ABSTRACT
Interkingdom communication between bacteria and host organisms is one of the most interesting research topics in biology. Quorum sensing molecules produced by Gram-negative bacteria, such as acylated homoserine lactones and quinolones, have been shown to interact with host cell receptors, stimulating innate immunity and bacterial clearance. To our knowledge, there is no evidence that these molecules influence CNS function. Here, we have found that low micromolar concentrations of the Pseudomonas aeruginosa quorum sensing autoinducer, 2-heptyl-3-hydroxy-4-quinolone (PQS), inhibited polyphosphoinositide hydrolysis in mouse brain slices, whereas four selected acylated homoserine lactones were inactive. PQS also inhibited forskolin-stimulated cAMP formation in brain slices. We therefore focused on PQS in our study. Biochemical effects of PQS were not mediated by the bitter taste receptors, T2R4 and T2R16. Interestingly, submicromolar concentrations of PQS could be detected in the serum and brain tissue of adult mice under normal conditions. Levels increased in five selected brain regions after single i.p. injection of PQS (10 mg/kg), peaked after 15 min, and returned back to normal between 1 and 4 h. Systemically administered PQS reduced spontaneous locomotor activity, increased the immobility time in the forced swim test, and largely attenuated motor response to the psychostimulant, methamphetamine. These findings offer the first demonstration that a quorum sensing molecule specifically produced by Pseudomonas aeruginosa is centrally active and influences cell signaling and behavior. Quorum sensing autoinducers might represent new interkingdom signaling molecules between ecological communities of commensal, symbiotic, and pathogenic microorganisms and the host CNS.
Subject(s)
Behavior, Animal/drug effects , Brain/drug effects , Cyclic AMP/metabolism , Phosphatidylinositol Phosphates/metabolism , Pseudomonas aeruginosa/metabolism , Quinolones/pharmacology , Quorum Sensing , Signal Transduction/drug effects , Animals , Brain/metabolism , Host-Pathogen Interactions , Hydrolysis , In Vitro Techniques , Locomotion/drug effects , Male , Mice , Morris Water Maze Test/drug effects , Motor Activity/drug effects , Quinolones/metabolismABSTRACT
Myosin Va (MyoVa) is an actin-based molecular motor abundantly found at the centrosome. However, the role of MyoVa at this organelle has been elusive due to the lack of evidence on interacting partners or functional data. Herein, we combined yeast two-hybrid screen, biochemical studies and cellular assays to demonstrate that MyoVa interacts with RPGRIP1L, a cilia-centrosomal protein that controls ciliary signaling and positioning. MyoVa binds to the C2 domains of RPGRIP1L via residues located near or in the Rab11a-binding site, a conserved site in the globular tail domain (GTD) from class V myosins. According to proximity ligation assays, MyoVa and RPGRIP1L can interact near the cilium base in ciliated RPE cells. Furthermore, we showed that RPE cells expressing dominant-negative constructs of MyoVa are mostly unciliated, providing the first experimental evidence about a possible link between this molecular motor and cilia-related processes.
Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Myosin Heavy Chains/metabolism , Myosin Type V/metabolism , Adaptor Proteins, Signal Transducing/chemistry , Adaptor Proteins, Signal Transducing/genetics , Amino Acid Sequence , Binding Sites , Centrosome/metabolism , Cilia/genetics , Cilia/metabolism , Conserved Sequence , Humans , Models, Molecular , Myosin Heavy Chains/chemistry , Myosin Heavy Chains/genetics , Myosin Type V/chemistry , Myosin Type V/genetics , Protein Binding , Protein Conformation , Protein Interaction Domains and Motifs , Recombinant ProteinsABSTRACT
The human proteins FEZ1 (fasciculation and elongation protein zeta 1) and FEZ2 are orthologs of the protein UNC-76 from C. elegans, involved in the growth and fasciculation of the worms axon. Pull down assays showed that the protein FEZ1 interacts with other proteins (e.g., the protein SCOCO, short coiled-coil protein), mitochondria, and vesicles. These components may therefore represent cargoes to be transported along the microtubule, and the transport may be mediated through FEZ1 reported binding to kinesins (KIF3A). We previously showed that FEZ1 dimerizes in its N-terminal region and interacts with other proteins, including the candidate cargoe proteins, through its C-terminus. Here, we studied the fragment FEZ1(92-194) as well as full-length 6xHis-FEZ1 (1-392) in vitro and endogenous FEZ1 isolated from HEK 293 cells and were able to demonstrate the formation of an intermolecular disulfide bond through FEZ1 Cys-133, which appears to be essential for dimerization. This disulfide bond may be important for the FEZ1 role as a dimeric and bivalent transport adaptor molecule, since it establishes a strong link between the monomers, which could be a prerequisite for the simultaneous binding of two cargoes.
Subject(s)
Adaptor Proteins, Signal Transducing/chemistry , Disulfides/chemistry , Nerve Tissue Proteins/chemistry , Proteomics/methods , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Amino Acid Sequence , Cell Adhesion Molecules/chemistry , Cell Adhesion Molecules/metabolism , Cell Line , Conserved Sequence , Disulfides/metabolism , Humans , Mass Spectrometry , Models, Molecular , Molecular Sequence Annotation , Molecular Sequence Data , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Protein Multimerization , Protein Structure, Tertiary , Protein Transport , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Scattering, Small Angle , Sequence Alignment , X-Ray DiffractionABSTRACT
BACKGROUND: The established pathologic criteria for minor salivary gland (MSG) involvement in chronic graft-vs.-host disease (cGVHD) could play a role in monitoring response to therapy. METHODS: We evaluated MSG sequential biopsies during cGVHD therapy in 14 allogeneic bone marrow transplantation (BMT) patients. Nine patients that did not develop GVHD after BMT entered the control group. Biopsies were examined using hematoxylin-eosin, Periodic acid-Schiff (PAS) and leukocyte common antigen staining. RESULTS: A significant loss of PAS+ acinar volume was observed at the diagnosis of cGVHD as much as at the end of treatment when compared with the control group. In the second evaluation, the inflammatory infiltrate was still greater than control group. CONCLUSIONS: The results suggest that persistent xerostomia after cGVHD treatment is because of maintenance of lymphocytic infiltrate and consequent absence of MSG secretory unit recovery. This data may be useful to provide improved insight into the histopathology of this organ involvement.
