Changes in protein expression during honey bee larval development

Abstract

Background: The honey bee (Apis mellifera), besides its role in pollination and honey production, serves as a model for studying the biochemistry of development, metabolism, and immunity in a social organism. Here we use mass spectrometry-based quantitative proteomics to quantify nearly 800 proteins during the 5- to 6-day larval developmental stage, tracking their expression profiles.

Results: We report that honey bee larval growth is marked by an age-correlated increase of protein transporters and receptors, as well as protein nutrient stores, while opposite trends in protein translation activity and turnover were observed. Levels of the immunity factors prophenoloxidase and apismin are positively correlated with development, while others surprisingly were not significantly age-regulated, suggesting a molecular explanation for why bees are susceptible to major age-associated bee bacterial infections such as American Foulbrood or fungal diseases such as chalkbrood. Previously unreported findings include the reduction of antioxidant and G proteins in aging larvae.

Conclusion: These data have allowed us to integrate disparate findings in previous studies to build a model of metabolism and maturity of the immune system during larval development. This publicly accessible resource for protein expression trends will help generate new hypotheses in the increasingly important field of honey bee research.

Figure 1

The peptide ratios within an experiment are roughly normally distributed and show no labeling bias. Using replicate number 1 of day 1 versus day 3 solid tissue quantification data as an example, the peptide ratios are displayed as a histogram, sorted into natural-log unit bins (bin size = 1).

Figure 2

PAGE of honey bee larvae (a) hemolymph and (b) solid tissue. Age is shown in days post-hatching. Molecular weight markers are shown on the left.

Figure 3

Developmental changes of larval hemolymph. The left axis denotes the volume of hemolymph per larva (diamonds; μl) or hemolymph protein concentration (squares; μg/μl), while the right axis describes the mass of total protein per larva (triangles; μg). Measurements were made by pooling 5-120 larva (n = 3 separate pools) depending on age (x-axis, in days) and size. (Error bars represent 2 standard deviations.)

Figure 4

Average-linkage clustering of proteins quantified in the honey bee larvae. Proteins that were quantified in either or both the (a) tissue or (b) hemolymph for at least four out of five tested days were arranged by hierarchical clustering using software described in [46]. All expression values, shown relative to day 3 (= 0, black), have been natural log-transformed (>0, red; <0, green; no data, grey). These proteins, which have been manually annotated with a function and category, are calculated for enrichment within a node (results in Table 3) if the node correlation value is >0.8 (see thick bar on scale).

Figure 5

Expression profiles of four selected proteins during larval development. Expression levels (y-axis, expressed in natural log scale) over 5 days of larval growth (x-axis) are shown for 3 proteins discussed in the text: (a) imaginal disc growth factor [GenBank:66514614], (b) odorant binding protein 14 [GenBank:94158822], (c) apismin [GenBank:58585112]. Error bars represent one standard deviation.