanistic basis of this divergence, we searched for differences in the kinetics of phosphorylation and/or in the subcellular localization of ERKs in response to 17beta-estradiol in the two cell types. In contrast to its transient effect on ERK phosphorylation in osteocytic cells (return to base line by 30 min), 17beta-estradiol-induced ERK phosphorylation in osteoclasts was sustained for at least 24 h following exposure to the hormone. Conversion of sustained ERK phosphorylation to transient, by means of cholera toxin-induced activation of the adenylate cyclase/cAMP/protein kinase A pathway, abrogated the pro-apoptotic effect of 17beta-estradiol on osteoclasts. Conversely, prolongation of ERK activation in osteocytes, by means of leptomycin B-induced inhibition of ERK export from the nucleus or overexpression of a green fluorescent protein-ERK2 mutant that resides permanently in the nucleus, converted the anti-apoptotic effect of 17beta-estradiol to a pro-apoptotic one. These findings indicate that the kinetics of ERK phosphorylation and the length of time that phospho-ERKs are retained in the nucleus are responsible for pro-versus anti-apoptotic effects of estrogen on different cell types of bone and perhaps their many other target tissues.
Highlights
Sex steroids exert anti-apoptotic effects on osteoblasts/osteocytes but exert pro-apoptotic effects on osteoclasts, in both cases requiring activation of the extracellular signal-regulated kinases (ERKs)
Sustained Versus Transient Nuclear Accumulation of ERK2 in Osteoclasts and Osteocytes—In osteoblastic/osteocytic cells, estradiol or 5␣-dihydrotestosterone induces rapid phosphorylation of ERKs and causes rapid translocation of the phosphorylated kinase into the nucleus [13, 14]. This finding, together with evidence that cell survival can depend on the duration of ERK nuclear accumulation [21, 29], prompted us to compare the kinetics of this phenomenon in osteoclasts and osteocytes (Fig. 2)
Osteoclasts derived from the RAW264.7 monocytic cell line as well as MLO-Y4 cells were transiently co-transfected with constructs containing GFPERK2, mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK), and nRFP
Summary
Vol 280, No 6, Issue of February 11, pp. 4632–4638, 2005 Printed in U.S.A. Transient Versus Sustained Phosphorylation and Nuclear Accumulation of ERKs Underlie Anti- Versus Pro-apoptotic Effects of Estrogens*. A shortening of the life span of boneforming cells in combination with prolongation of the life span of bone-resorbing cells represent critical pathophysiologic changes in most acquired metabolic bone diseases, including the osteoporosis that results from sex steroid deficiency and from glucocorticoid excess or old age (4, 6 –9) Other agents, such as parathyroid hormone and bisphosphonates, used commonly for the treatment of metabolic bone diseases, exert their beneficial effects on bone by regulating the rate of birth of new osteoblasts or osteoclasts or their apoptosis (10 –12). The duration of the nuclear accumulation of ERKs is determined, at least in part, by the action of nuclear phosphatases, which dephosphorylate ERKs and thereby promote their export back to the cytoplasm [30]
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