Team:TU-Delft/part1
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In S. cerevisiae two GPCR cascades have been identified: a glucose sensing pathway and a mating pathway [5]. There are two sexes of yeast cells, MATa and MATα. Whenever pheromones (small peptides) of the opposite sex are bound to the specific G-protein coupled receptors (Ste2 p or Ste3p), the MAP kinase cascade is turned on, leading to induction of mating genes such as Fus1 and growth arrest mediated by the Far1 promoter. This mating response can be seen in the form of a morphological change, called shmoo formation. In figure an overview of the pheromone and glucose signaling pathways in S. cerevisiae is shown. | In S. cerevisiae two GPCR cascades have been identified: a glucose sensing pathway and a mating pathway [5]. There are two sexes of yeast cells, MATa and MATα. Whenever pheromones (small peptides) of the opposite sex are bound to the specific G-protein coupled receptors (Ste2 p or Ste3p), the MAP kinase cascade is turned on, leading to induction of mating genes such as Fus1 and growth arrest mediated by the Far1 promoter. This mating response can be seen in the form of a morphological change, called shmoo formation. In figure an overview of the pheromone and glucose signaling pathways in S. cerevisiae is shown. | ||
+ | <h2>INTRODUCTION OF A NEW OLFACTORY RECEPTOR</h2> | ||
+ | Previously it was found that that the yeast pheromone signaling pathway can be coupled to a mammalian olfactory receptor. Minic et al. succeeded in functional express the rat 17 OR and its trafficking to the plasma membrane in S. cerevisiae. Between the three GPCRs that are known in S. cerevisiae, Ste2, Ste3 and Gpr1, the sequence similarity is limited. Except for pheromone receptors in Schizosaccharomyces pombe and Kluyveromyces lactis, Ste2 and Ste3 are largely unrelated in sequence to other GPCRs. [5]. Nevertheless, the yeast pheromone receptors can be functionally replaced by several mammalian GPCRs so that the pheromone pathway can be activated by the corresponding ligands [4]. | ||
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+ | <h2>Chimeric design</h2> | ||
+ | A major hindrance for functional expression of ORs has been that the receptors did not localize in the membrane or that the downstream coupling of the receptor to the Gα did not work properly. It has been shown that the rat olfactory receptor 17 (R17) that responds to octanal can be functionally expressed in many different cell types, including S. cerevisiae. [6]. Earlier research investigated on the question whether the RI7 sequence can be used to functionally express other ORs. Sequence analysis of ORs have shown that the N termini of the receptor are involved in plasma membrane localization, whereas the C termini generally define the specificity for G protein interaction [Chemical sensing of DNT]. Based on this observations Radhika et al. functionally expressed a chimeric OR with the N terminus and the C terminus of the RI7 sequence. A schematic picture is shown in figure. | ||
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Revision as of 10:29, 25 September 2012