DMP1 mutations in autosomal
implicate a bone matrix protein
in the regulation of phosphate
Bettina Lorenz-Depiereux1,12, Murat Bastepe2,12,
Anna Benet-Page `s1, Mustapha Amyere3, Janine Wagenstaller1,
Ursula Mu ¨ller-Barth4, Klaus Badenhoop5, Stephanie M Kaiser6,
Roger S Rittmaster6, Alan H Shlossberg6, Jose ´ L Olivares7,
Ce ´sar Loris8, Feliciano J Ramos7, Francis Glorieux9,
Miikka Vikkula3, Harald Ju ¨ppner2,10& Tim M Strom1,11
Hypophosphatemia is a genetically heterogeneous disease.
Here, we mapped an autosomal recessive form (designated
ARHP) to chromosome 4q21 and identified homozygous
mutations in DMP1 (dentin matrix protein 1), which encodes
a non-collagenous bone matrix protein expressed in osteoblasts
and osteocytes. Intact plasma levels of the phosphaturic
protein FGF23 were clearly elevated in two of four affected
individuals, providing a possible explanation for the
phosphaturia and inappropriately normal 1,25(OH)2D levels
and suggesting that DMP1 may regulate FGF23 expression.
Despite its broad biological importance, the control of phosphate
homeostasis remains incompletely understood. Most of the genes
contributing to its normal regulation have been identified by studying
genetic defects leading to different hypophosphatemic disorders. The
most frequent of these disorders is X-linked hypophosphatemia (XLH;
OMIM 307800), caused by inactivating mutations in a gene encoding
a putative endopeptidase (PHEX)1. A much rarer disorder, autosomal
dominant hypophosphatemic rickets (ADHR; OMIM 193100) is
caused by mutations in FGF23 that render the phosphaturic factor
encoded by this gene resistant to proteolytic cleavage by subtilisin-like
proprotein convertases2,3. Wild-type FGF23 is abundantly expressed in
certain mesenchymal tumors, causing increased serum FGF23 levels
that induce renal phosphate-wasting and osteomalacia (tumor-
induced osteomalacia or TIO)4–6.
We have now investigated three multiplex families in which the
affected individuals showed clinical, biochemical and histomorpho-
metric parameters that were similar to those observed in XLH and
ADHR (Table 1 and Supplementary Methods online). However,
inspection of the pedigrees suggested an autosomal recessive mode
of inheritance (hence the abbreviation ARHP), thus excluding XLH
and ADHR. In order to identify the molecular defect, we performed a
genome-wide linkage analysis using SNP array genotyping. Assuming
that the disease alleles could be identical by descent in each family, we
analyzed the data by homozygosity mapping and identified a 4.6-Mb
candidate region on chromosome 4q21 between SNPs rs340204 and
rs722937. Parametric LOD score calculations under the conservative
assumption of second-cousin marriages in family 1 and 2 resulted in
maximum LOD scores of 3.1 and 2.4, respectively, and 4.2 in family 3
with established consanguinity.
The candidate region for ARHP contained a cluster of genes coding
for a class of tooth and bone noncollagenous matrix proteins that are
referred to as SIBLING proteins (small integrin-binding ligand,
N-linked glycoproteins). Members of this protein family include
dentin sialophosphoprotein (DSPP), dentin matrix protein 1
(DMP1), integrin-binding sialoprotein (IBSP), matrix extracellular
phosphoglycoprotein (MEPE) and osteopontin (also named secreted
phosphoprotein 1, SPP1). These polyanionic proteins are believed to
have key biological roles in mineralization of osteoid and dentin7and
thus were plausible candidates. We therefore searched by direct
sequencing for mutations in their exons and relevant flanking intronic
regions (except for the last exon of DSPP and MEPE).
In the affected members of all three investigated families, we
identified different homozygous, presumably loss-of-function muta-
tions in DMP1; each of the unaffected parents was heterozygous for
the respective mutation (Fig. 1 and Supplementary Fig. 1 online).
