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

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.

TRPM7 channels mediate spontaneous Ca 2+ fluctuations in growth plate chondrocytes that promote bone development

“…We attempted to address the signaling cascades activated downstream of TRPM7-mediated Ca 2+ fluctuations in growth plate chondrocytes. Ca 2+ entry generally triggers intracellular signaling by activating Ca 2+ -dependent enzymes, and previous studies suggest the involvement of Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) and calcineurin (CaN) in chondrogenesis (36)(37)(38). In our metatarsal culture system, the CaMKII inhibitors KN93 and cell-permeable autocamtide-2-related inhibitory peptide II (pAIP-II) dose-dependently inhibited bone outgrowth, whereas the CaN inhibitor FK506 exerted no significant effects at doses typically used for cell culture experiments (fig.…”

Loss of the WNT9a ligand aggravates the rheumatoid arthritis-like symptoms in hTNF transgenic mice

“…Whole protein lysates from SFBs were prepared as previously described 40 . 12.5 µg lysate was run on a 10% SDS-PAGE and transferred to a 0.45-μm PVDF-membrane by semi-dry transfer (PerfectBlue™, PeqLab).…”

Elucidation of the Physiological Functions of Membrane Proteins as Novel Drug Target Candidate Molecules

“…49) We crossed Trpm7 fl/fl mice with 11Enh-Cre mice, which express Cre recombinase under the control of the Col11a2 promoter and enhancer and thus delete loxP-flanked genes in a chondrocyte-specific manner. 50) Chondrocyte-specific knockout of Trpm7 resulted in significantly weakened spontaneous Ca 2+ fluctuations with reduced autophosphorylation of Ca 2+ /calmodulin-dependent protein kinase II (CaMKII), which is one of the major Ca 2+ -dependent enzymes that triggers downstream signals [51][52][53] (Figs. 3A, B).…”