PP2A catalytic subunit silence by microRNA-429 activates AMPK and protects osteoblastic cells from dexamethasone
Introduction
Long-term and/or excessive clinical use of glucocorticoids could lead to osteoporosis or even osteonecrosis [1]. Dexamethasone (Dex) and other glucocorticoids will induce direct damages to osteoblasts [1], [2], [3]. Indeed, osteoblast cell apoptosis is often detected in bones of Dex-taking patients [4], [5]. Our group [6], [7], [8], [9] and others [10], [11], [12] have been adding Dex to the cultured osteoblasts/osteoblastic cells, which mimics glucocorticoid-induced cell injuries [4], [7], [8], [13], [14]. This cellular model is utilized to explore the pathological mechanisms of Dex-induced osteoblast cell damages, which shall help to develop possible intervention strategies [6], [7], [8], [9].
AMP-activated protein kinase (AMPK) is a highly-conserved serine/threonine protein kinase [15]. It is a key energy sensor, which controls cell metabolism during environmental stresses [15]. Emerging evidences have implied that AMPK activation could also promote cell survival [16]. Studies, including ours [9], [17], have investigated the potential function of AMPK in osteoblasts [9], [18], [19], [20]. It was found that genetic or pharmacological activation of AMPK could efficiently protect osteoblasts from Dex [9], [18], [19], [20].
Studies have suggested that protein phosphatase 2A (PP2A) is a key AMPK phosphatase [21], [22]. Inhibition of PP2A, with a selective PP2A inhibitor (okadaic acid) or PP2A knockdown, could provoke or restore AMPK activation [21], [22]. PP2A is composed of a scaffold (PP2A-a) and a catalytic subunit (PP2A-c) [21], [22]. The 36 kDa PP2A-c is tightly associated with the 65 kDa PP2A-c to form the core structure of PP2A [21], [22]. In the current study, we aim to identify microRNA (“miRNA”) that could possibly activate AMPK via silencing PP2A-c. Our results here discover that microRNA-429 (“miR-429”) selectively targets 3’-untranslated region (UTR) of PP2A-c mRNA. Significantly, expression of miR-429 silences PP2A-c to activate AMPK signaling, therefore protecting osteoblastic cells from Dex.
Section snippets
Chemicals and reagents
Dex, neomycin and puromycin were obtained from Sigma Aldrich (Shanghai, China). Cell culture reagents were provided by Gibco (Shanghai, China). Antibodies of this study were purchased from Cell Signaling Technology (Danvers, MA).
Cell culture
The OB-6 [4] and hFOB1.19 [23] human osteoblastic cell lines were obtained from the Cell Bank of Shanghai Institute of Biological Science (Shanghai, China). Osteoblastic cells were cultured as described [4], [23].
miR-429 transfection
Osteoblastic cells were initially transfected with 30 nM
Expression of miR-429 downregulates PP2A catalytic subunit and activates AMPK in human osteoblastic cells
Our previous studies [9], [17] have demonstrated that AMPK activation could inhibit Dex-induced osteoblast cell death. In the current study, we aim to provoke AMPK activation via miRNA-mediated downregulating its phosphatase PP2A [21], [22]. TargetScan miRNA database was searched. We discovered that miR-429 putatively targets the 3′UTR of PP2A-c mRNA (Fig. 1A). To study the potential effect of miR-429 on PP2A-c, miR-429 was introduced to hFOB1.19 osteoblastic cells, and stable hFOB1.19 cell
Discussions
AMPK is mainly composed of the catalytic α subunit and β/γ regulatory subunits [27], [28]. Multiple isoforms and transcriptional variants of these subunits have been characterized [27], [28]. Thr172 phosphorylation of AMPKα1 subunit is key to AMPK activation [27], [29], [30]. Different groups have been focusing on the mechanisms of AMPKα1 phosphorylation [31]. Several AMPKα1 kinases have been identified, including LKB1 (Liver kinase B1) [30], CaMKK (calcium/calmodulin-dependent protein kinase
Conflicts of interest
The listed authors have no conflict of interests.
Acknowledgments
This work is supported by the National Natural Science Foundation (No. 81672170. To: Feng Ji).
References (38)
- et al.
Glucocorticoids in osteonecrosis of the femoral head: a new understanding of the mechanisms of action
J. Steroid Biochem. Mol. Biol.
(2009) - et al.
Dexamethasone-induced apoptosis of osteocytic and osteoblastic cells is mediated by TAK1 activation
Biochem. Biophys. Res. Commun.
(2015) - et al.
MHY1485 activates mTOR and protects osteoblasts from dexamethasone
Biochem. Biophys. Res. Commun.
(2016) - et al.
K6PC-5, a novel sphingosine kinase 1 (SphK1) activator, alleviates dexamethasone-induced damages to osteoblasts through activating SphK1-Akt signaling
Biochem. Biophys. Res. Commun.
(2015) - et al.
alpha-Melanocyte stimulating hormone attenuates dexamethasone-induced osteoblast damages through activating melanocortin receptor 4-SphK1 signaling
Biochem. Biophys. Res. Commun.
(2016) - et al.
Activating AMP-activated protein kinase by an alpha1 selective activator compound 13 attenuates dexamethasone-induced osteoblast cell death
Biochem. Biophys. Res. Commun.
(2016) - et al.
EGFR trans-activation mediates pleiotrophin-induced activation of Akt and Erk in cultured osteoblasts
Biochem. Biophys. Res. Commun.
(2014) - et al.
Activating AMP-activated protein kinase by an alpha1 selective activator Compound 13 attenuates dexamethasone-induced osteoblast cell death
Biochem. Biophys. Res. Commun.
(2016) - et al.
Activation of AMPK protects against hydrogen peroxide-induced osteoblast apoptosis through autophagy induction and NADPH maintenance: new implications for osteonecrosis treatment?
Cell Signal
(2014) - et al.
Activation of protein phosphatase 2A by palmitate inhibits AMP-activated protein kinase
J. Biol. Chem.
(2007)
Hepcidin increases intracellular Ca2+ of osteoblast hFOB1.19 through L-type Ca2+ channels
Regul. Pept.
Belinostat-induced apoptosis and growth inhibition in pancreatic cancer cells involve activation of TAK1-AMPK signaling axis
Biochem. Biophys. Res. Commun.
Co-activation of AMPK and mTORC1 induces cytotoxicity in acute myeloid leukemia
Cell Rep.
Advances in glucocorticoid-induced osteoporosis
Curr. Rheumatol. Rep.
Clinical practice. Glucocorticoid-induced bone disease
N. Engl. J. Med.
Glucocorticoid induces apoptosis of osteoblast cells through the activation of glycogen synthase kinase 3beta
J. Bone Min. Metab.
P53 dependent mitochondrial permeability transition pore opening is required for dexamethasone-induced death of osteoblasts
J. Cell Physiol.
Glucocorticoid receptor and sequential P53 activation by dexamethasone mediates apoptosis and cell cycle arrest of osteoblastic MC3T3-E1 cells
PLoS One
EGFR-AKT-mTOR activation mediates epiregulin-induced pleiotropic functions in cultured osteoblasts
Mol. Cell Biochem.
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