We observed only known sequence variations in DSPP and IBSP but
did not identify any variations in MEPE and SPP1 (Supplementary
Table 1 online). The affected siblings in family 1 carried a homozygous
1-bp deletion in exon 6 (362delC) leading to a premature stop codon
after 120 unrelated amino acids. The two affected brothers in family 2
had a homozygous mutation in the canonical splice acceptor sequence
(55-1G-C) of intron 2. The affected individuals in family 3 carried a
missense mutation in exon 2 that changed the initiator codon ATG to
GTG (1A-G, leading to M1V). Protein blot analysis of the super-
natant of HEK293 cells transiently expressing a plasmid encoding a
Received 1 May; accepted 3 August; published online 8 October 2006; doi:10.1038/ng1868
1Institute of Human Genetics, GSF National Research Center for Environment and Health, 85764 Munich-Neuherberg, Germany.2Endocrine Unit, Massachusetts
General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.3Human Molecular Genetics (GEHU), Christian de Duve Institute & Universite ´
catholique de Louvain, 1200 Brussels, Belgium.4Medical Genetics, 63450 Hanau, Germany.5Division of Endocrinology, Diabetes and Metabolism, University
Hospital, 60590 Frankfurt am Main, Germany.6Division of Endocrinology and Metabolism, Queen Elizabeth II Health Sciences Centre, Halifax, Nova Scotia, Canada.
7Department of Pediatrics, University Hospital Lozano Blesa, Faculty of Medicine, University of Zaragoza, 50009 Zaragoza, Spain.8Service of Nephrology, University
Children’s Hospital Miguel Servet, 50009 Zaragoza, Spain.9Shriners Hospital for Children, Genetics Unit, Montreal, Canada.10Pediatric Nephrology Unit,
Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.11Institute of Human Genetics, Klinikum rechts der Isar, Technical
University, 81675 Munich, Germany.12These authors contributed equally to this work. Correspondence should be addressed to T.M.S. (TimStrom@gsf.de) or
1248 VOLUME 38 [ NUMBER 11 [ NOVEMBER 2006 NATURE GENETICS
© 2006 Nature Publishing Group http://www.nature.com/naturegenetics
His-tagged DMP1 carrying this mutation showed a 23-kDa protein
band but not the 80-kDa and 57-kDa bands that were observed in cells
expressing His-tagged wild-type DMP1. Furthermore, we searched for
DMP1 mutations in 18 individuals with hypophosphatemia for
whom PHEX and FGF23 mutations had been excluded and in three
individuals with tumoral calcinosis for whom FGF23 and GALNT3
mutations had been excluded. In addition to several common
sequence variants (Supplementary Table 1), we identified a non-
synonymous, heterozygous variant (349G-A, D117N) in an indivi-
dual with tumoral calcinosis that was not present in 666 normal
DMP1 and its mouse and rat orthologs are expressed in tooth,
bone, brain and salivary gland. Targeted ablation of both Dmp1 alleles
in mice resulted in shorter bones and vertebrae, a highly expanded
zone of proliferating and hypertrophic chondrocytes in the growth
plate of younger mice and broad sclerotic long bones in older
animals8. These skeletal findings, which were initially thought to
represent a form of chondrodysplasia, led to the conclusion that
DMP1 is required for normal postnatal bone and tooth formation.
Recently, however, decreased serum phosphate and calcium levels have
been observed9, demonstrating the similarity of this phenotype with
different hypophosphatemia-induced forms of rickets.
Various in vitro and in vivo studies suggested that the genes mutated
in different forms of hypophosphatemia constitute a previously
unrecognized pathway involved in regulating phosphate homeostasis.
