Metabolic interplay between cytosolic phospho<i>enol</i>pyruvate carboxylase and mitochondrial alternative oxidase in thermogenic skunk cabbage, <i>Symplocarpus renifolius</i>

<p>Skunk cabbage (<i>Symplocarpus renifolius</i>) blooms in early spring and its inflorescence, referred to as the spadix, can produce enough heat to melt snow. Here, we investigated glycolytic carbon flow at the PEP branch-point in thermogenic spadices. Our analyses revealed that petals and pistils in thermogenic florets exhibited higher expression of <i>SrPEPC</i> and <i>SrAOX</i> transcripts than those of <i>SrPK, SrPEPCK</i>, and <i>SrPEPtase</i>. Moreover, enzymatic analyses showed high activities of PEPC in the extracts from thermogenic florets. Finally, mitochondria from thermogenic florets showed low respiratory activities when pyruvate was used as a substrate, although a significant malate-mediated cyanide-insensitive respiration was observed. Collectively, these results suggest that PEP metabolism, primarily catabolized by PEPC, plays a critical role in thermogenesis in <i>S. renifolius</i>.</p

Expression of the <i>AmGR10</i> candidate in nurse and forager bees.

<p>(A) Photographs show HPGs from a nurse bee 7 days after adult emergence (upper) and from a forager bee collected randomly outside of the hive (hive 1) (lower). Scale bars = 0.5 mm. (B) RT-PCR analyses of <i>AmGR10</i> expression in nurse and forager bees and at each developmental stage. Upper gels show results from both adult castes in (A). Lower gels show results from 300 eggs within 24 h after oviposition, and 50 larvae and 10 pupae collected randomly between June and October of 2007 and 2008 (hive 1). HPGs taken from 20 nurse bees at 7 days after mergence and 20 forager bees collected randomly outside of the hive were dissected and pooled sample used for RT-PCR analyses. (C) RT-PCR analyses of <i>AmGR10</i> expression in HPGs during the transition between nurse and forager. HPGs were dissected from 20 workers of each indicated age (painted workers collected 0 to 14 days after mergence were identified as nurses, and those collected at 19, 24, and 29 days were identified as foragers) and pooled for RT-PCR analyses. We performed at least three RT-PCR analyses for each pooled sample.</p

Expression and function of <i>AmGR10</i> in honeybees.

<p>(A) Artificial expression of <i>AmGR10</i> in the nursing state. In October 2007, we collected nurse bees 0 and 7 days old (the latter from hives 1 and 2). In early November, we collected 29-day-old workers in an extended nursing state within the hive and foragers outside of the hive at the same time (hive 2). Twenty HPGs from each group were pooled for RT-PCR analyses. (B) Organ distribution of <i>AmGR10</i> expression in 20 HPGs, 150 ovaries, 150 mandibular glands, 150 salivary glands, 20 midguts, and 20 cerebral brains from 7-day-old nurse bees between July and October of 2007 and 2008 (hive 2). Organ preparations from each group were pooled for RT-PCR analyses. (C) Knockdown of <i>AmGR10</i> mRNA in the HPGs of 7-day-old nurse bees. One hundred bees in 4 independent experiments (25 bees × 4) injected with <i>ds AmGR10</i> (8.2 μg⁄2 μL, “RNAi”), 80 bees in 4 independent experiments (20 bees × 4) injected with nuclease-free water (2 μL, “MQ”), and 80 untreated bees in 4 independent experiments (20 bees × 4) (“Mock”) were placed back into their hives (hives 1, 2, and 3) immediately after treatment and were collected again 24 h later for qRT-PCR. For qRT-PCR analyses, 3 bees were used in 4 independent experiments (3 bees × 4) and the remaining bees were allowed to forage. All values are means ± SEM. *** <i>P</i> < 0.001 compared with each control. (D) Effect of RNAi on foraging behavior. We ascertained the age in each group from paint markings: 22 bees × 4 in “RNAi”, 17 bees × 4 in “MQ”, and 17 bees × 4 in “Mock”. We observed the bees over 3 consecutive days outside of the same hive (see RNAi section in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142917#sec002" target="_blank">Materials and Methods</a>) and their flights to ensure the onset of flight hazard were shown as cumulative numbers of bees initiating foraging, but thereafter we did not follow their survival or labors. All values are means ± SEM. *** <i>P</i> < 0.001 compared with each control.</p

Expression of <i>AmGR10</i> of the Gustatory Receptor Family in Honey Bee Is Correlated with Nursing Behavior

<div><p>We investigated the association between the expression of a gene encoding gustatory receptor (G10) and division of labor in the honey bee, <i>Apis mellifera</i>. Among 10 GR genes encoding proteins 15% ~ 99% amino acid identity in the honey bee, we found that <i>AmGR10</i> with 99% identity is involved in nursing or brood care. Expression of <i>AmGR10</i> was restricted to organs of the hypopharyngeal gland, brain, and ovary in the nurse bee phase. Members of an extended nursing caste under natural conditions continued to express this gene. RNAi knockdown of <i>AmGR10</i> accelerated the transition to foraging. Our findings demonstrate that this one gene has profound effects on the division of labor associated with the development and physiology of honeybee society.</p></div