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Daily rhythms in antennal protein and olfactory sensitivity in the malaria mosquito Anopheles gambiae.

Rund SS, Bonar NA, Champion MM, Ghazi JP, Houk CM, Leming MT, Syed Z, Duffield GE - Sci Rep (2013)

Bottom Line: Further, electrophysiological investigations demonstrate time-of-day specific differences in olfactory sensitivity of antennae to major host-derived odorants.The pre-dusk/dusk peaks in OBPs and takeout gene expression correspond with peak protein abundance at night, and in turn coincide with the time of increased olfactory sensitivity to odorants requiring OBPs and times of increased blood-feeding behavior.This suggests an important role for OBPs in modulating temporal changes in odorant sensitivity, enabling the olfactory system to coordinate with the circadian niche of An. gambiae.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biological Sciences and Eck Institute for Global Health, Galvin Life Science Center, University of Notre Dame, Notre Dame, IN 46556 [2].

ABSTRACT
We recently characterized 24-hr daily rhythmic patterns of gene expression in Anopheles gambiae mosquitoes. These include numerous odorant binding proteins (OBPs), soluble odorant carrying proteins enriched in olfactory organs. Here we demonstrate that multiple rhythmically expressed genes including OBPs and takeout proteins, involved in regulating blood feeding behavior, have corresponding rhythmic protein levels as measured by quantitative proteomics. This includes AgamOBP1, previously shown as important to An. gambiae odorant sensing. Further, electrophysiological investigations demonstrate time-of-day specific differences in olfactory sensitivity of antennae to major host-derived odorants. The pre-dusk/dusk peaks in OBPs and takeout gene expression correspond with peak protein abundance at night, and in turn coincide with the time of increased olfactory sensitivity to odorants requiring OBPs and times of increased blood-feeding behavior. This suggests an important role for OBPs in modulating temporal changes in odorant sensitivity, enabling the olfactory system to coordinate with the circadian niche of An. gambiae.

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Temporal differences to a hydrophobic odorant are maintained in dark treated mosquitoes, but there are no time-of-day specific changes to hexanoic acid.(a) Time-of-day differences were retained between mosquitoes at ZT16 and mosquitoes at ZT8 pretreated with dark for 4 hr (two way ANOVA: t, F1,36 = 15.4, p < 0.001; c, F2,36 = 192.7, p < 0.001; i, F2,36 = 1.9, n.s.). For nonanal (−1), Tukey post hoc tests also revealed a difference between ZT8 and ZT16 (p < 0.05). n = 5–9 recordings per group. (b) Mosquito EAG responses to hexanoic acid were concentration dependent, but not time-of-day dependent (two way ANOVA: t, F3,77 = 0.8, n.s.; c, F2,77 = 106.0, p < 0.001; i, F6,77 = 0.6, n.s.). Traces represent, the mean ± S.E.M. EAG responses at all three concentrations and all four tested time points. n = 6–8 recordings per group. The nonanal control showed expected time-of-day specific changes in sensitivity (ANOVA: F3,26 = 5.9, p < 0.01) and Tukey post hoc test results are shown. Bar charts represent mean ± S.E.M values. *p < 0.05, **p < 0.01.
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f5: Temporal differences to a hydrophobic odorant are maintained in dark treated mosquitoes, but there are no time-of-day specific changes to hexanoic acid.(a) Time-of-day differences were retained between mosquitoes at ZT16 and mosquitoes at ZT8 pretreated with dark for 4 hr (two way ANOVA: t, F1,36 = 15.4, p < 0.001; c, F2,36 = 192.7, p < 0.001; i, F2,36 = 1.9, n.s.). For nonanal (−1), Tukey post hoc tests also revealed a difference between ZT8 and ZT16 (p < 0.05). n = 5–9 recordings per group. (b) Mosquito EAG responses to hexanoic acid were concentration dependent, but not time-of-day dependent (two way ANOVA: t, F3,77 = 0.8, n.s.; c, F2,77 = 106.0, p < 0.001; i, F6,77 = 0.6, n.s.). Traces represent, the mean ± S.E.M. EAG responses at all three concentrations and all four tested time points. n = 6–8 recordings per group. The nonanal control showed expected time-of-day specific changes in sensitivity (ANOVA: F3,26 = 5.9, p < 0.01) and Tukey post hoc test results are shown. Bar charts represent mean ± S.E.M values. *p < 0.05, **p < 0.01.

