Histology and Embryology Program, Department of Morphology, Piracicaba Dental School, State University of Campinas, Piracicaba, SP, Brazil.
J Appl Oral Sci. 2010 Sep-Oct;18(5):482-6. doi: 10.1590/s1678-77572010000500009.
PAX9 belongs to the Pax family of transcriptional factor genes. This gene is expressed in embryonic tissues such as somites, pharyngeal pouch endoderm, distal limb buds and neural crest-derived mesenchyme. Polymorphisms in the upstream promoter region of the human PAX9 have been associated with human non-syndromic tooth agenesis. In the present study, we verified the in vitro mRNA expression of this gene and the luciferase activity of two constructs containing promoter sequences of the PAX9 gene.
Embryonic tissues were obtained from digits, face, and midbrain/hindbrain regions. Fragments containing PAX9 promoter sequences were cloned into reporter plasmids and were transfected into the different cell cultures. mRNA were extracted from primary cell cultures.
The semi-quantitative RT-PCR results showed that in vitro E13.5 limb bud and CNS cells express PAX9, but cells derived from the facial region do not. Moreover, the luciferase assay showed that protein activity of the constructed vector was weaker than pgl3 -basic alone.
The present results suggest that the promoter sequences analyzed are not sufficient to drive PAX9 gene transcription.
PAX9 属于 Pax 转录因子基因家族。该基因在胚胎组织中表达，如体节、咽囊内胚层、远端肢芽和神经嵴衍生的间充质。人类 PAX9 上游启动子区域的多态性与人类非综合征性牙齿缺失有关。在本研究中，我们验证了该基因的体外 mRNA 表达和包含 PAX9 基因启动子序列的两个构建体的荧光素酶活性。
胚胎组织取自指、面和中脑/后脑区域。含有 PAX9 启动子序列的片段被克隆到报告质粒中，并转染到不同的细胞培养物中。从原代细胞培养物中提取 mRNA。
半定量 RT-PCR 结果表明，体外 E13.5 肢芽和中枢神经系统细胞表达 PAX9，但来自面部区域的细胞不表达。此外，荧光素酶测定表明构建载体的蛋白活性比单独的 pgl3-basic 弱。
本研究结果表明，分析的启动子序列不足以驱动 PAX9 基因转录。
