Molecular and Biochemical Analysis of Periplastidial Starch Metabolism in the Cryptophyte Guillardia theta

Abstract

Starch in synchronously grown Guillardia theta cells accumulates throughout the light phase, followed by a linear degradation during the night. In contrast to the case for other unicellular algae such as Chlamydomonas reinhardtii, no starch turnover occurred in this organism under continuous light. The gene encoding granulebound starch synthase (GBSS1), the enzyme responsible for amylose synthesis, displays a diurnal expression cycle. The pattern consisted of a maximal transcript abundance around the middle of the light phase and a very low level during the night. This diurnal regulation of GBSS1 transcript abundance was demonstrated to be independent of the circadian clock but tightly light regulated. A similar yet opposite type of regulation pattern was found for two ␣-amylase isoforms and for one of the two plastidic triose phosphate transporter genes investigated. In these cases, however, the transcript abundance peaked in the night phase. The second plastidic triose phosphate transporter gene had the GBSS1 mRNA abundance pattern. Quantification of the GBSS1 activity revealed that not only gene expression but also total enzyme activity exhibited a maximum in the middle of the light phase. To gain a first insight into the transport processes involved in starch biosynthesis in cryptophytes, we demonstrated the presence of both plastidic triose phosphate transporter and plastidic ATP/ADP transporter activities in proteoliposomes harboring either total membranes or plastid envelope membranes from G. theta. These molecular and biochemical data are discussed with respect to the environmental conditions experienced by G. theta and with respect to the unique subcellular location of starch in cryptophytes.

The Toxoplasma Apicoplast Phosphate Translocator Links Cytosolic and Apicoplast Metabolism and Is Essential for Parasite Survival

“…In contrast, the red alga Galdieria sulphuraria possesses only a TPT and a PPT (Linka et al, 2008). Equivalent transport activities have also been detected in the plastid of the cryptophyte Guillardia theta which is also derived from red algae by secondary endosymbiosis, however, these activities have not yet been assigned to a particular protein (Haferkamp et al, 2006). Given the presence of both a PPT and TPT in red algae, it is likely that the ancestor of the apicoplast inherited multiple, specific pPTs.…”

Shotgun proteomic analysis of the unicellular alga Ostreococcus tauri

“…For example, starch content in Chlamydomonas reinhardtii oscillates even in constant darkness, suggesting that this is a clock-regulated system [28]. However, in the unicellular alga Guillardia theta, this free-running behaviour is not observed, and an evening-phased day-dark transition has been shown to be essential to trigger starch synthesis [32]. Differentially treated O. tauri cultures were grown in media containing either 14 N or 15 N, and pooled at equal densities at harvest for metabolic labeling experiments.…”

Intracellular distribution of the reductive and oxidative pentose phosphate pathways in two diatoms

“…The same phenotype was observed in a presequence:GFP fusion of the cryptophyte Guillardia theta UDP glucose-starch glycosyl transferase (UGGtransferase), when heterologously ex-pressed in the diatom P. tricornutum [65]. UGGtransferase links glucose to starch, and in cryptophytes, starch is stored in the periplastidic space as the main carbohydrate storage product [69,70]. This indicates that the "blob"-like structure phenotype might represent an accumulation of GFP in the periplastidic space of diatoms [65].…”

“…This process would also provide energy for metabolite transport (as proton anti-or symport) across plastid membranes. In cryptophytes, starch is synthesised and stored within the periplastidic compartment [69,70], but diatoms do not accumulate starch [14]. The identification of a PGDH that is putatively targeted to the periplastidic space further stimulates speculations on the role of the periplastidic space in carbon concentration and metabolite delivery to the plastid.…”