A Specific Glycogen Mobilization Strategy Enables Rapid Awakening of Dormant Cyanobacteria from Chlorosis

Abstract

Many organisms survive stressful conditions via entry into a dormant state that can be rapidly exited when the stressor disappears; this ability provides a strong selective advantage. In the cyanobacterium sp. PCC 6803, the exit from nitrogen chlorosis takes less than 48 h and is enabled by the impressive metabolic flexibility of these cyanobacteria, which pass through heterotrophic and mixotrophic phases before reentering photoautotrophic growth. Switching between these states requires delicate coordination of carbohydrate oxidation, CO fixation, and photosynthesis. Here, we investigated the contribution of the different carbon catabolic routes by assessing mutants of these pathways during nitrogen chlorosis and resuscitation. The addition of nitrate to nitrogen-starved cells rapidly starts the awakening program. Metabolism switches from maintenance metabolism, characterized by residual photosynthesis and low cellular ATP levels, to an initial heterotrophic phase, characterized by respiration and an immediate increase in ATP levels. This respiration relies on glycogen breakdown catalyzed by the glycogen phosphorylase GlgP2. In the following transient mixotrophic phase, photosynthesis and CO fixation restart and glycogen is consumed. During the mixotrophic phase, parallel operation of the oxidative pentose phosphate cycle and the Entner-Doudoroff pathway is required for resuscitation to proceed; the glycolytic route via the Embden-Meyerhof-Parnas pathway has minor importance. Our data suggest that, during resuscitation, only the Entner-Doudoroff and oxidative pentose phosphate pathways supply the metabolic intermediates necessary for the anabolic reactions required to reconstitute a vegetative cell. Intriguingly, the key enzymes for glycogen catabolism are already expressed during the preceding chlorotic phase, in apparent preparation for rapid resuscitation.

Nitrogen chlorosis in unicellular cyanobacteria – a developmental program for surviving nitrogen deprivation

“…The 77 K spectroscopy revealed substantial PSI fluorescence, whereas PSII fluorescence is reduced to almost undetectable levels in chlorotic cells (Heilmann et al, 2017;Spät et al, 2018). Accordingly, the chlorotic cells maintain constant ATP levels corresponding to about 20%-25% of the level of vegetatively growing cells (Doello et al, 2018). The rod-subunits of the phycobilisomes were reduced to about 0.3% of their initial value, whereas APC subunits, comprising the PBS core, were present at residual levels of about 2%.…”

“…When nitrate is added to fully developed chlorotic cells, the first measurable response is a sudden increase in the level of ATP, which doubles within 15 min (Doello et al, 2018), as well as an almost immediate conversion of nitrate to nitrite (Spät et al, 2018). The very low residual photosynthetic activity, which keeps cells alive during chlorosis (Sauer et al, 2001), is turned off while the cells switch on respiration, regardless of illumination conditions.…”

“…Therefore, the initiation of the resuscitation program is characterized by a switch to a heterotrophic type of metabolism, where glycogen serves as energy and carbon source. Mutant analysis revealed that the cells use a specific set of glycogen catabolic enzymes for the purpose of resuscitation (Doello et al, 2018): only glycogen phosphorylase Glg2 (Slr1367) is able to mobilize glycogen, and the breakdown products are further metabolized via a combination of oxidative pentose phosphate cycle and Entner-Doudoroff (ED) pathways, whereas the glycolytic route via the Emden-Mayerhof-Parnas (EMP) pathway seems to be dispensable. Most of the glycogen catabolic genes are induced upon the induction of chlorosis and are kept at increased levels but in an inactive state throughout chlorosis.…”

“…The molecular biology of chlorotic cells was studied in detail in Synechocystis (Klotz et al, 2016;Heilmann et al, 2017;Doello et al, 2018;Spät et al, 2018). In the chlorotic state, cell growth is arrested and, accordingly, anabolic reactions are almost completely shut down.…”

“…Furthermore, they imply that chlorotic Synechocystis cells maintain viability through cyclic electron transport around PSI. Accordingly, the chlorotic cells maintain constant ATP levels corresponding to about 20%-25% of the level of vegetatively growing cells (Doello et al, 2018). A remarkable and unexpected feature of chlorotic cells is the relatively high abundance of RNA transcripts, in contrast to the almost complete absence of protein synthesis.…”