**3. Response of the photosynthetic apparatus to micronutrient depletion**

The photosynthetic machinery comprises several protein–pigment complexes with specific cofactors including iron, copper, manganese and iron–sulfur centers. Under conditions of limitation of one of these micronutrients, the photosynthetic machinery displays a remarkable plasticity and ability to adapt to its new environment.

#### **3.1. Copper deficiency**

When *Chlamydomonas* cells face copper deficiency, the copper-binding protein plastocyanin that acts as an essential electron carrier between the Cyt*b*6*f* complex and PSI is degraded and replaced with Cyt *c6* [100]. In this way, the cells can maintain photosynthetic electron flow. Besides Cyt *c6*, Cpx (coproporphyrinogen oxidase) is also induced by copper deficiency at the transcriptional level [101]. The increase of Cpx1 expression may meet the demand for heme, the cofactor of Cyt *c6.* Crd1, another target besides Cyt *c6* and Cpx1 of this signal transduction pathway responsive to copper depletion, was identified through a genetic screen for a copperconditional phenotype*.* Crd1 is a thylakoid diiron membrane protein which is required for the maintenance of PSI and LHCI in copper-deficient cells. It has an isoform, Cth1, which accu‐ mulates in copper-sufficient oxygenated cells whereas Crd1 accumulates in a reciprocal manner in copper-deficient cells or under anaerobiosis [101]. Crd1 and Cth1 are two isoforms of a subunit of the aerobic cyclase in chlorophyll biosynthesis with overlapping functions in the biosynthesis of Chl proteins [102].

#### **3.2. Iron deficiency**

Iron deficiency occurs often in nature and poses a challenge for photosynthetic organisms because of the abundance and importance of iron in the primary photosynthetic reactions. With its three 4Fe-4S centers, PSI is a prime target under these conditions. Under conditions of iron limitation, the level of PSI decreases when *Chlamydomonas* cells are grown in the presence of a carbon source such as acetate. Eventually, these Fe-deficient cells become chlorotic because of proteolytically-induced loss of both photosystems and Cyt*b*6*f* [103]. Before chlorosis occurs, a graded response is induced, in which the LHCI antenna is dissociated from PSI. This dissociation appears to be caused by the decrease of the amount of the peripheral chlorophyll-binding PsaK subunit of PSI which is required for the functional connection of LHCI to PSI. Interestingly, loss of Crd1, the Fe-requiring aerobic oxidative cyclase in coppersufficient cells, also leads to a lower accumulation of PsaK and to uncoupling of LHCI from PSI. It was proposed that a change in plastid iron content is sensed by the diiron enzyme Crd1 through the occupancy of its Fe-containing active site which determines its activity [103]. In turn, this would affect the flux through the chlorophyll biosynthetic pathway and PsaK stability. This response to Fe deficiency and also to light quantity and quality further involves a remodeling of the antenna complexes with the degradation of specific subunits and the synthesis of new ones leading to a new state of the photosynthetic apparatus which allows for optimal photosynthetic function and minimal photooxidative damage. The protective value of this antenna remodeling is further confirmed by the observation that the light sensitivity of a PsaF-deficient mutant [104] is alleviated in a *psaF-crd1* double mutant [101]. The proposed mechanism can be placed within a general frame for explaining the causal link between chlorosis induced by iron deficiency and loss of photosynthetic function.

D1 degradation is significantly retarded in the *stn8* and *stn7 stn8* mutants of *Arabidopsis* in which the thylakoid membrane architecture is affected [52]. In the absence of STN8, grana diameter is increased and there are fewer grana stacks. This observation is particularly intriguing as it suggests that PSII core phosphorylation is important for maintaining grana size, a parameter which is highly conserved in land plants and algae [11]. Loss of STN8 also affects partitioning of FtsH between grana and stromal membranes and limits its access to the grana, and migration of D1 from the grana to the stroma lamellae is slowed down during the PSII repair cycle [52]. These observations are thus compatible with the view that PSII core phosphorylation has a strong impact on thylakoid membrane folding and architecture mediated most likely by electrostatic repulsion between membrane layers as proposed earlier

**3. Response of the photosynthetic apparatus to micronutrient depletion**

The photosynthetic machinery comprises several protein–pigment complexes with specific cofactors including iron, copper, manganese and iron–sulfur centers. Under conditions of limitation of one of these micronutrients, the photosynthetic machinery displays a remarkable

When *Chlamydomonas* cells face copper deficiency, the copper-binding protein plastocyanin that acts as an essential electron carrier between the Cyt*b*6*f* complex and PSI is degraded and replaced with Cyt *c6* [100]. In this way, the cells can maintain photosynthetic electron flow. Besides Cyt *c6*, Cpx (coproporphyrinogen oxidase) is also induced by copper deficiency at the transcriptional level [101]. The increase of Cpx1 expression may meet the demand for heme, the cofactor of Cyt *c6.* Crd1, another target besides Cyt *c6* and Cpx1 of this signal transduction pathway responsive to copper depletion, was identified through a genetic screen for a copperconditional phenotype*.* Crd1 is a thylakoid diiron membrane protein which is required for the maintenance of PSI and LHCI in copper-deficient cells. It has an isoform, Cth1, which accu‐ mulates in copper-sufficient oxygenated cells whereas Crd1 accumulates in a reciprocal manner in copper-deficient cells or under anaerobiosis [101]. Crd1 and Cth1 are two isoforms of a subunit of the aerobic cyclase in chlorophyll biosynthesis with overlapping functions in

