Methods:Human PLAG1 cDNA was cloned from salivary gland tumor or placenta tissues by RT-PCR. Ubiquitous expression vector pCMV-EGFP/PLAG1 driven by CMV promoter and tissue-specific expression vector pMMTV-PLAG1 driven by MMTV LTR were constructed. NIH3T3 cells transiently transfected with pCMV-EGFP/PLAG1 show high expression of PLAG1 in nucleus. Transgenes were microinjected into pronucleus of zygotes to generate transgenic mice.
Results:Human PLAG1 cDNA cloned from several salivary gland tumor and normal placenta tissues consistently show a variation of a single nucleotide at same position compared with the human PLAG1 cDNA sequence in Genbank (Accession No. U65002), which leads to T458P at amino acid level. It might be a Single Nucleotide Polymorphism (SNP) locus . Fused EGFP/PLAG1 protein is localized in nucleus of NIH3T3 cells transiently transfected with pCMV-EGFP/PLAG1. We got several pCMV-EGFP/PLAG1 and pMMTV-PLAG1 transgenic mouse lines, respectively. As we expected, pMMTV-PLAG1 transgenic mice spontaneously develop salivary gland tumors in three independent lines. Among them, line 42 shows tumorigenic phenotype in almost 100% of transgenic mice within three months after birth.
Conclusions:Overexpression of PLAG1 gene plays a crucial role in tumorigenesis of salivary gland tumors.