Chemosensory mechanisms of bitter taste receptors (T2Rs) in mediating host-pathogen interactions in airways

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Jaggupilli, Appalaraju
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Bitter taste receptors (T2Rs) belong to G protein-coupled receptor superfamily. In humans, 25 T2Rs perform a chemosensory function with little information on their extraoral role. Recent studies suggest the expression of T2Rs in different tissues and their interaction with quorum-sensing molecules (QSMs). This thesis has two independent hypotheses involving T2Rs: First hypothesis, human airways show differential expression of T2Rs in pathophysiological conditions such as cystic fibrosis (CF). The second hypothesis, bacterial QSMs and bitter compounds mediate host-pathogen interactions through T2Rs in airways. To test these hypotheses, I analyzed the expression patterns of T2Rs at transcript and protein level in normal and CF airway cells. The results suggest a specific pattern of T2Rs with no differential expression in the samples analyzed. Next to assess T2R functionality, I pursued calcium mobilization assays after stimulation of cells with various bitter compounds. The inhibition of Gβγ-subunit and phospholipase-C (PLC) showed the calcium mobilized in these cells predominantly takes place through T2R-Gαβγ-PLC pathway. To test the second hypothesis, combination of molecular and pharmacological approaches were used to study the amino acid interactions in selected T2Rs. In these experiments, commonly used antibiotics in CF treatment, and major QSMs secreted by CF bacteria were tested. The results suggest that antibiotics and QSMs activate multiple T2Rs with different potencies. Extracellular loop-2 in T2Rs performs a key function in binding to the tested compounds. To address the lack of properly characterized bitter blockers, I pursued the characterization of novel T2R4 blockers derived from plant and meat products. A Schild regression analysis on the plant hormone abscisic acid determined its T2R4 antagonism as surmountable. Further testing with the naturally occuring (+)-ABA isomer indicated that individual isomers do not induce calcium mobilization. Subsequently, I identified and characterized advanced glycation end-products as T2R4 antagonists. Glyoxal-derived lysine dimer inhibited the quinine response while carboxy methyl lysate showed subtle inhibition. In conclusion, these structure-function and mechanistic studies on T2Rs identify as new targets for antibiotics and QSMs. This work provides new insights into the host-pathogen interactions in CF and may lead to novel therapeutic approaches for CF that target T2Rs.
bitter taste receptors, cystic fibrosis, antibiotics, GPCR, quorum sensing