Molecular Genetics and Genomics

, Volume 284, Issue 4, pp 273–287 | Cite as

Antler development and coupled osteoporosis in the skeleton of red deer Cervus elaphus: expression dynamics for regulatory and effector genes

  • Viktor Stéger
  • Andrea Molnár
  • Adrienn Borsy
  • István Gyurján
  • Zoltán Szabolcsi
  • Gábor Dancs
  • János Molnár
  • Péter Papp
  • János Nagy
  • László Puskás
  • Endre Barta
  • Zoltán Zomborszky
  • Péter Horn
  • János Podani
  • Szabolcs Semsey
  • Péter Lakatos
  • László Orosz
Original Paper

Abstract

Antlers of deer display the fastest and most robust bone development in the animal kingdom. Deposition of the minerals in the cartilage preceding ossification is a specific feature of the developing antler. We have cloned 28 genes which are upregulated in the cartilaginous section (called mineralized cartilage) of the developing (“velvet”) antler of red deer stags, compared to their levels in the fetal cartilage. Fifteen of these genes were further characterized by their expression pattern along the tissue zones (i.e., antler mesenchyme, precartilage, cartilage, bone), and by in situ hybridization of the gene activities at the cellular level. Expression dynamics of genes col1A1, col1A2, col3A1, ibsp, mgp, sparc, runx2, and osteocalcin were monitored and compared in the ossified part of the velvet antler and in the skeleton (in ribs and vertebrae). Expression levels of these genes in the ossified part of the velvet antler exceeded the skeletal levels 10–30-fold or more. Gene expression and comparative sequence analyses of cDNAs and the cognate 5′ cis-regulatory regions in deer, cattle, and human suggested that the genes runx2 and osx have a master regulatory role. GC–MS metabolite analyses of glucose, phosphate, ethanolamine-phosphate, and hydroxyproline utilizations confirmed the high activity of mineralization genes in governing the flow of the minerals from the skeleton to the antler bone. Gene expression patterns and quantitative metabolite data for the robust bone development in the antler are discussed in an integrated manner. We also discuss the potential implication of our findings on the deer genes in human osteoporosis research.

Keywords

Antler cycle Antler microarray Physiological osteoporosis Mineralization genes Red deer Osteocalcin Runx2 Osx 

Abbreviations

AGC

Antler growth center

BMD

Bone mineral density

BMP

Bone morphogenetic protein

BGLAP

Human ortholog of osteocalcin

CGRP

Calcitonin gene-related peptide

Col1A1

Collagen alpha-1(I) chain

CVA

Canonical variates analysis/discriminant analysis

ECM

Extracellular matrix

FGF2

Fibroblast growth factor 2

GC–MS

Gas chromatography–mass spectrometry

Oc

Osteocalcin, human ortholog BGLAP

Osx

Osterix

PBS

Phosphate-buffered saline

PCA

Principal components analysis

PNP

Non-osteoporotic patients

PP

Patients affected with age-related osteoporosis

PTH

Parathyroid hormone

PTHrP

Parathyroid hormone-related protein

Runx2

Runt-related transcription factor 2

TGF

Transforming growth factor

TGFβ

Transforming growth factor beta

Notes

Acknowledgments

The authors are indebted to Magdolna Tóth Péli, Csilla Sánta Török, Kornélia Szóráth Gálné for excellent technical assistance, to MSc students András Berta and Attila Hegedűs (ELTE, Budapest) for enthusiastic help. Thanks are due to Péter Orosz and Natalia Polgár for critical reading of the manuscript, and to Sankar Adhya (NIH, Bethesda), László Sugár (U. Kaposvár), Ernő Duda (BRC, Szeged) and János Szabad (U. Szeged), Sándor Spisák, Bernadett Balla and János Kósa (SOTE, Budapest) for their constant interest. This work was supported by grants OTKA T032205 to L.O., OM 0320/2004 to L.O., NKFP 1A/007/2004 to L.O. and P.L., 454/2003 and 006/2009 from the Ministry of Health, Social and Family Affairs, ETT-ESKI to L.O., OTKA PD75496 to S.S., and by the János Bolyai fellowship of the HAS to S.S.

