Expression of the isiA gene is essential for the survival of the cyanobacterium Synechococcus sp. PCC 7942 by protecting photosystem II from excess light under iron limitation

Iron deficiency is known to suppress primary productivity in both marine and freshwater ecosystems. In response to iron deficiency, certain cyanobacteria induce a chlorophyll (Chl)-protein complex, CP43', which is encoded by the isiA gene. The deduced amino-acid sequence of CP43' predicts some structural similarity to the CP43 polypeptide of photosystem II, but the function of CP43' remains uncertain. In order to assess its physiological role, the isiA gene of a cyanobacterium, Synechococcus sp. PCC7942, was inactivated by insertion mutagenesis (giving isiA cells). Compared with isiA cells, under iron deprivation, wild-type cells showed both lower rates of photosystem II-mediated O2 evolution at limiting light irradiances and decreased yields of room temperature Chl fluorescence at various irradiances. These observations strongly suggest that the decreased photosystem II activity in wild-type cells with CP43' is attributable to increased non-radiative dissipation of light energy. In agreement with this hypothesis, isiA cells were more susceptible to photoinhibition of photosynthesis than wild-type cells, resulting in much slower growth rates under iron limitation. Based on these results, we suggest that CP43' functions as a non-radiative dissipator of light energy, thus protecting photosystem II from excessive excitation under iron-deficient conditions.     

pkn22 (alr2502) encoding a putative Ser/Thr kinase in the cyanobacterium Anabaena sp. PCC 7120 is induced by both iron starvation and oxidative stress and regulates the expression of isiA

In cyanobacteria, the isiA gene is required for cell adaptation to oxidative damage caused by the absence of iron. We show here that a putative Ser/Thr kinase gene, pkn22 (alr2052), is activated by iron deficiency and oxidative damage in Anabaena sp. PCC 7120. A pkn22 insertion mutant is unable to grow when iron is limiting. pkn22 regulates the expression of isiA (encoding CP43'), but not of isiB (encoding flavodoxin) and psbC (CP43). Fluorescence measurement at 77 K reveals the absence of the typical signature of CP43' associated with photosystem I in the mutant under iron-limiting conditions. We propose that Pkn22 is required for the function of isiA/CP43' and constitutes a regulatory element necessary for stress response.     

Differential regulation of nramp and irt metal transporter genes in wild type and iron uptake mutants of tomato

Metal transporters regulated by iron can transport a variety of divalent metals, suggesting that iron regulation is important for specificity of iron transport. In plants, the iron-regulated broad-range metal transporter IRT1 is required for uptake of iron into the root epidermis. Functions of other iron-regulated plant metal transporters are not yet established. To deduce novel plant iron transport functions we studied the regulation of four tomato metal transporter genes belonging to the nramp and irt families with respect to environmental and genetic factors influencing iron uptake. We isolated Lenramp1 and Lenramp3 from tomato and demonstrate that these genes encode functional NRAMP metal transporters in yeast, where they were iron-regulated and localized mainly to intracellular vesicles. Lenramp1 and Leirt1 revealed both root-specific expression and up-regulation by iron deficiency, respectively, in contrast to Leirt2 and Lenramp3. Lenramp1 and Leirt1, but not Lenramp3 and Leirt2, were down-regulated in the roots of fer mutant plants deficient in a bHLH gene regulating iron uptake. In chloronerva mutant plants lacking the functional enzyme for synthesis of the plant-specific metal chelator nicotianamine Leirt1 and Lenramp1 were up-regulated despite sufficient iron supply independent of a functional fer gene. Lenramp1 was expressed in the vascular root parenchyma in a similar cellular pattern as the fer gene. However, the fer gene was not sufficient for inducing Lenramp1 and Leirt1 when ectopically expressed. Based on our results, we suggest a novel function for NRAMP1 in mobilizing iron in the vascular parenchyma upon iron deficiency in plants. We discuss fer/nicotianamine synthase-dependent and -independent regulatory pathways for metal transporter gene regulation.     

