Biological stenciling of mineralization in the skeleton: Local enzymatic removal of inhibitors in the extracellular matrix

September 30, 2020

N. Reznikov(a), B. Hoac(b), D.J. Buss(c), W.N. Addison(d), N.M.T. Barros(e), M.D. McKee(b,c)
Bone, Volume 138, September 2020, 115447 (


Mineralized tissues: Bone; Phosphate-regulating endopeptidase homolog X-linked (PHEX); Tissue-nonspecific alkaline phosphatase (TNAP, TNSALP, ALPL); Pyrophosphate; Osteopontin; Stenciling principle; Hypophosphatasia; X-linked hypophosphatemia; Review; Mineralization; Biomineralization


Biomineralization is remarkably diverse and provides myriad functions across many organismal systems. Biomineralization processes typically produce hardened, hierarchically organized structures usually having nanostructured mineral assemblies that are formed through inorganic-organic (usually protein) interactions. Calcium‑carbonate biomineral predominates in structures of small invertebrate organisms abundant in marine environments, particularly in shells (remarkably it is also found in the inner ear otoconia of vertebrates), whereas calcium-phosphate biomineral predominates in the skeletons and dentitions of both marine and terrestrial vertebrates, including humans. Reconciliation of the interplay between organic moieties and inorganic crystals in bones and teeth is a cornerstone of biomineralization research. Key molecular determinants of skeletal and dental mineralization have been identified in health and disease, and in pathologic ectopic calcification, ranging from small molecules such as pyrophosphate, to small membrane-bounded matrix vesicles shed from cells, and to noncollagenous extracellular matrix proteins such as osteopontin and their derived bioactive peptides. Beyond partly knowing the regulatory role of the direct actions of inhibitors on vertebrate mineralization, more recently the importance of their enzymatic removal from the extracellular matrix has become increasingly understood. Great progress has been made in deciphering the relationship between mineralization inhibitors and the enzymes that degrade them, and how adverse changes in this physiologic pathway (such as gene mutations causing disease) result in mineralization defects. Two examples of this are rare skeletal diseases having osteomalacia/odontomalacia (soft bones and teeth) – namely hypophosphatasia (HPP) and X-linked hypophosphatemia (XLH) – where inactivating mutations occur in the gene for the enzymes tissue-nonspecific alkaline phosphatase (TNAP, TNSALP, ALPL) and phosphate-regulating endopeptidase homolog X-linked (PHEX), respectively. Here, we review and provide a concept for how existing and new information now comes together to describe the dual nature of regulation of mineralization – through systemic mineral ion homeostasis involving circulating factors, coupled with molecular determinants operating at the local level in the extracellular matrix. For the local mineralization events in the extracellular matrix, we present a focused concept in skeletal mineralization biology called the Stenciling Principle – a principle (building upon seminal work by Neuman and Fleisch) describing how the action of enzymes to remove tissue-resident inhibitors defines with precision the location and progression of mineralization.

Author Affiliation

(a) Object Research Systems Inc., 760 St. Paul West, Montreal, Quebec H3C 1M4, Canada
(b) Faculty of Dentistry, McGill University, 3640 University St., Montreal, Quebec H3A 0C7, Canada
(c) Department of Anatomy and Cell Biology, McGill University, 3640 University St., Montreal, Quebec H3A 0C7, Canada
(d) Department of Molecular Signaling and Biochemistry, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka, Japan
(e) Departamento de Biofísica, São Paulo, Departamento de Ciências Biológicas, Universidade Federal de São Paulo, Diadema, Brazil