Expression And Localization Of Dental Matrix Protein 1(DMP1) In Mouse Submandibular Glands

Dental matrix protein 1(DMP1),an acidic phosphoprotein,belongs to the family of small integrin-binding ligand,N-linked glycoprotein(SIBLING).It has been demonstrated that DMP1 may serve as an important regulator in the formation of tooth and bone.In addition to the mineralized tissues,DMP1 is also expressed in some unmineralized tissues,such as brain,pancreas,kidney and salivary glands.As the largest pair of salivary glands,mouse submandibular glands produce and secrete about 70 percent of the total saliva.Moreover,as a special type of conduct in mouse submandibular glands,granular convoluted tubule(GCT)is capable of secreting diverse kinds of bioactive peptides.Under the regulation of thyroid hormones and androgen,GCTs between male and female mouse submandibular glands have sexually dimorphic differences.In addition,GCTs appear in mouse submandibular glands at the postnatal 2 weeks.The above characteristics of GCTs differ from the other conducts in mouse submandibular glands.Although it has been demonstrated that DMP1 is expressed in mouse submandibular glands,the expression and localization of DMP1 fragments in mouse submandibular glands remains to be unknown.Recently,the facts that DMP1 is significantly elevated in some malignant tumors suggest that DMP1 may have other functions in addition to the regulation role in mineralized tissues.In this study,with multiple approaches including reverse transcription PCR(RT-PCR),quantitative real time PCR(q RT-PCR),Western immunoblotting,immunohistochemistry(IHC),in situ hybridization(ISH)and β-galactosidase(Lac Z)expression assay,we systematically analyzed the expression and localization of DMP1 fragment in mouse submandibular glands.The results showed that DMP1 was expressed in male and female mouse submandibular glands,but the level of DMP1 m RNA in mouse submandibular glands was lower than that in mouse incisors.In mouse submandibular glands,DMP1 existed as its proteolytically processed fragments including the N-terminal fragment of DMP1(DMP1-N)and the C-terminal fragment of DMP1(DMP1-C).In contrast to 57 kDa of DMP1-C and 37 kDa of DMP1-N in mineralized tissues,DMP1 was primarily present as the 46 kDa of DMP1-C and 37 kDa of DMP1-N.DMP1-C was mainly localized in the nuclei of acinar and ductal cells,while DMP1-N was restricted to the cytoplasm of ductal cells except for GCT cells.At the postnatal 4 weeks and 12 weeks,the mRNA levels of DMP1 in male mice were significantly higher than those in female mice.This study showed the expression of DMP1 in the GCTs of male mice with the experiments of qRT-PCR,IHC,ISH and β-galactosidase(Lac Z)expression assay,a novel finding different from the result of previous reports.Collectively,the differential localization patterns of DMP1 fragments indicate that different forms of DMP1 may play distinct roles in the submandibular glands such as transcriptional regulation of other genes and increasing invasion of tumor cells through the high affinity with matrix metalloproteinase 9.