1561 Expression of FAM20A, causative gene for Amelogenesis Imperfecta in ameloblasts

Saturday, March 24, 2012: 9:45 a.m. - 11 a.m.
Presentation Type: Poster Session
Y. OHYAMA1, C. AN2, A. ALMEHMADI1, M. SNEAD3, S. KUMABE4, Y. IWAI2, H. HOTTA5, and Y. MOCHIDA1, 1Department of Periodontology and Oral Biology, Boston University, Henry M. Goldman School of Dental Medicine, Boston, MA, 2Department of Oral Anatomy, Osaka Dental University, Osaka, Japan, 3University of Southern California, Los Angeles, CA, 4Oral Anatomy, Osaka Dental University, Hirakata, Osaka, Japan, 5Division of Microbiology, Kobe University Graduate School of Medicine, Kobe, Japan
Objective: Patients with Amelogenesis Imperfecta (AI) caused by FAM20A mutation display several dental phenotypes including hypoplastic enamel, intrapulpal calcification, delayed tooth eruption, failure of tooth development and gingival hyperplasia. It has been initially reported that FAM20A is a member of three related “family with sequence similarity 20” proteins, i.e. FAM20A, FAM20B and FAM20C, with unknown function and proposed that they are secreted proteins. The localization of FAM20 proteins were re-examined and reported that the immunoreactivity of FAM20A was not clearly observed in the enamel matrix. To characterize the biological function of FAM20A, as a preliminary step, the present study is sought to investigate the expression and localization of endogenous Fam20a in an ameloblast cell line, LS8.

Method: The gene expression was analyzed using several mouse tissues by real time PCR. The expression and localization of Fam20a protein were examined using anti-Fam20a antibody by immunofluorescence-based staining and immunoprecipitation (IP) - Western Blot (WB) techniques in LS8 cells.

Result:  Fam20a was highly expressed in lung, liver and tooth at mRNA levels. The expression was also confirmed in the LS8 ameloblast cell line. The IP-WB analysis was performed using cultured media and cell lysates from LS8 cells and the protein expression was clearly observed in the cell lysate fraction. Cell lysates from LS8 cells were further fractionated and the WB analysis demonstrated that the endogenous Fam20a protein was present in membrane fraction. The subcellular localization of endogenous Fam20a protein was detected at the cytoplasm.

Conclusion: We demonstrated the intracellular expression of FAM20A in LS8 ameloblasts. Our data suggest that FAM20A is a novel intracellular modulator of enamel formation/mineralization. 

Supported by NIH grant DE019527 and Boston University School of Dental Medicine.

This abstract is based on research that was funded entirely or partially by an outside source: NIH grant DE019527

Keywords: Ameloblasts, Amelogenesis Imperfecta, Gene expression and Teeth