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Characterization of Reproductive Dormancy in Male Drosophila melanogaster

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ABSTRACT

Insects are known to respond to seasonal and adverse environmental changes by entering dormancy, also known as diapause. In some insect species, including Drosophila melanogaster, dormancy occurs in the adult organism and postpones reproduction. This adult dormancy has been studied in female flies where it is characterized by arrested development of ovaries, altered nutrient stores, lowered metabolism, increased stress and immune resistance and drastically extended lifespan. Male dormancy, however, has not been investigated in D. melanogaster, and its physiology is poorly known in most insects. Here we show that unmated 3–6 h old male flies placed at low temperature (11°C) and short photoperiod (10 Light:14 Dark) enter a state of dormancy with arrested spermatogenesis and development of testes and male accessory glands. Over 3 weeks of diapause we see a dynamic increase in stored carbohydrates and an initial increase and then a decrease in lipids. We also note an up-regulated expression of genes involved in metabolism, stress responses and innate immunity. Interestingly, we found that male flies that entered reproductive dormancy do not attempt to mate females kept under non-diapause conditions (25°C, 12L:12D), and conversely non-diapausing males do not mate females in dormancy. In summary, our study shows that male D. melanogaster can enter reproductive dormancy. However, our data suggest that dormant male flies deplete stored nutrients faster than females, studied earlier, and that males take longer to recover reproductive capacity after reintroduction to non-diapause conditions.

No MeSH data available.


Dormant flies display alterations in carbohydrate and lipid stores. (A) Hemolymph glucose is increased after 3 weeks of dormancy (D3) and 3 weeks of recovery from diapause (R3). (B) Hemolymph trehalose is not altered during dormancy. (C) Whole body (stored) glucose is low after 1 week of (D1) and then increases (D3) during dormancy. (D) Whole body trehalose is not significantly affected by dormancy, but elevated in recovering flies (R3). (E) Stored glycogen is lower in flies dormant for 1 week (D1). (F) Triacylglycerid (TAG) content first increases (D1) and then decreases (D3) during dormancy. After 1 (R1) and 3 (R3) weeks of recovery TAG levels increased back to control levels. Data significantly different from that in the C1 control flies or between groups indicated with connectors are shown: *p < 0.05, **p < 0.01, ***p < 0.001 (ANOVA followed with Tukey test) or alternatively with #p < 0.05, ##p < 0.01 (Kruskal–Wallis test followed by pairwise comparisons using Wilcoxon rank sum test). N.S. Not significant. Data represent 6 replicates with 10–15 flies in each replicate, n = 60–90 flies.
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Figure 4: Dormant flies display alterations in carbohydrate and lipid stores. (A) Hemolymph glucose is increased after 3 weeks of dormancy (D3) and 3 weeks of recovery from diapause (R3). (B) Hemolymph trehalose is not altered during dormancy. (C) Whole body (stored) glucose is low after 1 week of (D1) and then increases (D3) during dormancy. (D) Whole body trehalose is not significantly affected by dormancy, but elevated in recovering flies (R3). (E) Stored glycogen is lower in flies dormant for 1 week (D1). (F) Triacylglycerid (TAG) content first increases (D1) and then decreases (D3) during dormancy. After 1 (R1) and 3 (R3) weeks of recovery TAG levels increased back to control levels. Data significantly different from that in the C1 control flies or between groups indicated with connectors are shown: *p < 0.05, **p < 0.01, ***p < 0.001 (ANOVA followed with Tukey test) or alternatively with #p < 0.05, ##p < 0.01 (Kruskal–Wallis test followed by pairwise comparisons using Wilcoxon rank sum test). N.S. Not significant. Data represent 6 replicates with 10–15 flies in each replicate, n = 60–90 flies.

Mentions: Extended lifespan during diapause is enabled by a strong regulation of metabolism and energy storage (Hahn and Denlinger, 2011). In addition, increased nutrient reserves are critical for post-diapause fitness (Hahn and Denlinger, 2007, 2011). Previously we found that dormant D. melanogaster females have lower body mass and display increased circulating and stored carbohydrates and triacylglycerides (TAG) compared to non-dormant ones (Kubrak et al., 2014). We find here that D. melanogaster males kept in dormancy for the same duration display a lower body weight (Figure S2) and a slight hyperglycemia, with hemolymph glucose levels higher than in C1 controls (Figure 4A). However, the circulating trehalose concentration was not affected by dormancy (Figure 4B). Whole-body levels of glucose are higher after 3 weeks of dormancy than in controls, but decreased to the control level during recovery (R1, R3) (Figure 4C). Whole body trehalose is slightly higher in males after 3 weeks of dormancy and, in contrast to glucose, did not decrease after recovery (Figure 4D). The main stored carbohydrate, glycogen, is lower at the beginning of dormancy (D1), compared to non-dormant males, then increased somewhat, but still remained at a slightly lower level than in control flies (Figure 4E). Surprisingly, the dormant male D3 flies exhausted stored TAG (Figure 4F), while females kept high levels of TAG over 12 weeks of diapause conditions (Kubrak et al., 2014). Rapid use of stored nutrients might result in a shorter dormancy in males than in females. Indeed, a shorter duration of male adult diapause was seen in several insect species (reviewed in Pener, 1992).


