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キーワード:
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要旨:
Matrix metalloproteases (MMPs) play a crucial role in tissue homeostasis. Profuse literature has studied their roles in cell migration and tissue invasion during cancer metastasis, as well as in inflammatory processes. Although the literature covering MMPs function is abundant, the intracellular trafficking of these proteins remains poorly understood. The aim of the present work was to identify the molecular mechanism of intracellular trafficking of MMPs, with particular focus on MMP2. A novel mass spectrometry approach revealed nucleobindin-1 (NUCB1), a major regulator of Ca2+ homeostasis at the Golgi, as a potential candidate for the regulation of MMP transport. Using a synchronized cargo trafficking assay, it was possible to demonstrate that in the absence of NUCB1 the intracellular trafficking of MMP2 is delayed. Moreover, this work reveals that NUCB1-dependent MMP2 trafficking is restricted to the Golgi, exclusively delaying its intra-Golgi trafficking at the cis compartment and, as a consequence, decreasing MMP2 mediated cell migration and matrix invasion. Furthermore, my findings show that not only MMP2, but also MT1-MMP intra-Golgi trafficking is impaired, implying that this mechanism could also influence the trafficking of other MMPs. Interestingly, experiments performed with a NUCB1 Ca2+-binding deficient mutant showed that Ca2+ is required, both for the interaction, as well as for proper MMP2 trafficking, suggesting that a specific impairment of cis-Golgi Ca2+ homeostasis, rather than an overall Ca2+ deficiency, is essential for proper MMP2 intra-Golgi trafficking. Taken together the results of this thesis contributed to enlighten the mechanism of MMP2 intracellular trafficking by identifying NUCB1 as a critical player in MMP transport. Importantly, this work highlights the requirement of Ca2+ for proper trafficking, not just at the TGN, as has been documented, but also at the cis-Golgi. Although this is a big step towards the understanding of MMP intracellular trafficking, further investigations are required to gain a better understanding of the retention mechanism of NUCB1 at the cis-Golgi lumen and a deeper insight into the regulation of intra-Golgi protein trafficking.
