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The glutamate transporter excitatory amino acid carrier (EAAC1/EAAT3) mediates the absorption of dicarboxylic amino acids in epithelial cells as well as the uptake of glutamate from the synaptic cleft. Its cell-surface density is regulated by interaction with accessory proteins which remain to be identified. We detected a consensus sequence for interaction with post-synaptic density-95/Discs large/Zonula occludens (PDZ) proteins (-SQF) and a tyrosine-based internalization signal (-YVNG-) in the C-terminus of EAAC1, and investigated their role in the transporter localization. We demonstrated that PDZ interactions are required for the efficient delivery to and the retention in the plasma membrane of EAAC1 and we identified PDZK1/NHERF3 (Na+/H+-exchanger regulatory factor 3) as a novel EAAC1 interacting protein. Expression of PDZK1 in Madin-Darby canine kidney (MDCK) cells tethered EAAC1 to filopodia and increased its surface activity. Removal of the PDZ-target motif promoted the EAAC1 binding to α-adaptin and clathrin and the transporter internalization in endocytic/degradative compartments. This defect was largely prevented by hypertonic treatment or overexpression of the dominant-negative µ2-W421A-subunit of AP-2 clathrin-adaptor. The rate of transporter endocytosis was attenuated following tyrosine mutagenesis in the internalization signal, thus indicating that this motif can regulate the transporter endocytosis. We suggest that EAAC1 density is controlled by balanced interactions with PDZK1 and adaptor protein 2 (AP2): the former promotes the transporter expression at the cell surface, and the latter mediates its constitutive endocytosis., (© 2010 John Wiley & Sons A/S.)

In this study, we examined the role of the L27 [(LIN2-LIN7) domain] and PDZ domain (domain previously found in PSD95-DlgA-ZO-1) for protein-protein interaction of the scaffold protein LIN7 in tight junction (TJ) assembly in Madin-Darby canine kidney (MDCK) cells and found that the stable expression of a LIN7 mutant lacking the L27 domain (DeltaL27 mutant) acts as a dominant interfering protein by inhibiting TJ localization of endogenous LIN7. The loss of LIN7 did not alter the localization of the PALS1 (protein associated with LIN7) partner of the L27 domain but prevented TJ localization of the insulin receptor substrate p53 (IRSp53), a partner of the PDZ domain of LIN7. The function of both L27 and PDZ domains of LIN7 in IRSp53 localization to TJs has been further demonstrated by reducing the expression of LIN7 (LIN7 small hairpin RNA experiments) and by expression of IRSp53 deleted of its motif for PDZ interaction (IRSp53Delta5) or fused to the L27 domain of LIN7 (L27-IRSp53Delta5). Cell lines with decreased localization of LIN7 and IRSp53 to TJs showed defects during assembly of TJs and cyst polarization and failed to activate Rac1, a member of the Rho guanosine triphosphatases family crucially involved in actin organization and orientation of apicobasal polarity. These data therefore indicate that LIN7-IRSp53 association plays a role during assembly of functional TJs and surface polarization in epithelial cells.

Background: Pseudomonas aeruginosa is responsible for a wide range of infections. In immunocompromised patients with cancer, the emergence of multidrug resistant P. aeruginosa may have grave consequences., Methods: Patients with cancer who were infected with multidrug-resistant P. aeruginosa with polyclonal DNA restriction patterns were used as the case group. Two control groups were used: one group of cancer patients who were infected with multidrug-susceptible P. aeruginosa and another group of cancer patients who had the same underlying disease and the same intensive care unit exposure as patients in the case group but who were not infected or colonized by P. aeruginosa., Results: Risk factors that were associated significantly with multidrug-resistant P. aeruginosa infection were the use of carbapenem for > or = 7 days, a history of P. aeruginosa infection during the preceding year, and a history of chronic obstructive pulmonary disease (P < 0.01)., Conclusions: Carbapenems may need to be used more judiciously as first-line empirical therapy for cancer patients with prior pseudomonal infection or chronic obstructive pulmonary disease who require hospitalization, and alternative, antipseudomonal antibiotic regimens may need to be considered, especially in this patient population.

Background: Invasive aspergillosis (IA) has emerged as a common cause of morbidity and mortality among immunocompromised patients. At The University of Texas M. D. Anderson Cancer Center (Houston, TX), Aspergillus terreus is second to A. fumigatus as the most common cause of IA. In the current study, the authors compared the risk factors and outcomes associated with IA caused by A. terreus and IA caused by A. fumigatus., Methods: The authors retrospectively reviewed the medical records of 300 patients who received care at our institution between 1995 and 2001 and who had cultures that were positive for Aspergillus infection, including 90 patients whose cultures were positive for A. fumigatus and 70 patients whose cultures were positive for A. terreus., Results: Thirty-two patients with IA caused by A. terreus and 33 patients with IA caused by A. fumigatus were evaluated. The two groups were comparable in terms of age, gender, and underlying disease. Leukemia was the most common underlying malignancy (84%). More than 40% of patients in each group had undergone bone marrow transplantation. There was a trend toward a higher frequency of neutropenia among patients with IA caused by A. terreus (P = 0.12). IA caused by A. terreus was considered to be nosocomial in origin significantly more frequently compared with IA caused by A. fumigatus (P = 0.03). In vitro, A. terreus was found to be more resistant to amphotericin B (minimal inhibitory concentration [MIC90], 4.0 microg/mL) than to antifungal therapy (MIC90, 1.0 Hg/mL) in the isolates that were tested (< 50% of all isolates). The overall rate of response to antifungal therapy was 39% for patients with A. fumigatus infection, compared with 28% for patients with A. terreus infection (P = 0.43)., Conclusions: Despite the decreased in vitro susceptibility of A. terreus (relative to A. fumigatus) to amphotericin B, the two groups within the current patient population had comparably poor responses to amphotericin B preparation and somewhat improved responses to posaconazole., ((c) 2004 American Cancer Society.)