The role of annexin 2 in osteoblastic mineralization

Abstract

While the basic cellular contributions to bone differentiation and mineralization are widely accepted, the regulation of these processes at the intracellular level remains inadequately understood. Our laboratory recently identified annexin 2 as a protein involved in osteoblastic mineralization. Annexin 2 was overexpressed twofold in SaOSLM2 osteoblastic cells as a fusion protein with green fluorescent protein. The overexpression of annexin 2 led to an increase in alkaline phosphatase activity as well as an increase in mineralization. Our data suggest that the increase in alkaline phosphatase activity does not result from increased alkaline phosphatase transcript or protein levels; therefore we evaluated mechanism of action. We determined that both annexin 2 and alkaline phosphatase activity were localized to membrane microdomains called lipid rafts in osteoblastic cells. Annexin 2 overexpression resulted in an increase in alkaline phosphatase activity that was associated with lipid microdomains in a cholesterol-dependent manner. Furthermore, disruption of lipid rafts with a cholesterol sequestering agent or reduction of annexin 2 expression by specific antisense oligonucleotides each resulted in diminished mineralization. Therefore, intact lipid rafts containing annexin 2 appear to be important for alkaline phosphatase activity and may facilitate the osteoblastic mineralization process.

Lipid Segregation and Membrane Budding Induced by the Peripheral Membrane Binding Protein Annexin A2*

“…A scaffolding function of AnxA2 that can cluster PI(4,5)P 2 and other interacting lipids into larger domains is likely to be of relevance in several membrane related signaling and transport events. These include the formation of actin assembly sites at the plasma membrane (14,47), the organization of lipid domains (rafts) in sarcolemmal differentiation and during osteoblastic mineralization (56,57), and the formation of rafts required for efficient exocytosis of dense core granules in neuroendocrine cells (16). Thus, the novel properties reported here for AnxA2 and its derivatives using artificial membrane systems could represent the molecular basis of AnxA2 activities observed in cells.…”

Fetuin-A uptake in bovine vascular smooth muscle cells is calcium dependent and mediated by annexins

“…If either fetuin-A is not carrying calcium, or annexins are blocked, then neither calcium nor fetuin-A enters the cells. This is plausible in that annexins are known to facilitate membrane conformational changes and the formation of lipid rafts that facilitate endocytosis in osteoblasts (7). Several studies have demonstrated that annexins are present on endosomal compartments and involved in endocytosis (6).…”

Inactivation of anoctamin-6/Tmem16f, a regulator of phosphatidylserine scrambling in osteoblasts, leads to decreased mineral deposition in skeletal tissues

“…(46)(47)(48)(49)(50) Furthermore, at least in hypertrophic chondrocytes, the release of mineralization-competent matrix vesicles coincides with the translocation of PS from the inner to the outer leaflet of the plasma membrane. (47) This process, denoted as phospholipid scrambling, is regulated by members of the phospholipid scramblase protein family.…”