Normal tissue repair involves inflammation, granulation tissue formation, and tissue remodeling. In chronic wounds, these events are disrupted, resulting in delay or lack of repair. Effective wound repair requires extracellular matrx (ECM) remodeling to allow cell migration and capillary morphogenesis. ECM remodeling and signaling cascades derived from this process dictate whether or to what extent tissue repair progresses or is chronically impaired, and remodeling depends on precisely controlled interactions between cells and the ECM. The matrix metalloproteinase (MMP) family regulates ECM remodeling. In concert, the integrin family of cell surface receptors mediates adhesive interactions between cells and the ECM. Thus, interaction and crosstalk between cells, MMPs, integrins, and other players determines remodeling and vascularization during wound healing. Current understanding of key transcription factors regulating ECM remodeling is limited. Our lab has discovered a novel role of the nuclear transcription co-factor, cardiac ankyrin repeat protein (CARP), in the formation and abundance of new blood vessels during wound healing. The central hypothesis of this application is that CARP protein regulates the transcription of genes coding for matrix-degrading proteases, cell adhesion molecules, and other genes associated with the process of ECM remodeling and capillary morphogenesis during the wound repair process. In support, deletion of CARP gene, ankrd1, results in decreased expression of 1 integrin subunit. Interestingly, expression of other integrin subunits does not change by lack of CARP. Furthermore, overexpression of CARP results in decreased MMP- 13 promoter activity, and deletion of CARP results in increased mRNA expression of MMP-13 in vitro and in vivo. Knockout of ankrd1 results in higher MMP-13 protein levels in unwounded skin and leads to a higher abundance of MMP-13 in mouse excisional wounds 4 days after injury. The goal of this application is to identify and characterize CARP target genes that are important for ECM remodeling during wound repair. To address this goal two Specific Aims are proposed.
Aim one will determine the involvement of CARP in regulation of matrix metalloproteinase-13 (MMP-13).
This aim will be achieved by identifying factors, e.g. nucleolin, that interact with CARP to regulate MMP-13 activity as well as characterizing how exactly CARP regulates MMP-13 using electrophoretic mobility shift assay (EMSA) and ChIP analysis.
Aim two is designed to delineate the role of CARP in regulation of 11 integrin. For this aim, a combination of in vitro studies and animal wound healing models will be used to investigate how CARP regulates the integrin 1 subunit and the consequences of this regulation. The proposed research will investigate the regulation of integrins and matrix metalloproteinases by the transcriptional nuclear co-factor, CARP. Understanding the mechanisms that CARP utilizes for its pro-angiogenic activities can lead to the development of a therapy that improves wound healing in chronic wounds, especially in diabetic patients where wound repair is severely compromised.
Data from cardiac ankyrin repeat protein (CARP) overexpression and deletion show CARP involvement in neovascularization, a critical aspect of the wound healing process that is impaired in the diabetic state. I propose to examine how CARP affects tissue repair through provisionally-identified targets such as integrins and matrix metalloproteinases. Results derived from this study can lead to the delineation of new mechanisms that regulate wound angiogenesis.
|AlmodÃ³var-GarcÃa, Karinna; Kwon, Minjae; Samaras, Susan E et al. (2014) ANKRD1 acts as a transcriptional repressor of MMP13 via the AP-1 site. Mol Cell Biol 34:1500-11|