Gelatinases Cleave Dentin Sialoprotein Intracellularly

Area of Science:

• Biochemistry
• Cell Biology
• Dentinogenesis

Background:

• Dentin sialoprotein (DSP), a key component of dentin, is derived from dentin sialophosphoprotein (DSPP).
• DSP undergoes intracellular processing within odontoblasts, the cells responsible for dentin formation.
• Extracellular cleavage of DSPP by gelatinases (MMP2, MMP9) in tooth structures is known.

Purpose of the Study:

• To investigate the hypothesis that gelatinases cleave DSP intracellularly within odontoblasts.
• To elucidate the role of FURIN in the activation of MMP9 and subsequent DSP processing.

Main Methods:

• Co-expression and interaction analysis using double immunofluorescence and in situ proximity ligation assay (PLA).
• Assessment of intracellular gelatinase activity via gelatin zymography and in situ zymography.
• Western blot analysis of odontoblastic cell lysates treated with MMP inhibitors or from Mmp9 knockout cells.
• Investigation of FURIN-MMP9 interaction and proMMP9 cleavage in HEK293E cells.

Main Results:

• DSP was shown to co-localize and physically interact with MMP2 and MMP9 within odontoblasts.
• Active gelatinase activity was detected intracellularly, and DSP cleavage was confirmed by western blotting.
• FURIN was found to co-localize and interact with MMP9, and its inhibition increased proMMP9 levels.
• FURIN was demonstrated to cleave proMMP9 at the R⁹⁶TPR⁹⁹ motif, activating it for intracellular processing of DSP.

Conclusions:

• DSP is intracellularly processed by gelatinases (MMP2, MMP9) in odontoblasts.
• FURIN plays a critical role in the intracellular activation of proMMP9 via cleavage of its R⁹⁶TPR⁹⁹ motif.
• This study reveals a novel mechanism for DSP processing and MMP9 activation during dentinogenesis.