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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

New insights in osteogenic differentiation revealed by mass spectrometric assessment of phosphorylated substrates in murine skin mesenchymal cells

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Author(s):
Halcsik, Erik [1] ; Forni, Maria Fernanda [1] ; Fujita, Andre [2] ; Verano-Braga, Thiago [3] ; Jensen, Ole Norregaard [3] ; Sogayar, Mari Cleide [1]
Total Authors: 6
Affiliation:
[1] Univ Sao Paulo, Sch Med, Cell & Mol Therapy Ctr NUCEL NETCEM, Chem Inst, Dept Biochem, BR-05508000 Sao Paulo - Brazil
[2] Univ Sao Paulo, Inst Math & Stat, Dept Comp Sci, BR-05508090 Sao Paulo - Brazil
[3] Univ Southern Denmark, Dept Biochem & Mol Biol, Odense - Denmark
Total Affiliations: 3
Document type: Journal article
Source: BMC CELL BIOLOGY; v. 14, OCT 22 2013.
Web of Science Citations: 9
Abstract

Background: Bone fractures and loss represent significant costs for the public health system and often affect the patients quality of life, therefore, understanding the molecular basis for bone regeneration is essential. Cytokines, such as IL-6, IL-10 and TNF alpha, secreted by inflammatory cells at the lesion site, at the very beginning of the repair process, act as chemotactic factors for mesenchymal stem cells, which proliferate and differentiate into osteoblasts through the autocrine and paracrine action of bone morphogenetic proteins (BMPs), mainly BMP-2. Although it is known that BMP-2 binds to ActRI/BMPR and activates the SMAD 1/5/8 downstream effectors, little is known about the intracellular mechanisms participating in osteoblastic differentiation. We assessed differences in the phosphorylation status of different cellular proteins upon BMP-2 osteogenic induction of isolated murine skin mesenchymal stem cells using Triplex Stable Isotope Dimethyl Labeling coupled with LC/MS. Results: From 150 mu g of starting material, 2,264 proteins were identified and quantified at five different time points, 235 of which are differentially phosphorylated. Kinase motif analysis showed that several substrates display phosphorylation sites for Casein Kinase, p38, CDK and JNK. Gene ontology analysis showed an increase in biological processes related with signaling and differentiation at early time points after BMP2 induction. Moreover, proteins involved in cytoskeleton rearrangement, Wnt and Ras pathways were found to be differentially phosphorylated during all timepoints studied. Conclusions: Taken together, these data, allow new insights on the intracellular substrates which are phosphorylated early on during differentiation to BMP2-driven osteoblastic differentiation of skin-derived mesenchymal stem cells. (AU)