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Endopolyploidy changes with age-related polyethism in the honey bee, Apis mellifera.

Rangel J, Strauss K, Seedorf K, Hjelmen CE, Johnston JS - PLoS ONE (2015)

Bottom Line: We found a nonsignificant increase in ploidy levels with age (P < 0.1) in the most highly endopolyploid secretory cells, the Malpighian tubules.Endopolyploidy decreased the least amount (nonsignificant) in neural (brain) cells and the stinger (P < 0.1).There was a significant reduction of endopolyploidy with age in leg (P < 0.05) and thoracic (P < 0.001) muscles.

View Article: PubMed Central - PubMed

Affiliation: Department of Entomology, Texas A&M University, College Station, Texas 77843, United States of America.

ABSTRACT
Honey bees (Apis mellifera) exhibit age polyethism, whereby female workers assume increasingly complex colony tasks as they age. While changes in DNA methylation accompany age polyethism, other DNA modifications accompanying age polyethism are less known. Changes in endopolyploidy (DNA amplification in the absence of cell division) with increased larval age are typical in many insect cells and are essential in adults for creating larger cells, more copies of essential loci, or greater storage capacity in secretory cells. However, changes in endopolyploidy with increased adult worker age and polyethism are unstudied. In this study, we examined endopolyploidy in honey bee workers ranging in age from newly emerged up to 55 days old. We found a nonsignificant increase in ploidy levels with age (P < 0.1) in the most highly endopolyploid secretory cells, the Malpighian tubules. All other cell types decreased ploidy levels with age. Endopolyploidy decreased the least amount (nonsignificant) in neural (brain) cells and the stinger (P < 0.1). There was a significant reduction of endopolyploidy with age in leg (P < 0.05) and thoracic (P < 0.001) muscles. Ploidy in thoracic muscle dropped from an average of 0.5 rounds of replication in newly emerged workers to essentially no rounds of replication (0.125) in the oldest workers. Ploidy reduction in flight muscle cells is likely due to the production of G1 (2C) nuclei by amitotic division in the multinucleate striated flight muscles that are essential to foragers, the oldest workers. We suggest that ploidy is constrained by the shape, size and makeup of the multinucleate striated muscle cells. Furthermore, the presence of multiple 2C nuclei might be optimal for cell function, while higher ploidy levels might be a dead-end strategy of some aging adult tissues, likely used to increase cell size and storage capacity in secretory cells.

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Least square estimates of mean ploidy levels post-emergence (intercept), and change of ploidy over time (slope) for five different worker honey bee (Apis mellifera) tissues: Brain, thoracic (flight) muscle, leg muscle, Malpighian tubules, and stinger.Adult honey bee workers were collected at different days post emergence (See Methodsfor details).
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pone.0122208.g001: Least square estimates of mean ploidy levels post-emergence (intercept), and change of ploidy over time (slope) for five different worker honey bee (Apis mellifera) tissues: Brain, thoracic (flight) muscle, leg muscle, Malpighian tubules, and stinger.Adult honey bee workers were collected at different days post emergence (See Methodsfor details).

Mentions: Cell nuclei were isolated from each worker’s honey bee stinger and tissue samples as described in Scholes et al. [9]. In brief, tissues were isolated and then ground in 1 mL of Galbraith buffer (45 mM magnesium chloride, 30 mM sodium citrate, 20 mM 4-morpho-G line propane sulfonate, and Triton X-100 (1 mg/mL)) [35] using ten gentle strokes in a 2 mL Kontes Dounce homogenizer using the “A” pestle. The tissues were then filtered using a 40U nylon mesh to remove debris. The resultant solution was brought to 1 mL with additional buffer and was stained using 25 μL Propidium Iodide for 20 minutes in the dark at 4°C. A Partec Cyflow cytometer (Partec America, Swedesboro, NJ) was used to score relative fluorescence and thereby the amount of DNA in the nuclei isolated from each stinger and from each of the other tissue samples. The cytometer was used to score number of nuclei based on amount of PI fluorescence. Histograms were then produced with multiple discrete peaks that represented different fluorescence levels associated with different ploidy levels (Fig 1). The cytometer was adjusted to show diploid (2C) nuclei at channel 25, which made it possible to display 2C, 4C, 8C, 16C, and 32C ploidy levels. The number of nuclei under each ploidy peak was scored using standard Partec software. The few nuclei with ploidy of 64C+ accumulated and were counted in channel 500.


