RhoA GTPase interacts with beta-catenin signaling in clinorotated osteoblasts

Abstract

Bone is a dynamic tissue under constant remodeling in response to various signals including mechanical loading. A lack of proper mechanical loading induces disuse osteoporosis that reduces bone mass and structural integrity. β-catenin signaling together with a network of GTPases is known to play a primary role in load-driven bone formation, but little is known about potential interactions of β-catenin signaling and GTPases in bone loss. In this study, we addressed a question: Does unloading suppress an activation level of RhoA GTPase and β-catenin signaling in osteoblasts? If yes, what is the role of RhoA GTPase and actin filaments in osteoblasts in regulating β-catenin signaling? Using a fluorescence resonance energy transfer (FRET) technique with a biosensor for RhoA together with a fluorescent T-cell factor/lymphoid enhancer factor (TCF/LEF) reporter, we examined the effects of clinostat-driven simulated unloading. The results revealed that both RhoA activity and TCF/LEF activity were downregulated by unloading. Reduction in RhoA activity was correlated to a decrease in cytoskeletal organization of actin filaments. Inhibition of β-catenin signaling blocked unloading-induced RhoA suppression, and dominant negative RhoA inhibited TCF/LEF suppression. On the other hand, a constitutively active RhoA enhanced unloading-induced reduction of TCF/LEF activity. The TCF/LEF suppression by unloading was enhanced by co-culture with osteocytes, but it was independent on organization of actin filaments, myosin II activity, or a myosin light chain kinase. Collectively, the results suggest that β-catenin signaling is required for unloading-driven regulation of RhoA, and RhoA, but not actin cytoskeleton or intracellular tension, mediates the responsiveness of β-catenin signaling to unloading.

RhoGTPases as Key Players in Mammalian Cell Adaptation to Microgravity

“…This may indicate, even indirectly, the activation of an adipogenic program under microgravity [ 125 ]. Moreover, Wan et al [ 126 ] recently demonstrated a changed RhoA and β -catenin signaling after 1 and 2.5 h, respectively, in clinorotated osteoblasts. They revealed that both the RhoA activity and the TCF/LEF (T-cell factor-1 and lymphoid enhancing factor-1) activity, a β -catenin recruiter, were downregulated by unloading.…”

“…However, the inhibition of β -catenin signaling blocked the unloading-induced RhoA suppression, and dominant negative RhoA inhibited the TCF/LEF suppression, revealing a regulation loop between β -catenin, RhoA, and TCF/LEF. Furthermore, while β -catenin signaling seemed to be required for microgravity regulation of RhoA, this response was not mediated by the actin cytoskeleton or intracellular tension [ 126 ]. The same was observed for Rac1/ β -catenin signaling [ 91 ].…”

The role of the cytoskeleton in sensing changes in gravity by nonspecialized cells

“…This particular model is based on observed changes in activity of Rho‐a after 7 d of exposure to simulated microgravity (52). More recently, Wan et al (87) showed changed Rho‐a and β‐catenin signaling after 1 and 2.5 h, respectively, of simulated microgravity on a slowly revolving clinostat. Furthermore, their experiments using actin‐disrupting drugs suggest that β‐catenin signaling is independent of actin cytoskeleton structure.…”

Different bone remodeling levels of trabecular and cortical bone in response to changes in Wnt/β‐catenin signaling in mice

“…Accumulating evidence showed that Wnt/β‐catenin signaling pathway plays a central role in bone development, bone growth and bone remodeling . Thus we hypothesized that the expression levels of molecules in the Wnt/β‐catenin signaling pathway in the cortical and trabecular compartments are different and that the different expression levels lead to different bone remodeling levels between cortical and trabecular bone.…”

“…2 Accumulating evidence showed that Wnt/b-catenin signaling pathway plays a central role in bone development, bone growth and bone remodeling. 11,[17][18][19][20][21][22][23][24][25] Thus we hypothesized that the expression levels of molecules in the Wnt/b-catenin signaling pathway in the cortical and trabecular compartments are different and that the different expression levels lead to different bone remodeling levels between cortical and trabecular bone. In the present study, the transcriptional and protein expression levels of Wnt/b-catenin signaling-related molecules were examined in the metaphyseal trabecular and diaphyseal cortical bone of the tibia in mice.…”