Project description:In this RNA-seq study, we compared the antennal transcriptomes of sexually mature drones (males) and time-trained foragers (females) of Apis mellifera collected at different times of day and different activity states. The goals of our project was to provide a more comprehensive description of gene expression differences between drone and forager antennae. Most studies on insect antennal transcriptome still focus on identifying and reporting genes involved in odorant binding and detection. In contrast, we also aimed at identifying so far unrecognised molecules, not directly involved in odorant detection, but likely playing an important role in peripheral olfactory processing. We also wanted to explore whether daily changes in gene expression and correlations between gene expression levels and behavioral activity might be a fruitful approach to identify additional genes involved in odorant reception. Apis mellifera drones perform mating flight in the afternoon around 14:00 hour (h) in Bangalore, India. During their daily mating flight activity, drones were caught at the hive entrance and color marked on the thorax. On the next day color-marked drones were collected at two different time points: 9:00 (inactive) and 14:00 h (active). Honey bee foragers can be trained to visit a food source at a specific time of the day. In this study, an A. mellifera colony was transferred in an enclosed outdoor flight cage to train the foragers to visit a feeder at a specific time of the day. A sucrose reward (1M sucrose solution) was presented either from 8:00 to 10:00 h (morning training) or from 13:00 to 15:00 h (afternoon training) for 10 consecutive days. On the 8th, 9th and 10th day of training, foragers visiting the feeder were marked on their thorax with different colors, one type of color each day, to identify the frequently visiting foragers. On the 11th day, the feeder was not presented and the foragers that had all 3 color marks were collected at 9:00 and 14:00 h. All the samples were immediately flash frozen in liquid nitrogen. Collected samples were transferred from liquid nitrogen onto dry ice and the entire antennae (i.e. scape, pedicel and flagellum) were cut off. We pooled 10 antennae from 5 bees per sample and extracted total RNA using Trizol method. Antennal transcriptomes of drones (n=3 per time point), morning-trained foragers (n=2 per time point) and afternoon-trained foragers (n=2 per time point) were sequenced at 2 different time points (9:00 h and 14:00 h).

Project description:Pyrroloquinoline quinone (PQQ) is a naturally occurring compound and known to improve growth and reproductive performance when added to diets of PQQ-deprived rodents. To understand its mechanism action, changes in hepatic gene expression were measured in rats fed diets with or without added PQQ. Gene expression changes for PQQ supplemented (EC50 ~ 3 nmol PQQ/Kg diet) or deficient rats were assayed by expression microarray analysis and compared to those for known transcriptional regulators (epicatechin; Epi; dexamethasone, Dex; clofibrate, Clo; phenobarbital, Pb). Two principal expression clusters were observed; one for Dex and another for Pb, Epi, Clo, and PQQ. Within the latter, a PQQ subcluster containing a unique group of genes for cell signaling and transport functions. Next, short and long term PQQ depletion and repletion protocols (48 or 36 h, respectively) were performed, leading to changes in hepatic gene expression for both time periods. Of the ~10,000 genes and ESTs analyzed, 4.7% of the transcripts were sensitive to changes in PQQ dietary status. PQQ deprivation generally caused down regulation of genes associated with mitochondriogenesis, cell differentiation, and immune function. These gene expression changes provide the basis for most of the previously published functional observations associated with PQQ deficiency and PQQ administered in pharmacological amounts. To assess PQQ’s potential functions, we used gene expression profiling through microarray technology as part of a comprehensive approach to identify potential pathways and mechanisms. Given that the systemic effects of PQQ deprivation are influenced at levels of dietary intake in the micromolar range, highly purified diets were used to reduce expression from other bioactive factors and xenobiotics, such as those found in typical rodent chow diets. To establish if the changes in gene expression in response to PQQ exposure will follow a similar pattern as other xenobiotics, the responses to PQQ exposure were contrasted with those from exposure to epicatechin (Epi), dexamethasone (Dex), clofibrate (Clo), or phenobarbital (Pb). Dietary conditions were also chosen to clarify the response to short- and longer-term PQQ deprivation. A goal was to determine if specific changes in dietary protocol or patterns could be used to identify genes important to the function of PQQ. Because it has been observed that mitochondrial-related functions are influenced by PQQ, we hypothesized that changes in genes important to fatty acid and amino acid metabolism and mitochondrial function would be likewise affected by dietary levels of PQQ. The overall study was carried out using two experiments. In this specific experiment, the gene expression profiles of Sprague Dawley (SD) rats treated with different xenobiotics were compared to the gene expression profile of PQQ treated SD rats.

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