Supplementary material

438_2010_565_MOESM1_ESM.tif (692 kb)
ESM Fig. S1 (A) Antlerogenesis cycle of red deer stag, Cervus elaphus , (B) Bone structure at three time of the antler cycle : cross sections of flying ribs (left), corresponding scanning electron microscopic picture (right) (TIFF 691 kb)
438_2010_565_MOESM2_ESM.tif (328 kb)
ESM Fig. S2 Northern blot analyses of gene expressions. Antler tissues: FC foetal cartilage , RM reserve mesenchyme, PC precartilage , C cartilage , AB antler bone; Skeletal bones : VB vertebral bone , RB rib bone, ibsp* example for longer exposition. Note the robust expressions in AB versus VB and RB and the cartilage specific expression of col2A1. Stag 1 was in velvet antler development and skeletal osteoporosis status; Stag 2, in the period of late autumn dwell status when mineral mobilization and deposition are dynamically equilibrated – BMD is in steady state. Stag 3, in the velvet shedding, skeletal regenerating status. (For more information see Materials and Methods, ESM Fig. S1 and Borsy et al. 2009) (TIFF 327 kb)
438_2010_565_MOESM3_ESM.tif (418 kb)
ESM Fig. S3In situ hybridization of gene expressions for col3A1 and col10A1 (mRNA-s) in velvet antler tissues. Analyses of 8 μm thick longitudinal sections (RM) from mesenchymal region, (PC) from precartilage, (C) from cartilage. Note the very specific expression of col10A1 in hiperthrophic cartilage cells and the prominent expression of col3A1 in the mesenchyme and cartilage cells. Bars: 0.25 mm and 0.1 mm in the corners lower left (the higher magnifications). (TIFF 418 kb)
438_2010_565_MOESM4_ESM.doc (118 kb)
ESM Fig. S4 The DNA sequences for the 1 Kb promoter regions and the 1st exons plus 1245 intronic region of orthologs col1A1 genes of human, bovine and deer with Runx2 binding sites underlined. (DOC 117 kb)
438_2010_565_MOESM5_ESM.doc (104 kb)
ESM Fig. S5 The DNA sequences for the 1 Kb promoter regions and the 1st exons plus 1245 intronic region of orthologs col1A1 genes of human, bovine and deer with Osterix binding sites underlined. (DOC 104 kb)
438_2010_565_MOESM6_ESM.doc (34 kb)
Supplementary material 6 (DOC 35 kb)

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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Viktor Stéger
    • 1
    • 2
  • Andrea Molnár
    • 1
    • 2
    • 9
  • Adrienn Borsy
    • 1
    • 2
    • 10
  • István Gyurján
    • 1
    • 2
  • Zoltán Szabolcsi
    • 1
    • 2
  • Gábor Dancs
    • 2
  • János Molnár
    • 6
  • Péter Papp
    • 2
  • János Nagy
    • 3
  • László Puskás
    • 4
  • Endre Barta
    • 2
    • 8
  • Zoltán Zomborszky
    • 3
  • Péter Horn
    • 3
  • János Podani
    • 7
  • Szabolcs Semsey
    • 1
  • Péter Lakatos
    • 5
  • László Orosz
    • 1
    • 2
  1. 1.Department of GeneticsEötvös Loránd UniversityBudapestHungary
  2. 2.Institute of GeneticsAgricultural Biotechnology CenterGödöllőHungary
  3. 3.Department of Fish and Pet Animal Breeding, Faculty of Animal ScienceUniversity of KaposvárKaposvárHungary
  4. 4.Laboratory of Functional Genomics, Biological Research CenterHungarian Academy of SciencesSzegedHungary
  5. 5.1st Department of Internal MedicineSemmelweis UniversityBudapestHungary
  6. 6.BIOMI Ltd.GödöllőHungary
  7. 7.Department of Plant Taxonomy and EcologyEötvös Loránd UniversityBudapestHungary
  8. 8.Apoptosis and Genomics Research Group of the Hungarian Academy of SciencesUniversity of DebrecenDebrecenHungary
  9. 9.Institute of Experimental Medicine of the Hungarian Academy of SciencesBudapestHungary
  10. 10.Institute of BiochemistryBiological Research Center of the Hungarian Academy of SciencesSzegedHungary

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