Two-hybrid analysis of human salivary mucin MG2 (MUC7) interactions

Abstract

Saliva has been long associated with the maintenance of the oral health. The complex mixture of salivary constituents provides an effective set of systems for lubricating and protecting the soft and hard tissues. Among the salivary constituents responsible for these functions are the mucus glycoproteins. It is generally accepted that mucins in saliva occur in two different molecular weight forms: a low molecular weight (MG2) and a high molecular weight (MG1). MG2 is the low molecular weight monomeric mucin secreted by submandibular, sublingual and minor salivary glands. This mucin has been implicated in the non-immune host defense system in the oralcavity since it binds, agglutinates and exerts direct microbicidal effects on a variety of oral bacteria and fungi. MG2 has been also implicated in the formation of heterotypic complexes with other proteins present in salivary secretions such as secretory IgA and lactoferrin. Interactions between proteins occur in many levels within cells and tissues as welI as in glandular secretions and other body fluids under physiological conditions and might result in modified biological activity. To investigate interactions, a submandibular gland prey library was screened with baits encoding the N- and C-terminal regions of MG2 using the yeast two-hybrid system. The N-terminal region interacted with a group of secretory salivary proteins whereas the C-terminal region did not. Interacting proteins included amylase, acidic proline-rich protein 2, basic proline-rich protein 3, lacrimal proline-rich protein 4, statherin and histatin 1.Formation of complexes between these proteins and the N-terminal region of MG2 was confirmed in Far Western blotting experiments and co-immunoprecipitation studies. These results show that protein-protein interactions occur between mucin and non-mucin proteins in the secretory salivary protein "complexome". Interactions could protect complex partners from proteolysis, modulate the biological activity of complexed proteins or serve as a delivery system for distribution of secretory salivary proteins throughout the oral cavity.