Endopolyploidy changes with age-related polyethism in the honey bee, Apis mellifera.

Rangel J, Strauss K, Seedorf K, Hjelmen CE, Johnston JS - PLoS ONE (2015)

Least square estimates of mean ploidy levels post-emergence (intercept), and change of ploidy over time (slope) for five different worker honey bee (Apis mellifera) tissues: Brain, thoracic (flight) muscle, leg muscle, Malpighian tubules, and stinger.Adult honey bee workers were collected at different days post emergence (See Methodsfor details).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400096&req=5

pone.0122208.g001: Least square estimates of mean ploidy levels post-emergence (intercept), and change of ploidy over time (slope) for five different worker honey bee (Apis mellifera) tissues: Brain, thoracic (flight) muscle, leg muscle, Malpighian tubules, and stinger.Adult honey bee workers were collected at different days post emergence (See Methodsfor details).
Mentions: Cell nuclei were isolated from each worker’s honey bee stinger and tissue samples as described in Scholes et al. [9]. In brief, tissues were isolated and then ground in 1 mL of Galbraith buffer (45 mM magnesium chloride, 30 mM sodium citrate, 20 mM 4-morpho-G line propane sulfonate, and Triton X-100 (1 mg/mL)) [35] using ten gentle strokes in a 2 mL Kontes Dounce homogenizer using the “A” pestle. The tissues were then filtered using a 40U nylon mesh to remove debris. The resultant solution was brought to 1 mL with additional buffer and was stained using 25 μL Propidium Iodide for 20 minutes in the dark at 4°C. A Partec Cyflow cytometer (Partec America, Swedesboro, NJ) was used to score relative fluorescence and thereby the amount of DNA in the nuclei isolated from each stinger and from each of the other tissue samples. The cytometer was used to score number of nuclei based on amount of PI fluorescence. Histograms were then produced with multiple discrete peaks that represented different fluorescence levels associated with different ploidy levels (Fig 1). The cytometer was adjusted to show diploid (2C) nuclei at channel 25, which made it possible to display 2C, 4C, 8C, 16C, and 32C ploidy levels. The number of nuclei under each ploidy peak was scored using standard Partec software. The few nuclei with ploidy of 64C+ accumulated and were counted in channel 500.

Bottom Line: We found a nonsignificant increase in ploidy levels with age (P < 0.1) in the most highly endopolyploid secretory cells, the Malpighian tubules.Endopolyploidy decreased the least amount (nonsignificant) in neural (brain) cells and the stinger (P < 0.1).There was a significant reduction of endopolyploidy with age in leg (P < 0.05) and thoracic (P < 0.001) muscles.

View Article: PubMed Central - PubMed

Affiliation: Department of Entomology, Texas A&M University, College Station, Texas 77843, United States of America.

ABSTRACT
Honey bees (Apis mellifera) exhibit age polyethism, whereby female workers assume increasingly complex colony tasks as they age. While changes in DNA methylation accompany age polyethism, other DNA modifications accompanying age polyethism are less known. Changes in endopolyploidy (DNA amplification in the absence of cell division) with increased larval age are typical in many insect cells and are essential in adults for creating larger cells, more copies of essential loci, or greater storage capacity in secretory cells. However, changes in endopolyploidy with increased adult worker age and polyethism are unstudied. In this study, we examined endopolyploidy in honey bee workers ranging in age from newly emerged up to 55 days old. We found a nonsignificant increase in ploidy levels with age (P < 0.1) in the most highly endopolyploid secretory cells, the Malpighian tubules. All other cell types decreased ploidy levels with age. Endopolyploidy decreased the least amount (nonsignificant) in neural (brain) cells and the stinger (P < 0.1). There was a significant reduction of endopolyploidy with age in leg (P < 0.05) and thoracic (P < 0.001) muscles. Ploidy in thoracic muscle dropped from an average of 0.5 rounds of replication in newly emerged workers to essentially no rounds of replication (0.125) in the oldest workers. Ploidy reduction in flight muscle cells is likely due to the production of G1 (2C) nuclei by amitotic division in the multinucleate striated flight muscles that are essential to foragers, the oldest workers. We suggest that ploidy is constrained by the shape, size and makeup of the multinucleate striated muscle cells. Furthermore, the presence of multiple 2C nuclei might be optimal for cell function, while higher ploidy levels might be a dead-end strategy of some aging adult tissues, likely used to increase cell size and storage capacity in secretory cells.

Show MeSH
Related in: MedlinePlus