Iron deficiency occurs often in nature and poses a challenge for photosynthetic organisms because of the abundance and importance of iron in the primary photosynthetic reactions. With its three 4Fe-4S centers, PSI is a prime target under these conditions. Under conditions of iron limitation, the level of PSI decreases when *Chlamydomonas* cells are grown in the presence of a carbon source such as acetate. Eventually, these Fe-deficient cells become chlorotic because of proteolytically-induced loss of both photosystems and Cyt*b*6*f* [103]. Before

plasticity and ability to adapt to its new environment.

[99].

**3.1. Copper deficiency**

34 Applied Photosynthesis - New Progress

the biosynthesis of Chl proteins [102].

**3.2. Iron deficiency**

Marine organisms can face iron limitation in the oceans. A deep-sea/ low light strain of the marine green alga *Ostreococcus* has lower photosynthetic activity due to the limited accumu‐ lation of PSI [105]. Interestingly, in this strain, electron flow from PSII is shuttled to a plastid plastoquinol terminal oxidase thereby bypassing electron transfer through the Cyt*b*6*f* complex. This water-to-water cycle allows for the pumping of additional protons to the lumen thylakoid space and thus facilitates ATP production and enhances qE in the case of absorption of excess light excitation energy.

Micronutrient limitation can also act at the level of the biosynthesis of the photosynthetic apparatus which is mediated by the concerted action of the nuclear and chloroplast genetic systems. It is well established that subunits of the photosynthetic complexes originate from these two systems. In addition, a large number of nucleus-encoded factors are required for chloroplast gene expression that act at various plastid posttranscriptional steps comprising RNA processing and stability, translation and assembly of the photosynthetic complexes. Many of these factors have unique gene targets in the plastid and often interact directly or indirectly with specific 5′-untranslated RNA sequences [106]. One of these factors, Taa1 is specifically required for the translation of the PsaA PSI reaction center subunit in *C. reinhard‐ tii* [107]. Under iron limitation, this protein is down-regulated through a posttranscriptional process and it reaccumulates upon restoration of iron. Another recently identified factor is Mac1 which is necessary for stabilization of the *psaC* mRNA [108]. Under iron limitation, both Mac1 protein and *psaC* mRNA are reduced 2-fold and PsaC and PSI are destabilized. Thus, PSI abundance appears to be regulated by iron availability through at least two of these nucleus-encoded plastid factors specifically involved in PSI biosynthesis. Another intriguing observation is that Mac1 is differentially phosphorylated in response to changes in the redox state of the electron transport chain raising questions to what extent posttranslational protein changes modulate the assembly of photosynthetic complexes.

Similar findings have been reported for Mca1 and Tca1, two nucleus-encoded proteins that are required for the stability and translation of the *petA* mRNA encoding the Cyt *f* subunit in *C. reinhardtii*. Nitrogen deprivation leads to the proteolytic degradation of these factors and in turn to the loss of the Cyt*b*6*f* complex [109,110]. The response to nitrogen starvation also involves other factors required for the assembly of the Cyt*b*6*f* complex and its hemes [111].

#### **3.3. Sulfur deprivation and hydrogen production**

Many soil-dwelling algae like *Chlamydomonas* experience anoxic conditions especially during the night and are able to rapidly acclimate to anaerobiosis by shifting from aerobic to fermen‐ tative metabolism and can thus sustain energy production in the absence of photosynthesis [112–114]. These anaerobic conditions lead to the expression of the oxygen-sensitive hydro‐ genase which catalyzes the production of hydrogen from protons and electrons derived from the photosynthetic electron transport chain. Sulfur deprivation of *Chlamydomonas* cells leads to a significant decline in photosynthetic activity within 24 h although there is no proportional concomitant decline in the levels of the major photosynthetic complexes [115,116]. This decline in electron transport activity is due to the conversion of PSII centers from the QB-reducing to a QB nonreducing center [117]. This system has been used for improving hydrogen production in *Chlamydomonas* cells [118]. These cells as well other microalgal species possess a chloroplast (FeFe)-hydrogenase which acts as an additional sink when the photosynthetic electron transport chain is overreduced under anaerobic conditions. Upon sulfur deprivation, photo‐ synthetic oxygen evolution decreases whereas respiration is maintained resulting in an anaerobic environment in a closed culture system. Although the exact physiological role of algal hydrogenases is not known, they are likely to play a significant role in redox poise, photoprotection and fermentative energy production [114].
