Dataset Information

MMP10 degradomics in keratinocytes

ABSTRACT: Matrix metalloproteinases (MMPs) are important players in skin homeostasis, wound repair, and in the pathogenesis of skin cancer. One member – MMP10 – is specifically expressed in wound keratinocytes and in epithelial cancer cells, but its function is unclear and epidermal substrates are poorly characterized. To unravel the role of MMP10 in the skin, we employed Terminal Amine Isotopic Labeling of Substrates (TAILS), an iTRAQ-based quantitative proteomics technique for the discovery of protease substrates and their cleavage sites, to monitor MMP10-dependent proteolysis over time in secretomes from keratinocytes. By time-resolved abundance clustering of neo-N termini we revealed a cleavage site specificity preference of MMP10 for glutamate in the P1 position and identified the α6 integrin subunit, cysteine-rich angiogenic inducer 61 and dermokine as novel MMP10 substrates. Moreover, we confirmed the same MMP10-dependent processing of dermokine in vivo by TAILS analysis of epidermis from transgenic mice that overexpress a constitutively active mutant of MMP10 in basal keratinocytes. Since these substrates have been associated with cell adhesion, differentiation and senescence, MMP10 might contribute to modulation of these processes during skin repair.

INSTRUMENT(S): LTQ Orbitrap Velos, Q Exactive

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Cell Culture, Keratinocyte, Skin

SUBMITTER: Ulrich auf dem Keller

LAB HEAD: Ulrich auf dem Keller

PROVIDER: PXD002474 | Pride | 2015-10-19

REPOSITORIES: Pride

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Publications

Matrix Metalloproteinase 10 Degradomics in Keratinocytes and Epidermal Tissue Identifies Bioactive Substrates With Pleiotropic Functions.

Schlage Pascal P Kockmann Tobias T Sabino Fabio F Kizhakkedathu Jayachandran N JN Auf dem Keller Ulrich U

Molecular & cellular proteomics : MCP 20151016 12

Matrix metalloproteinases (MMPs) are important players in skin homeostasis, wound repair, and in the pathogenesis of skin cancer. It is now well established that most of their functions are related to processing of bioactive proteins rather than components of the extracellular matrix (ECM). MMP10 is highly expressed in keratinocytes at the wound edge and at the invasive front of tumors, but hardly any non-ECM substrates have been identified and its function in tissue repair and carcinogenesis is ...[more]

PMID: 26475864

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Project description:Proteases control complex tissue responses by modulating inflammation, cell proliferation and migration, and matrix remodeling. All these processes are orchestrated in cutaneous wound healing to restore the skinâ€™s barrier function upon injury. Altered protease activity has been implicated in the pathogenesis of healing impairments, and proteases are important targets in diagnosis and therapy of this pathology. Global assessment of proteolysis at critical turning points after injury will define crucial events in acute healing that might be disturbed in healing disorders. As optimal biospecimens, wound exudates contain an ideal proteome to detect extracellular proteolytic events, are non-invasively accessible, and can be collected at multiple time points along the healing process from the same wound in the clinics. In this study, we applied multiplexed Terminal Amine Isotopic Labeling of Substrates (TAILS) to globally assess proteolysis in early phases of cutaneous wound healing. By quantitative analysis of proteins and protein N termini in wound fluids from a clinically relevant pig wound model, we identified more than 650 proteins and discerned major healing phases through distinctive abundance clustering of markers of inflammation, granulation tissue formation, and re-epithelialization. TAILS revealed a high degree of proteolysis at all time points after injury by detecting almost 1300 N-terminal peptides in ~450 proteins, most of which could not be assigned to known mature protein N termini. Quantitative positional proteomics mapped pivotal interdependent processing events in the blood coagulation cascade, detailed activating thrombin cleavages in vivo, and temporally discerned clotting and fibrinolysis during the healing process. Similarly, we found virtually all major cleavages in complement activation and inactivation and demonstrated time-dependent changes in the proteolytic potential of the wound milieu by detecting processing of complement C3 at distinct time points after wounding and by different proteases.

			Action	DRS
	20110812_04_PS_PO3.mgf	Mgf
	20110812_04_PS_PO3.raw	Raw
	20110812_05_PS_PO4.mgf	Mgf
	20110812_05_PS_PO4.raw	Raw
	20110812_06_PS_PO5.mgf	Mgf