Daylength and circadian effects on starch degradation and maltose metabolism

Abstract

Transitory starch is stored during the day inside chloroplasts and broken down at night for export. Maltose is the primary form of carbon export from chloroplasts at night. We investigated the influence of daylength and circadian rhythms on starch degradation and maltose metabolism. Starch breakdown was faster in plants of Arabidopsis (Arabidopsis thaliana) ecotype Wassilewskija growing in long days. Transcript levels of genes encoding enzymes involved in starch degradation and maltose metabolism showed a strong diurnal rhythm. Under altered photoperiods, the transcript levels and the rate of starch degradation changed within one day/night cycle. However, the amount of proteins involved in starch degradation was maintained relatively constant throughout the day/night cycle. To investigate whether the diurnal cycling of the transcript levels is only a response to light or is also regulated by a circadian clock, we measured the amount of messenger RNAs in Arabidopsis leaves under continuous light and continuous darkness. The expression of genes encoding starch degradation-related enzymes was under very strong circadian control in continuous light. Under continuous light, the amount of maltose also showed a strong endogenous rhythm close to 24 h, indicating that maltose metabolism is under circadian control. Light is necessary for the cycling of transcript levels and maltose levels. Under continuous darkness, these genes were barely expressed, and no cycling of maltose levels was observed.

Figure 1.

Diurnal changes of starch in LD, LS, SD, and SL. A, LD (black squares) and LS (black circles). B, SD (white squares) and SL (white circles). White bars and black bars on the top indicate days and nights, respectively. Values are mean ± se (n = 5). FW, fresh weight.

Figure 2.

Diurnal transcript amounts of starch degradation-related genes. A and B, RbcS1A; C and D, DPE2; E and F, AtPHS2; G and H, SEX1; I and J, CT-BMY. Left sections, LD (black squares) and LS (black circles). Right sections, SD (white squares) and SL (white circles). White bars and black bars on the top indicate days and nights, respectively. Asterisks near circles indicate that LS or SL samples (circles) are significantly different from LD or SD samples (squares) collected at the same local time (Student's t test, P = 0.05). Values are mean ± se (n = 3).

Figure 3.

Diurnal protein abundance of RbcS, SEX1, and DPE2. A, Coomassie stains of RbcS as control; B, Western blots of SEX1; C, Western blots of DPE2 in LD, LS, SD, and SL. Numbers on the top are sample-collecting time in hours.

Figure 4.

Diurnal changes of carbohydrate levels in LD, LS, SD, and SL. A and B, β-Maltose; C and D, Glc; E and F, Suc. Left sections, LD (black squares) and LS (black circles). Right sections, SD (white squares) and SL (white circles). White bars and black bars on the top indicate days and nights, respectively. Asterisks near circles indicate that LS or SL samples (circles) are significantly different from LD or SD samples (squares) collected at the same local time (Student's t test, P = 0.05). Values are mean ± se (n = 5).

Figure 5.

Linear regression of maximum nighttime carbohydrate levels with starch degradation rates. A, β-Maltose (r² = 0.996); B, Suc (r² = 0.945). Average rates of starch degradation were estimated by a linear regression of all the nighttime values of starch over the duration of the night. Values on the x axis are slopes and ses of the linear regressions when estimating the rate of starch degradation. Values on y axis are mean ± se (n = 5).

Figure 6.

Relative transcript abundance of starch degradation-related genes in continuous light or continuous darkness. A and B, RbcS1A; C and D, DPE2; E and F, AtPHS2; G and H, SEX1; I and J, CT-BMY. Left sections, Continuous light, white symbols. Right sections, Continuous darkness, black symbols. White bars and gray bars on the top are subjective days and nights. Values are mean ± se (n = 3).

Figure 7.

Protein levels of RbcS, SEX1, and DPE2 in continuous light or continuous darkness. A, Coomassie stains of RbcS; B, Western blots of SEX1; C, Western blots of DPE2. Numbers on the top are hours in continuous light or darkness.

Figure 8.

Carbohydrate levels in continuous light or continuous darkness. A and B, Starch; C and D, maltose; E and F, Glc; G and H, Suc. Left sections, Continuous light, white symbols. Right sections, Continuous darkness, black symbols. White bars and gray bars on the top are subjective days and nights. Values are mean ± se (n = 5).