Iron deficiency in cyanobacteria causes monomerization of photosystem I trimers and reduces the capacity for state transitions and the effective absorption cross section of photosystem I in vivo

The induction of the isiA (CP43') protein in iron-stressed cyanobacteria is accompanied by the formation of a ring of 18 CP43' proteins around the photosystem I (PSI) trimer and is thought to increase the absorption cross section of PSI within the CP43'-PSI supercomplex. In contrast to these in vitro studies, our in vivo measurements failed to demonstrate any increase of the PSI absorption cross section in two strains (Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803) of iron-stressed cells. We report that iron-stressed cells exhibited a reduced capacity for state transitions and limited dark reduction of the plastoquinone pool, which accounts for the increase in PSII-related 685 nm chlorophyll fluorescence under iron deficiency. This was accompanied by lower abundance of the NADP-dehydrogenase complex and the PSI-associated subunit PsaL, as well as a reduced amount of phosphatidylglycerol. Nondenaturating polyacrylamide gel electrophoresis separation of the chlorophyll-protein complexes indicated that the monomeric form of PSI is favored over the trimeric form of PSI under iron stress. Thus, we demonstrate that the induction of CP43' does not increase the PSI functional absorption cross section of whole cells in vivo, but rather, induces monomerization of PSI trimers and reduces the capacity for state transitions. We discuss the role of CP43' as an effective energy quencher to photoprotect PSII and PSI under unfavorable environmental conditions in cyanobacteria in vivo.     

A novel mammalian iron-regulated protein involved in intracellular iron metabolism

We have isolated and characterized a novel iron-regulated gene that is homologous to the divalent metal transporter 1 family of metal transporters. This gene, termed metal transporter protein (mtp1), is expressed in tissues involved in body iron homeostasis including the developing and mature reticuloendothelial system, the duodenum, and the pregnant uterus. MTP1 is also expressed in muscle and central nervous system cells in the embryo. At the subcellular level, MTP1 is localized to the basolateral membrane of the duodenal epithelial cell and a cytoplasmic compartment of reticuloendothelial system cells. Overexpression of MTP1 in tissue culture cells results in intracellular iron depletion. In the adult mouse, MTP1 expression in the liver and duodenum are reciprocally regulated. Iron deficiency induces MTP1 expression in the duodenum but down-regulates expression in the liver. These data indicate that MTP1 is an iron-regulated membrane-spanning protein that is involved in intracellular iron metabolism.     

Structural analysis of the photosystem I supercomplex of cyanobacteria induced by iron deficiency

Here we describe the three-dimensional structure of the newly discovered CP43'-photosystem I (PSI) supercomplex of cyanobacteria calculated by single-particle analysis of images obtained by electron cryomicroscopy (cryo-EM). This large membrane protein complex has a molecular mass of approximately 2 MDa and is found in cyanobacteria when grown in iron deficient media. It is composed of a reaction center trimer surrounded by 18 subunits of the chlorophyll a binding CP43'protein, encoded by the isiA gene, which increases the light harvesting capacity of PSI by approximately 70%. By modeling higher-resolution structural data obtained from X-ray crystallography into the three-dimensional (3D) cryo-EM map, we have been able to gain a better understanding of the structure and functional properties of this supermolecular complex. We have identified three separate clusters of chlorophyll molecules at the periphery of the PSI core which may aid energy transfer from the CP43' antenna ring to the reaction center. Moreover, it is shown that despite the replacement of ferredoxin with flavodoxin as an electron acceptor under iron stress conditions, the 3D map has density to accommodate the extrinsic proteins, PsaC, PsaD, and PsaE. The presence of these three proteins was also confirmed by immunoblotting.     

The role of cellular iron deficiency in controlling iron export

BackgroundIron export via the transport protein ferroportin (Fpn) plays a critical role in the regulation of dietary iron absorption and iron recycling in macrophages. Fpn plasma membrane expression is controlled by the hepatic iron-regulated hormone hepcidin in response to high iron availability and inflammation. Hepcidin binds to the central cavity of the Fpn transporter to block iron export either directly or by inducing Fpn internalization and lysosomal degradation. Here, we investigated whether iron deficiency affects Fpn protein turnover.MethodsWe ectopically expressed Fpn in HeLa cells and used cycloheximide chase experiments to study basal and hepcidin-induced Fpn degradation under extracellular and intracellular iron deficiency.Conclusions/General significanceWe show that iron deficiency does not affect basal Fpn turnover but causes a significant delay in hepcidin-induced degradation when cytosolic iron levels are low. These data have important mechanistic implications supporting the hypothesis that iron export is required for efficient targeting of Fpn by hepcidin. Additionally, we show that Fpn degradation is not involved in protecting cells from intracellular iron deficiency.