In brief, the endopeptidase PHEX seems to indirectly regulate levels of
the phosphaturic protein FGF23, which affects expression and inter-
nalization of the renal sodium-phosphate cotransporters SLC34A1
and SLC34A3. It is not yet known how DMP1 is involved in regulating
phosphate homeostasis. However, it has been shown that DMP1 is
processed at four different cleavage sites10and that cleavage can be
Table 1 Chemical and clinical data
Family 1 Family 2 Family 3Reference ranges
Individual12804 284291181027240 27243128
FGF23 (intact) (pg/ml)
FGF23 (C-t) (RU/ml)
U Ca/Cr (mg/mg)
Age (in years and months)
8 yrs 5 mo
15 yrs 9 mo
1 yr 10 mo
Age- and sex-dependent reference values are given in parentheses. FGF23 plasma levels were measured after 2 d of withdrawal of 1,25 vitamin D and phosphate substitution; for
some affected individuals, the results of two or more measurements are provided. FGF23: fibroblast growth factor 23, Ca: calcium, P: phosphorus, AP: alkaline phosphatase, PTH:
parathyroid hormone, 25OHD: 25-hydroxyvitamin D, 1,25(OH)2D: 1,25-dihydroxyvitamin D, U Ca/Cr: urine calcium/creatinine ratio, TRP: tubular reabsorption of phosphate, TmP/
GFR: maximal tubular phosphate reabsorption to glomerular filtration rate. The centiles for weight and height are given in parentheses.
aAt age 2 years 6 months. Chemical data for the heterozygous parents in Family 2 are shown in Supplementary Table 2 online.
Family 1Family 2 Family 3
12804 12805 28429 11810
c.1A>G, p.M1V c.55-1G>Cc.362delC
Figure 1 DMP1 mutations in three families with autosomal recessive
hypophosphatemia. All affected individuals were homozygous for the
mutated allele segregating in the corresponding family. The parents were
heterozygous for the respective mutation. Affected individuals are indicated
by filled symbols and heterozygous carriers by half-filled symbols.
Radiographs for affected members of each family are shown in
Supplementary Figure 3 online. Morphometric parameters of an iliac bone
biopsy specimen from an affected individual in family 3 are shown in
Supplementary Table 3 online. Primer sequences used for sequence
analysis are listed in Supplementary Table 4 online. Informed consent
was obtained from all study participants. The study was approved by the
institutional review boards of the Medical Department of the Technical
University of Munich and Massachusetts General Hospital.
NATURE GENETICS VOLUME 38 [ NUMBER 11 [ NOVEMBER 2006 1249
© 2006 Nature Publishing Group http://www.nature.com/naturegenetics
achieved by BMP1/tolloid-like proteinases in vitro11. Considering that Download full-text
PHEX and DMP1 are coexpressed (Supplementary Fig. 2 online) and
that PHEX and BMP1 require an aspartate at the P1¢ position of the
cleavage site12, it appeared conceivable that DMP1 might be processed
by PHEX10. To investigate this hypothesis, we expressed human
C-terminally tagged (DMP1/His) and mouse N-terminally tagged
(Flag/Dmp1) fusion proteins in HEK293 cells and incubated the
proteins with a recombinant secreted and soluble form of PHEX
(secPHEX)12. Under these in vitro conditions, none of the tagged
DMP1 or Dmp1 proteins showed specific cleavage or substantial
degradation in the presence of secPHEX (Supplementary Fig. 2).
However, we cannot exclude the possibility that intact DMP1 or
DMP1 fragments are PHEX substrates under physiological conditions.
Previous studies have described that PHEX binds to the C-terminal
part of MEPE, another member of the SIBLING family, without
cleaving it13. To investigate whether DMP1 binds to PHEX protein in
a similar manner, PHEX was incubated with a PHEX substrate,
PTHrP107–139(ref. 12). The addition of DMP1 or Dmp1 did not
alter the extent of degradation of PTHrP107–139by PHEX (Supple-
mentary Fig. 2), suggesting that physical interactions between DMP1
and PHEX do not affect PHEX activity.