Mentions: We next determined if the temporal rhythms described in protein levels correspond with olfactory sensitivity. We used electroantennogram (EAG) analysis to measure the olfactory responses induced by host-derived odorant chemicals. At ZT4 (morning), ZT8 (afternoon), ZT12 (dusk) and ZT16 (night), mosquito antennae were challenged with major host-derived odorant chemicals (nonanal, indole, geranyl acetone, and a mixture of hexanoic acid, geranyl acetone, nonanal, indole, 3-methylindole and p-cresol). As several of the odorants are hydrophobic, it is expected that OBPs will be necessary for the mosquito to detect these stimuli. Since OBPs are rhythmic in protein abundance, we predicted rhythmic sensitivity in detection of these compounds. Indeed, we found time-of-day specific olfactory responses to all four stimuli, with sensitivity peaking at ZT16 (night) and least at ZT4 or ZT8 (day) (Fig. 4a–b). In all cases, the time-of-day specific differences in EAG amplitudes were comparable to the differences observed due to 10-fold changes in odorant concentration (mean peak-to-nadir difference in each time series: Geranyl acetone 44%, indole 50% and nonanal 61%; mean increase in response with each 10× increase in odorant concentration: Geranyl acetone 15%, indole 67%, and nonanal 81%). It is therefore plausible that the ≤8.4-fold rhythmic changes observed in OBP abundance (antennae mean ± SEM fold-change, 3.4 ± 0.4; THAs, 3.2 ± 0.6) could account for such differences in EAG responses. This is especially so given that in some cases multiple OBPs can bind a single odorant, and vice versa11141517. These findings are broadly consistent with our observed OBP protein rhythms (Fig. 2, Fig. S3) and mosquito behaviors described here (Fig. 6) and by others15. Specifically, protein expression and olfactory sensitivity were also lowest at ZT4-ZT8 and highest at ~ZT16, and blood-feeding and flight behaviors were higher at night than during the daytime. These data are particularly interesting for indole as there is significant evidence that OBP1 contributes to antennal sensing of this odorant1117. OBP1 protein rhythm has a nadir at ZT4-ZT8 and a peak at ZT16 (Fig. 1, Fig. 2), which corresponds with the lowest and highest sensitivity to indole in our electrophysiological measurements.


Daily rhythms in antennal protein and olfactory sensitivity in the malaria mosquito Anopheles gambiae.

Rund SS, Bonar NA, Champion MM, Ghazi JP, Houk CM, Leming MT, Syed Z, Duffield GE - Sci Rep (2013)

Temporal differences to a hydrophobic odorant are maintained in dark treated mosquitoes, but there are no time-of-day specific changes to hexanoic acid.(a) Time-of-day differences were retained between mosquitoes at ZT16 and mosquitoes at ZT8 pretreated with dark for 4 hr (two way ANOVA: t, F1,36 = 15.4, p < 0.001; c, F2,36 = 192.7, p < 0.001; i, F2,36 = 1.9, n.s.). For nonanal (−1), Tukey post hoc tests also revealed a difference between ZT8 and ZT16 (p < 0.05). n = 5–9 recordings per group. (b) Mosquito EAG responses to hexanoic acid were concentration dependent, but not time-of-day dependent (two way ANOVA: t, F3,77 = 0.8, n.s.; c, F2,77 = 106.0, p < 0.001; i, F6,77 = 0.6, n.s.). Traces represent, the mean ± S.E.M. EAG responses at all three concentrations and all four tested time points. n = 6–8 recordings per group. The nonanal control showed expected time-of-day specific changes in sensitivity (ANOVA: F3,26 = 5.9, p < 0.01) and Tukey post hoc test results are shown. Bar charts represent mean ± S.E.M values. *p < 0.05, **p < 0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

