The plasminogen activator (PA) system plays an important role in cell adhesion, migration and invasion, and may require the coordinated expression of various proteins. The human urokinase-type plasminogen activator (uPA) receptor (uPAR) is a central protein component of the PA system. By binding its ligand uPA, uPAR can direct proteolysis of the extracellular matrix. Also, it is now apparent that uPAR can initiate proteolytic independent signal transduction to influence angiogenesis, inflammation, wound repair and tumour progression. To determine whether any novel proteins interacted with uPAR, a yeast two-hybrid screening analysis was undertaken using alternate uPAR domain constructs as baits. These included full-length three domain uPAR (uPAR-DIDIIDIII), two domain uPAR (uPAR-DIIDIII), and each individual uPAR domain (uPAR-DI, uPAR-DII and uPAR-DIII). A number of proteins were identified as putative candidate interactors for the alternate constructs, with two of special interest for uPAR-DIDIIDIII. These were the heat shock protein Mrj, and the extracellular matrix protein fibulin-2. The protein Mrj was shown to bind uPAR both in vitro and in vivo using GST-pull down and co-immunoprecipitation assays respectively. The GST-pull down assay identified the interaction between Mrj and uPAR dependent on the C-terminal domain of Mrj and DI of uPAR. Using in vivo co-immunoprecipitation analysis, Mrj also bound to uPAR. Preliminary data suggest the association between uPAR and Mrj may play a role in the regulation of apoptosis. In regard to the uPAR interactor of fibulin-2, a calcium dependent binding interaction with uPAR was identified using the GST-pull down assay. However due to the large molecular weight and stringent conditions needed to solubilise fibulin-2, it was not possible to co-immunoprecipitate both uPAR and fibulin-2. Together, the identification of both Mrj and fibulin-2 amongst other candidate interactors of uPAR presented here provides further insight into the intricate relationship between uPAR and other proteins which may influence a range of biological functions.

The urokinase receptor (uPAR) plays a central role in metastatic process. It???s evident uPAR is overexpressed across a variety of tumour cells and leads to the increased aggressiveness and poor prognosis of cancer. Inhibition of uPAR expression can block metastatic potential in many tumours. In addition, besides uPA, there are several other proteins which have been confirmed to interact with uPAR, such as vitronectin and integrins. These interactions also contribute to signal transduction and the functions of uPAR complex. Therefore, downregulation of uPAR expression by targeting uPAR mRNA or protein, or by regulating the uPAR partners would be potential therapeutic strategies for prevention of cancer metastasis. There are two main aspects contained in this thesis. Firstly, three specific DNAzymes targeting uPAR mRNA were designed to downregulate uPAR expression in vitro and their effects to decrease cancer cell invasion studied in a human osteosarcoma cell line Saos-2. The results showed that two of them (Dz483 and Dz720) cleaved uPAR transcript in vitro with high efficacy and specificity and the Dz720 inhibited uPAR protein levels by 55% in Saos-2 cells. Besides, the Dz720 significantly suppressed Saos-2 cell invasion using an in vitro matrigel assay. Secondly, two potential uPAR partners from yeast two-hybrid screening, a heat shock protein MRJ and an anti-apoptosis protein HAX-1, were characterised and their functions binding with uPAR investigated. The interactions were confirmed by co-immunoprecipitation, GST-pull down assay and confocal microscopy in cancer cells. In addition, there was a 50% increase in cell adhesion after transfection with MRJ. This increase in adhesion is dependent on the uPAR/full length MRJ interaction as cells transfected with the mutant construct containing only N-terminal region or C-terminal region of MRJ had no increase in cell adhesion. The observed increase in adhesion to vitronectin by MRJ was also blocked by an anti-uPAR domain I antibody suggesting that the induced adhesion is at least in part contributed by uPAR on the cell surface. Together, the identification of both MRJ and HAX-1 as uPAR interactors provides further insight into the intricate relationship between uPAR and other proteins which may develop potential approaches for cancer therapy.

The urokinase receptor (uPAR) plays a central role in metastatic process. It???s evident uPAR is overexpressed across a variety of tumour cells and leads to the increased aggressiveness and poor prognosis of cancer. Inhibition of uPAR expression can block metastatic potential in many tumours. In addition, besides uPA, there are several other proteins which have been confirmed to interact with uPAR, such as vitronectin and integrins. These interactions also contribute to signal transduction and the functions of uPAR complex. Therefore, downregulation of uPAR expression by targeting uPAR mRNA or protein, or by regulating the uPAR partners would be potential therapeutic strategies for prevention of cancer metastasis. There are two main aspects contained in this thesis. Firstly, three specific DNAzymes targeting uPAR mRNA were designed to downregulate uPAR expression in vitro and their effects to decrease cancer cell invasion studied in a human osteosarcoma cell line Saos-2. The results showed that two of them (Dz483 and Dz720) cleaved uPAR transcript in vitro with high efficacy and specificity and the Dz720 inhibited uPAR protein levels by 55% in Saos-2 cells. Besides, the Dz720 significantly suppressed Saos-2 cell invasion using an in vitro matrigel assay. Secondly, two potential uPAR partners from yeast two-hybrid screening, a heat shock protein MRJ and an anti-apoptosis protein HAX-1, were characterised and their functions binding with uPAR investigated. The interactions were confirmed by co-immunoprecipitation, GST-pull down assay and confocal microscopy in cancer cells. In addition, there was a 50% increase in cell adhesion after transfection with MRJ. This increase in adhesion is dependent on the uPAR/full length MRJ interaction as cells transfected with the mutant construct containing only N-terminal region or C-terminal region of MRJ had no increase in cell adhesion. The observed increase in adhesion to vitronectin by MRJ was also blocked by an anti-uPAR domain I antibody suggesting that the induced adhesion is at least in part contributed by uPAR on the cell surface. Together, the identification of both MRJ and HAX-1 as uPAR interactors provides further insight into the intricate relationship between uPAR and other proteins which may develop potential approaches for cancer therapy.

This master’s thesis is concerned with analysis of cabling and cabling systems in data center and with their projection as follows. The part of the work is aimed at the measurement parameters of metal cables which are used in data center with noise. The first part is focused on analyzing the structure of the data center and cabling used in it. The structure of the data center is based on the TIA-942 and consists of an entrance room, main distribution area MDA, horizontal distribution area HDA , zone distribution area ZDA and the equipment distribution area EDA. Divided into backbone cabling and horizontal cabling and use standard and pre-terminated cabling. The measurement results of structured metal cabling with noise are discussed in the next part. There were used these types of cables UTP, FTP and S/FTP. The measurement was performed in the electricity substation and in the laboratory of electromagnetic compatibility. The third part of this work deals with the actual design of structured cabling for the data center. In the first design we use the standard installation, the second preterminated cabling. For both types of installation cost statement is processed (CAPEX). Thesis is also completed with DC design drawings and rack drawings.