Mechanisms of Olfaction in Insects - ResearchSpace@Auckland ...
Mechanisms of Olfaction in Insects - ResearchSpace@Auckland ...
Mechanisms of Olfaction in Insects - ResearchSpace@Auckland ...
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Functional characterisation <strong>of</strong> Epiphyas postvittana odorant receptor 1 37<br />
Miura et al. (2009) co-expressed O. scapulalis OR1 (OscaOR1) together with its<br />
species-specific OR83b homologue <strong>in</strong> X. laevis oocytes and showed that it responded<br />
highly to E11-14:OH and a small response was shown to Z11-16:OAc (Miura et al.,<br />
2009). These compounds have been identified as pheromone components <strong>of</strong> other<br />
Ostr<strong>in</strong>ia species, while the sex pheromone components <strong>of</strong> O. scapulalis (E11- and<br />
Z11- 14:OAc) did not confer a response from OscaOR1, <strong>in</strong>dicat<strong>in</strong>g that even though<br />
OscaOR1 is homologous to the pheromone receptors <strong>of</strong> Ostr<strong>in</strong>ia species, it is not<br />
specific to O. scapulalis sex pheromone. Wanner et al. (2010) expressed Ostr<strong>in</strong>ia<br />
nubilalis OR1, and ORs 3–6 <strong>in</strong> X. laevis oocytes and showed that they bound all the<br />
components <strong>of</strong> the Ostr<strong>in</strong>ia sex pheromone, as shown <strong>in</strong> Table 1.1 (Wanner et al.,<br />
2010).<br />
Mitsuno et al. (2008) expressed the pheromone receptor together with their species<br />
specific OR83b homologue from three different moth species (P. xylostella, M.<br />
separata and D. <strong>in</strong>dica) <strong>in</strong> X. laevis oocytes and showed via electrophysiological<br />
record<strong>in</strong>gs that they all bound their respective sex pheromone components with an<br />
EC50 <strong>in</strong> the micromolar range and the lowest b<strong>in</strong>d<strong>in</strong>g threshold <strong>of</strong> 10 -7 M, values<br />
which are comparable with B. mori PR assays conducted by Nakagawa et al. (2005).<br />
Modified versions <strong>of</strong> HEK293 cells, conta<strong>in</strong><strong>in</strong>g a mouse Gα15 gene have also been<br />
used for successfully express<strong>in</strong>g and characteris<strong>in</strong>g moth ORs <strong>in</strong> fluorometric assays.<br />
Stable cell l<strong>in</strong>es <strong>of</strong> H. virescens HvOR13, HvOR14 and HvOR16 have been generated<br />
and characterised to b<strong>in</strong>d H. virescens sex pheromone components <strong>in</strong> HEK cells with<br />
high aff<strong>in</strong>ity. HvOR13 can recognise Z11-16:Al <strong>in</strong> DMSO with an EC50 <strong>of</strong> 1.2 x 10 -9<br />
M (Große-Wilde et al., 2007). These results are comparable to an earlier study <strong>in</strong><br />
which B. mori OR1 and OR3 were expressed and characterised <strong>in</strong> HEK293 cells.<br />
BmOR1 bound the sex pheromone bombykol with an EC50 <strong>of</strong> 10 -10 M (Große-Wilde<br />
et al., 2006). Together these results show that expression <strong>of</strong> moth ORs <strong>in</strong> HEK293<br />
cells yield functional ORs, that are highly responsive and sensitive to their ligands.<br />
Kiely et al. (2006) developed an <strong>in</strong>sect cell system for assay<strong>in</strong>g <strong>in</strong>sect ORs.<br />
Drosophila OR22a was transiently expressed <strong>in</strong> Sf9 cells, and after a 48 hour<br />
<strong>in</strong>cubation period, the cells were loaded with the calcium sensitive dye Fluo-4 and<br />
change <strong>in</strong> fluorescence <strong>of</strong> cells <strong>in</strong> response to stimulation by odorants was measured