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f5: Temporal differences to a hydrophobic odorant are maintained in dark treated mosquitoes, but there are no time-of-day specific changes to hexanoic acid.(a) Time-of-day differences were retained between mosquitoes at ZT16 and mosquitoes at ZT8 pretreated with dark for 4 hr (two way ANOVA: t, F1,36 = 15.4, p < 0.001; c, F2,36 = 192.7, p < 0.001; i, F2,36 = 1.9, n.s.). For nonanal (−1), Tukey post hoc tests also revealed a difference between ZT8 and ZT16 (p < 0.05). n = 5–9 recordings per group. (b) Mosquito EAG responses to hexanoic acid were concentration dependent, but not time-of-day dependent (two way ANOVA: t, F3,77 = 0.8, n.s.; c, F2,77 = 106.0, p < 0.001; i, F6,77 = 0.6, n.s.). Traces represent, the mean ± S.E.M. EAG responses at all three concentrations and all four tested time points. n = 6–8 recordings per group. The nonanal control showed expected time-of-day specific changes in sensitivity (ANOVA: F3,26 = 5.9, p < 0.01) and Tukey post hoc test results are shown. Bar charts represent mean ± S.E.M values. *p < 0.05, **p < 0.01.
Mentions: We next determined if the temporal rhythms described in protein levels correspond with olfactory sensitivity. We used electroantennogram (EAG) analysis to measure the olfactory responses induced by host-derived odorant chemicals. At ZT4 (morning), ZT8 (afternoon), ZT12 (dusk) and ZT16 (night), mosquito antennae were challenged with major host-derived odorant chemicals (nonanal, indole, geranyl acetone, and a mixture of hexanoic acid, geranyl acetone, nonanal, indole, 3-methylindole and p-cresol). As several of the odorants are hydrophobic, it is expected that OBPs will be necessary for the mosquito to detect these stimuli. Since OBPs are rhythmic in protein abundance, we predicted rhythmic sensitivity in detection of these compounds. Indeed, we found time-of-day specific olfactory responses to all four stimuli, with sensitivity peaking at ZT16 (night) and least at ZT4 or ZT8 (day) (Fig. 4a–b). In all cases, the time-of-day specific differences in EAG amplitudes were comparable to the differences observed due to 10-fold changes in odorant concentration (mean peak-to-nadir difference in each time series: Geranyl acetone 44%, indole 50% and nonanal 61%; mean increase in response with each 10× increase in odorant concentration: Geranyl acetone 15%, indole 67%, and nonanal 81%). It is therefore plausible that the ≤8.4-fold rhythmic changes observed in OBP abundance (antennae mean ± SEM fold-change, 3.4 ± 0.4; THAs, 3.2 ± 0.6) could account for such differences in EAG responses. This is especially so given that in some cases multiple OBPs can bind a single odorant, and vice versa11141517. These findings are broadly consistent with our observed OBP protein rhythms (Fig. 2, Fig. S3) and mosquito behaviors described here (Fig. 6) and by others15. Specifically, protein expression and olfactory sensitivity were also lowest at ZT4-ZT8 and highest at ~ZT16, and blood-feeding and flight behaviors were higher at night than during the daytime. These data are particularly interesting for indole as there is significant evidence that OBP1 contributes to antennal sensing of this odorant1117. OBP1 protein rhythm has a nadir at ZT4-ZT8 and a peak at ZT16 (Fig. 1, Fig. 2), which corresponds with the lowest and highest sensitivity to indole in our electrophysiological measurements.

Bottom Line: Further, electrophysiological investigations demonstrate time-of-day specific differences in olfactory sensitivity of antennae to major host-derived odorants.The pre-dusk/dusk peaks in OBPs and takeout gene expression correspond with peak protein abundance at night, and in turn coincide with the time of increased olfactory sensitivity to odorants requiring OBPs and times of increased blood-feeding behavior.This suggests an important role for OBPs in modulating temporal changes in odorant sensitivity, enabling the olfactory system to coordinate with the circadian niche of An. gambiae.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biological Sciences and Eck Institute for Global Health, Galvin Life Science Center, University of Notre Dame, Notre Dame, IN 46556 [2].

ABSTRACT
We recently characterized 24-hr daily rhythmic patterns of gene expression in Anopheles gambiae mosquitoes. These include numerous odorant binding proteins (OBPs), soluble odorant carrying proteins enriched in olfactory organs. Here we demonstrate that multiple rhythmically expressed genes including OBPs and takeout proteins, involved in regulating blood feeding behavior, have corresponding rhythmic protein levels as measured by quantitative proteomics. This includes AgamOBP1, previously shown as important to An. gambiae odorant sensing. Further, electrophysiological investigations demonstrate time-of-day specific differences in olfactory sensitivity of antennae to major host-derived odorants. The pre-dusk/dusk peaks in OBPs and takeout gene expression correspond with peak protein abundance at night, and in turn coincide with the time of increased olfactory sensitivity to odorants requiring OBPs and times of increased blood-feeding behavior. This suggests an important role for OBPs in modulating temporal changes in odorant sensitivity, enabling the olfactory system to coordinate with the circadian niche of An. gambiae.

Show MeSH