DMP1 has been shown to be expressed more abundantly in TIO
tumors than FGF23 (ref. 5). However, unlike FGF23, DMP1 showed
no phosphaturic effect in vivo5and did not inhibit phosphate uptake
in vitro14, suggesting that it does not have a direct role in renal
phosphate handling. Because inactivating DMP1 mutations, as shown
in this report, lead to hypophosphatemia, we asked whether the lack of
DMP1 increases plasma or serum FGF23 levels as in individuals with
XLH, TIO and ADHR6. Plasma and serum were available from four
individuals with homozygous DMP1 mutations, and two indepen-
dently collected samples from two individuals showed clearly elevated
FGF23 levels when measured by an ELISA that detects intact FGF23
alone and showed slightly elevated levels in one of these individuals
when using an ELISA that detects C-terminal and intact FGF23
(Table 1). In the other two individuals, intact FGF23 was slightly
elevated or in the upper normal range. These findings are similar to
those observed in XLH, in which many but not all affected individuals
show elevated FGF23 levels. The phosphaturia observed in ARHP
could thus also be FGF23 dependent. As FGF23 also inhibits 1-alpha-
hydroxylase, elevated FGF23 concentrations could furthermore pro-
vide an explanation for the inappropriately normal 1,25(OH)2D levels
observed in all affected individuals with ARHP whom we investigated
(Table 1). The details of the underlying regulatory mechanisms are
not yet known. It is interesting in this regard that DMP1 has been
reported to have different biological roles. If phosphorylated, it is
exported into the extracellular matrix, where it regulates nucleation of
hydroxyapatite. Otherwise, it is transported to the nuclear compart-
ment, where it acts as a transcription factor15. With the identification
of DMP1 mutations as the cause of ARHP, we have added a further
component to the growing list of genes involved in the regulation of
Note: Supplementary information is available on the Nature Genetics website.
We thank the families for participation in this study. We also thank S. Lo ¨secke
for technical assistance and H. Murdock, T. Neuhaus, M. Seton and G. El-Hajj
Fuleihan for their help in collecting DNA samples. This work was supported by
a grant of the Deutsche Forschungsgemeinschaft (STR304/2-2) and the National
Institutes of Health (R21 DK075856-01) and in part by the Fondo de
Investigacio ´n Sanitaria (FIS) of the Spanish Ministry of Health (Research
Network Programs C03/07 and G03/097) and the European Foundation for
the Study of Diabetes.
COMPETING INTERESTS STATEMENT
The authors declare that they have no competing financial interests.
Published online at http://www.nature.com/naturegenetics
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1. The HYP Consortium. Nat. Genet. 11, 130–136 (1995).
2. The ADHR Consortium. Nat. Genet. 26, 345–348 (2000).
3. White, K.E. et al. Kidney Int. 60, 2079–2086 (2001).
4. White, K.E. et al. J. Clin. Endocrinol. Metab. 86, 497–500 (2001).
5. Shimada, T. et al. Proc. Natl. Acad. Sci. USA 98, 6500–6505 (2001).
6. Jonsson, K.B. et al. N. Engl. J. Med. 348, 1656–1663 (2003).
7. Qin, C., Baba, O. & Butler, W.T. Crit. Rev. Oral Biol. Med. 15, 126–136 (2004).
8. Ye, L. et al. J. Biol. Chem. 280, 6197–6203 (2005).
9. Ling, Y. et al. J. Bone Miner. Res. 20, 2169–2177 (2005).
10. Qin, C. et al. J. Biol. Chem. 278, 34700–34708 (2003).
11. Steiglitz, B.M., Ayala, M., Narayanan, K., George, A. & Greenspan, D.S. J. Biol. Chem.
279, 980–986 (2004).
12. Boileau, G., Tenenhouse, H.S., Desgroseillers, L. & Crine, P. Biochem. J. 355,
13. Rowe, P.S. et al. Bone 36, 33–46 (2005).
14. Bowe, A.E. et al. Biochem. Biophys. Res. Commun. 284, 977–981 (2001).
15. Narayanan, K. et al. J. Biol. Chem. 278, 17500–17508 (2003).
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