Characterization of Reproductive Dormancy in Male Drosophila melanogaster
Dormant flies display alterations in carbohydrate and lipid stores. (A) Hemolymph glucose is increased after 3 weeks of dormancy (D3) and 3 weeks of recovery from diapause (R3). (B) Hemolymph trehalose is not altered during dormancy. (C) Whole body (stored) glucose is low after 1 week of (D1) and then increases (D3) during dormancy. (D) Whole body trehalose is not significantly affected by dormancy, but elevated in recovering flies (R3). (E) Stored glycogen is lower in flies dormant for 1 week (D1). (F) Triacylglycerid (TAG) content first increases (D1) and then decreases (D3) during dormancy. After 1 (R1) and 3 (R3) weeks of recovery TAG levels increased back to control levels. Data significantly different from that in the C1 control flies or between groups indicated with connectors are shown: *p < 0.05, **p < 0.01, ***p < 0.001 (ANOVA followed with Tukey test) or alternatively with #p < 0.05, ##p < 0.01 (Kruskal–Wallis test followed by pairwise comparisons using Wilcoxon rank sum test). N.S. Not significant. Data represent 6 replicates with 10–15 flies in each replicate, n = 60–90 flies.
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Figure 4: Dormant flies display alterations in carbohydrate and lipid stores. (A) Hemolymph glucose is increased after 3 weeks of dormancy (D3) and 3 weeks of recovery from diapause (R3). (B) Hemolymph trehalose is not altered during dormancy. (C) Whole body (stored) glucose is low after 1 week of (D1) and then increases (D3) during dormancy. (D) Whole body trehalose is not significantly affected by dormancy, but elevated in recovering flies (R3). (E) Stored glycogen is lower in flies dormant for 1 week (D1). (F) Triacylglycerid (TAG) content first increases (D1) and then decreases (D3) during dormancy. After 1 (R1) and 3 (R3) weeks of recovery TAG levels increased back to control levels. Data significantly different from that in the C1 control flies or between groups indicated with connectors are shown: *p < 0.05, **p < 0.01, ***p < 0.001 (ANOVA followed with Tukey test) or alternatively with #p < 0.05, ##p < 0.01 (Kruskal–Wallis test followed by pairwise comparisons using Wilcoxon rank sum test). N.S. Not significant. Data represent 6 replicates with 10–15 flies in each replicate, n = 60–90 flies.
Mentions: Extended lifespan during diapause is enabled by a strong regulation of metabolism and energy storage (Hahn and Denlinger, 2011). In addition, increased nutrient reserves are critical for post-diapause fitness (Hahn and Denlinger, 2007, 2011). Previously we found that dormant D. melanogaster females have lower body mass and display increased circulating and stored carbohydrates and triacylglycerides (TAG) compared to non-dormant ones (Kubrak et al., 2014). We find here that D. melanogaster males kept in dormancy for the same duration display a lower body weight (Figure S2) and a slight hyperglycemia, with hemolymph glucose levels higher than in C1 controls (Figure 4A). However, the circulating trehalose concentration was not affected by dormancy (Figure 4B). Whole-body levels of glucose are higher after 3 weeks of dormancy than in controls, but decreased to the control level during recovery (R1, R3) (Figure 4C). Whole body trehalose is slightly higher in males after 3 weeks of dormancy and, in contrast to glucose, did not decrease after recovery (Figure 4D). The main stored carbohydrate, glycogen, is lower at the beginning of dormancy (D1), compared to non-dormant males, then increased somewhat, but still remained at a slightly lower level than in control flies (Figure 4E). Surprisingly, the dormant male D3 flies exhausted stored TAG (Figure 4F), while females kept high levels of TAG over 12 weeks of diapause conditions (Kubrak et al., 2014). Rapid use of stored nutrients might result in a shorter dormancy in males than in females. Indeed, a shorter duration of male adult diapause was seen in several insect species (reviewed in Pener, 1992).

View Article: PubMed Central - PubMed

ABSTRACT

Insects are known to respond to seasonal and adverse environmental changes by entering dormancy, also known as diapause. In some insect species, including Drosophila melanogaster, dormancy occurs in the adult organism and postpones reproduction. This adult dormancy has been studied in female flies where it is characterized by arrested development of ovaries, altered nutrient stores, lowered metabolism, increased stress and immune resistance and drastically extended lifespan. Male dormancy, however, has not been investigated in D. melanogaster, and its physiology is poorly known in most insects. Here we show that unmated 3&ndash;6 h old male flies placed at low temperature (11&deg;C) and short photoperiod (10 Light:14 Dark) enter a state of dormancy with arrested spermatogenesis and development of testes and male accessory glands. Over 3 weeks of diapause we see a dynamic increase in stored carbohydrates and an initial increase and then a decrease in lipids. We also note an up-regulated expression of genes involved in metabolism, stress responses and innate immunity. Interestingly, we found that male flies that entered reproductive dormancy do not attempt to mate females kept under non-diapause conditions (25&deg;C, 12L:12D), and conversely non-diapausing males do not mate females in dormancy. In summary, our study shows that male D. melanogaster can enter reproductive dormancy. However, our data suggest that dormant male flies deplete stored nutrients faster than females, studied earlier, and that males take longer to recover reproductive capacity after reintroduction to non-diapause conditions.

No MeSH data available.