Cellular retinoic acid binding protein 2 inhibits osteogenic differentiation by modulating LIMK1 in C2C12 cells

Abstract

Cellular retinoic acid binding protein 2 (CRABP2) is essential for myoblast differentiation, however, little is known about its role in osteogenic differentiation. This study mainly aims to explore the biological functions and the underlying molecular mechanisms of CRABP2 in osteogenesis. Using quantitative polymerase chain reaction and western blot assays, we found that the expression of CRABP2 at both mRNA and protein levels were downregulated during osteogenesis. Furthermore, CRABP2 knockdown displayed significant changes in the cell phenotype and the actin filaments (F-actin) polymerization in C2C12 cells treated with BMP2. Moreover, the western blotting of osteogenic differentiation biomarkers, alkaline phosphatase (ALP) staining and Alizarin red staining showed that CRABP2 dramatically inhibited osteogenic differentiation. The following investigation of molecular mechanisms implicated that CARBP2 specifically interacted with LIMK1, a key factor in acin cytoskeletal rearrangements in osteogenesis, to interrupt its activity and stability in an ubiquitin-proteasome pathway to prevent C2C12 cells from osteogenic differentiation in response to BMP2. Above all, our data suggest a novel function of CRABP2 in regulating actin remodeling and osteogenic differentiation via LIMK1, thus presenting a possible molecular target for promoting the osteogenic differentiation in bone degenerative diseases.

Retinoic Acid Signal Negatively Regulates Osteo/Odontogenic Differentiation of Dental Pulp Stem Cells

“…CRABP2 knockdown enhances odontoblastic differentiation of human DPSCs. It is downregulated during osteogenic differentiation from myogenic progenitor cells and negatively regulates osteogenic differentiation [ 39 ]. Miniature pigs are similar to humans in their mandibular anatomy and diphyodont dentition [ 40 ].…”

Ubiquitination and Long Non-coding RNAs Regulate Actin Cytoskeleton Regulators in Cancer Progression

“…Rnf6 serves as a ubiquitin ligase and binds to LIMK1, contributing to Lys48-linked polyubiquitination of LIMK1 in the presence of UbcH5, which functions as an E2 enzyme. CRABP2 (cellular retinoic acid binding protein 2) participates in osteogenic differentiation via interacting with LIMK1 in a ubiquitin-proteasome pathway and compromises LIMK1 activity [118]. However, the exact domain of CRABP2 that interacts with LIMK1 and which ubiquitin ligase is involved in this process remain unclear.…”

Effect of CRABP2 on the proliferation and odontoblastic differentiation of hDPSCs

“…Furthermore, CRABP2 is involved in the RA, β-catenin and Notch signaling pathways, as well as in the interaction between epithelial and mesenchymal cells, all of which are reported to regulate tooth development. 12 Most importantly, CRABP2 regulates bone remodeling in murine degenerative joint disease models in vivo and osteogenic differentiation of myoblast cell lines (C2C12) in vitro, 13 which shares similar characteristics with odontoblastic differentiation. To the best of our knowledge, the expression and biological role of CRABP2 in hDPSCs have not been reported to date.…”

“…CRABP2 regulates osteogenic differentiation via LIMK to remodel cell morphology in vitro 13 . CRABP2 was also reported to regulate bone remodeling in murine degenerative joint disease models in vivo, 24 and to be modulated by the transcription factors MyoD and Sp1 to promote myoblast differentiation in C2C12 cells.…”