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Phenotypic Analysis of Dlx5 Overexpression in Post-natal Bone

J. Zhang^1,2,, J. Zhu^1,, P. Valverde¹, L. Li¹, S. Pageau¹, Q. Tu¹, R. Nishimura³, T. Yoneda³, P. Yang², W. Zheng⁴, W. Ma⁵, and J. Chen^1,*

¹ Division of Oral Biology, Department of General Dentistry, Tufts University School of Dental Medicine, One Kneeland Street, Boston, MA 02111, USA;
² College of Stomatology, Shandong University, Shandong Province, China;
³ Department of Biochemistry, Osaka University Graduate School of Dentistry, Osaka, Japan;
⁴ South Genomics Research Center, Guangzhou, 510800, China; and
⁵ Institute of Genetic Engineering, South Medical University, Guangzhou, 510515, China

^* corresponding author, jk.chen{at}tufts.edu

Dlx5 plays an important role in the embryonic development of mineralized tissues. We hypothesized that Dlx5 also functions in regulating post-natal bone formation in mice. To prove this hypothesis, we infected 5-day-old bone sialoprotein (BSP)/avian retroviral receptor gene (TVA) transgenic mice with replication-competent retroviral vectors expressing wild-type Dlx5 (RCAS-Dlx5WT) and mutated Dlx5 at arginine (R) 31 of its homeodomain (RCAS-Dlx5RH). Immunohistochemistry indicated that RCAS-Dlx5WT increased BSP and osteopontin (OPN) expression, whereas it decreased that of osteocalcin (OC). RCAS-Dlx5RH mediated opposite effects. Semi-quantitative RT-PCR confirmed these results. Ex vivo overexpression of RCAS-Dlx5WT in BSP/TVA calvarial cells promoted, whereas that of RCAS-Dlx5RH inhibited, mineralized nodule formation as compared with that in control cells. Our results suggest that Dlx5 promotes expression of early markers of osteogenic differentiation and increases mineralization post-natally.

KEY WORDS: Dlx5 (distal-less-related gene) • homeodomain • BSP/TVA transgenic mice • extracellular matrix proteins • mineralization