Immunogold Localization of Vitellogenin in the Ovaries, Hypopharyngeal Glands and Head Fat Bodies of Honeybee Workers, Apis Mellifera

Vitellogenin is a yolk precursor protein in most oviparous females. In the advanced eusocial honeybee, Apis mellifera (Hymenoptera: Apidae), vitellogenin has recently attracted much interest as this protein, in addition to a classical function in oocyte development in the reproductive queen caste, has evolved functions in the facultatively sterile female worker caste not documented in other species. However, research on the spatial dynamics of vitellogenin in various tissues is not easily performed with available tools. Here we present an immunogold staining procedure that visualizes honeybee vitellogenin in resin embedded tissue. To establish the protocol, we used ovaries of worker bees from colonies with and without a queen. Under the first condition, vitellogenin is assumed not to be present in the workers' ovaries. Under the second condition, the ovaries of worker bees become vitellogenic, with abundant opportunities for detection of complex patterns of vitellogenin uptake and storage. By use of this experimental setup, the staining method is shown to be both sensitive and specific. To demonstrate the functional significance of the protocol, it was subsequently used to identify vitellogenin protein in the hypopharyngeal glands (brood food producing head glands) of nursing worker bees and in adjacent head fat body cells for the first time. Localization of vitellogenin in these tissues supports previously hypothesized roles of vitellogenin in social behavior. This protocol thus provides deeper insights into the functions of vitellogenin in the honeybee

Somatic Maintenance Resources in the Honeybee Worker Fat Body Are Distributed to Withstand the Most Life-Threatening Challenges at Each Life Stage

<div><p>In a global transcriptome analysis of three natural and three manipulated honeybee worker phenotypes at different ages, we have investigated the distribution of investment in somatic maintenance of the fat body. Gene expression is modulated so that the bees are able to resist the most life-threatening challenges at the actual life stage. Different modes of maintenance and repair are regulated, apparently to meet the environmental challenges most detrimental to survival and reproductive potential for the hive. We observed a broad down-regulation of genomic and cellular maintenance in the short-lived foragers and nurse bees compared to the long-lived winter bees. Our results show that survival and reproduction of the entire hive is given priority over the individual bees, hence supporting the idea of the honeybee society as a superorganism. Our results also fit the disposable soma theory of aging.</p></div

Expressed genes within the insulin-insulin like signalling pathway.

<p>Comparison between worker phenotypes, only expressed genes with a fold change above 2 was considered. Abbreviations: F (foragers), N (nurse bees), Wb (winter bees).</p

Number of differently expressed genes with a cut-off of q = 10% in all comparisons of worker phenotypes.

<p>Number of differently expressed genes with a cut-off of q = 10% in all comparisons of worker phenotypes.</p

qPCR validation of SOLiD whole transcriptome data.

<p>There was a very good correlation between the experimental data and the qPCR in all comparisons apart from nurse bees versus <i>vg</i>knockdowns. Omitting this data point gave a correlation of 0.93435774 for the rest of the genes in this comparison.</p

The resulting extraction of 12 clusters from the unsupervised hierarchical clustering analysis of all 8640 transcripts.

<p>Only foragers, nurses and the winter bee pools were included in the analysis. The x axis denotes the fold change and the y axis the individual bees within each worker group. Two clusters were up-regulated in winter bee pools (a, e), whereas four clusters were down-regulated in this worker phenotype (f, g, h and i). Four clusters were up-regulated in nurse bees (b, c, j, and l); one cluster was up-regulated in foragers (k) and one down-regulated (d). Abbreviations: (F1-5) foragers, (N1-5) nurses bee, (Wp1-5) winter bee pools, (FC) fold change.</p