CaMKII Signaling Stimulates Mef2c Activity In Vitro but Only Minimally Affects Murine Long Bone Development in vivo.

Chandra S Amara,Lena I Wolff,Astrid Houben,Christine Hartmann,Christine Fabritius

doi:10.3389/fcell.2017.00020

Chandra S Amara, Lena I Wolff + Show 3 more

Open Access

https://doi.org/10.3389/fcell.2017.00020

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Abstract

The long bones of vertebrate limbs form by endochondral ossification, whereby mesenchymal cells differentiate into chondrogenic progenitors, which then differentiate into chondrocytes. Chondrocytes undergo further differentiation from proliferating to prehypertrophic, and finally to hypertrophic chondrocytes. Several signaling pathways and transcription factors regulate this process. Previously, we and others have shown in chicken that overexpression of an activated form of Calcium/calmodulin-dependent kinase II (CaMKII) results in ectopic chondrocyte maturation. Here, we show that this is not the case in the mouse. Although, in vitro Mef2c activity was upregulated by about 55-fold in response to expression of an activated form of CaMKII (DACaMKII), transgenic mice that expressed a dominant-active form of CaMKII under the control of the Col2a1 regulatory elements display only a very transient and mild phenotype. Here, only the onset of chondrocyte hypertrophy at E12.5 is accelerated. It is also this early step in chondrocyte differentiation that is temporarily delayed around E13.5 in transgenic mice expressing the peptide inhibitor CaM-KIIN from rat (rKIIN) under the control of the Col2a1 regulatory elements. Yet, ultimately DACaMKII, as well as rKIIN transgenic mice are born with completely normal skeletal elements with regard to their length and growth plate organization. Hence, our in vivo analysis suggests that CaMKII signaling plays a minor role in chondrocyte maturation in mice.

Highlights

Endochondral ossification is the process underlying the formation of the long bones in the vertebrate limbs (Erlebacher et al, 1995; Kronenberg, 2003)
Similar to what was observed in chicken, β-catenin was localized in the nucleus in mouse chondrocytes transfected with a DaCaMKII::GFP expression plasmid (Figure 1B, white arrows)
OSE2-luc reporter activity was upregulated in response to increasing amounts of DaCaMKII, while expression of rKIIN led to a downregulation of the reporter activity in a concentration independent manner (Figure 1C)

Summary

Introduction

Endochondral ossification is the process underlying the formation of the long bones in the vertebrate limbs (Erlebacher et al, 1995; Kronenberg, 2003). It starts with the condensation of mesenchymal cells that undergo chondrogenic differentiation, forming a cartilage template consisting of immature chondrocytes. These produce an extracellular matrix composed of proteoglycans, glycosaminoglycans, and glycoproteins. This template, which prefigures the future skeletal element, enlarges through chondrocyte proliferation (Akiyama and Lefebvre, 2011).

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