Mast Cell β-Tryptase Is Enzymatically Stabilized by DNA.

Sultan Alanazi,Fabio Rabelo Melo,Maria Lampinen,Gunnar Pejler,Christian P Sommerhoff,Ola Rollman,Mirjana Grujic

doi:10.3390/ijms21145065

Abstract

Tryptase is a tetrameric serine protease located within the secretory granules of mast cells. In the secretory granules, tryptase is stored in complex with negatively charged heparin proteoglycans and it is known that heparin is essential for stabilizing the enzymatic activity of tryptase. However, recent findings suggest that enzymatically active tryptase also can be found in the nucleus of murine mast cells, but it is not known how the enzmatic activity of tryptase is maintained in the nuclear milieu. Here we hypothesized that tryptase, as well as being stabilized by heparin, can be stabilized by DNA, the rationale being that the anionic charge of DNA could potentially substitute for that of heparin to execute this function. Indeed, we showed that double-stranded DNA preserved the enzymatic activity of human β-tryptase with a similar efficiency as heparin. In contrast, single-stranded DNA did not have this capacity. We also demonstrated that DNA fragments down to 400 base pairs have tryptase-stabilizing effects equal to that of intact DNA. Further, we showed that DNA-stabilized tryptase was more efficient in degrading nuclear core histones than heparin-stabilized enzyme. Finally, we demonstrated that tryptase, similar to its nuclear localization in murine mast cells, is found within the nucleus of primary human skin mast cells. Altogether, these finding reveal a hitherto unknown mechanism for the stabilization of mast cell tryptase, and these findings can have an important impact on our understanding of how tryptase regulates nuclear events.

Highlights

Mast cells are immune cells originating from the bone marrow [1]
When tryptase alone was incubated at room temperature in a neutral pH-buffer, we noted that it lost enzymatic activity in a time-dependent manner, with essentially a complete loss of activity seen after 6 h (Figure 1A)
When heparin was included in the incubation mixture, the enzymatic activity was completely preserved during the time interval studied; the addition of heparin potentiated the tryptase activity in comparison with the baseline level (Figure 1A)

Summary

Introduction

Mast cells are immune cells originating from the bone marrow [1] When they mature in local tissue environments, they acquire an abundance of secretory granules. These are filled with a plethora of preformed inflammatory mediators, including histamine, serotonin, cytokines, growth factors, lysosomal hydrolases, serglycin proteoglycan as well as large amounts of mast cell-restricted proteases [2]. Out of the mast cell proteases, β-tryptase (denoted tryptase in the following) is unique with respect to its tetrameric organization, where all of its active sites are facing a narrow, central pore [5]. It has been shown that heparin is necessary for stabilizing the tetrameric, active organization of tryptase, and that the absence of stabilizing heparin is accompanied by a rapid loss of enzymatic activity [12]

Methods

Results

Conclusion