Fig 1.
Regulatory components of iron homeostasis in non-graminaceous plant cells.
In the presence of iron deficiency the transcription factor FIT activates the excretion of H+ by ATPases AHA2/7, which leads to an increased solubility of Fe(III). FIT also induces the reduction of solubilized Fe(III) to Fe(II) by the membrane-bound enzyme FRO2 and activates the high-affinity transporter IRT1 and the uptake of Fe(II). Iron is stored in different organelles with the vacuole as a major store. Several transporters which move iron into organelles and the vacuole have been identified (see main text). Iron transport to other parts of the plant occurs in complexed form with the water flow, i.e. via the cytosolic symplast connected by plasmodesmata and perhaps also by a transport route using a vacuolar symplast. Transporter candidates for iron remobilization from the store are Nramp3/4.
Fig 2.
Negative Feedback Loops with and without Integral Control.
(a) Flow diagram illustrating the concept of integral control. The regulated value of A is compared with its set-point Aset and the integral E of the error e between A and its set-point is calculated. E is fed into the process to compensate uncontrolled inflow or outflow to and from A. (b) Scheme of an inflow controller, where integral control is represented by removing E with enzyme Eset, which is saturated with substrate E and reflected by a low value. (c) Illustration of robust homeostasis in A for different k1, k2 combinations with set-point
Eq (6). The change in k1 and k2 occurs at t = 50.0 and t = 100.0 time units indicated by the arrows. Rate constants: k3 = 1.0, k4 = 2.0,
, and
. Initial concentrations: A0 = 2.0, and E0 = 3.0. (d) Same negative feedback loop as in (b), but without integral control. The saturating kinetics of the E-removal is now replaced by a first-order process with respect to E with k5 = 1.0. The system is now not able to maintain robust homeostasis in A. Initial concentrations and the other rate constants are as in (c).
Fig 3.
Regulatory loop of high affinity iron uptake and cytosolic iron homeostasis based on an iron-dependent IRT1 degradation.
(a) Scheme of the control loop. Feext and Fecyt denote external and cytosolic iron, respectively. IRT1 and IRT1 denote mRNA and its protein, respectively. The extracellular iron concentration, Feext, is allowed to change to different but constant levels. (b) Homeostasis in cytosolic iron levels (Fecyt) with respect to sufficient and low external iron conditions. The set-point for the level of cytosolic iron is given by Eq (12) and is arbitrarily set to . In the first phase (time t = 0 to t = 50) Feext = 5.0 and relative high. To keep iron at its homeostatic set-point during this phase the required concentration of IRT1 is relative low. In the second phase starting at t = 50 (arrow) Feext is reduced to 0.5. Due to this reduction the IRT1 level is increased to keep the cytosolic iron concentration close at its set-point. In the third phase (t = 100 to t = 150) iron is resupplied and IRT1 levels decrease again. Other rate parameters remain unchanged during the three phases, i.e. k1 = 1.0, k2 = 2.0, k3 = 1.0 × 102, k4 = 1.0 × 102, k5 = 1.0 × 102,
,
, and
. Initial concentrations are Fecyt,0 = 1.0, IRT10 = 1.0, and IRT10 = 0.4. (c) Representation of results in a ‘blot-like’ manner. +Fe and −Fe denote sufficient and low external ion conditions, respectively. For each component (IRT1, IRT1) the gray levels (0–100%) reflect the relative IRT1/IRT1 concentrations at +Fe and −Fe conditions. (d) Experimental data, slightly rearranged from Fig 6A in Ref. [21].
Fig 4.
IRT1 overexpression leads to an increased iron set-point, iron overload and to saturation in the iron-dependent degradation of IRT1.
(a) Increase of IRT1 synthesis rate k3 at different external iron levels. At times t = 50 and t = 100 (indicated by arrows) the values of Feext and k3 are changed as indicated in the figure. As long as the IRT1 syntesis rate jIRT1-synth is lower than its degradation rate (t = 0 to t = 100), the cytosolic iron concentration is under homeostatic control at its new set-point Eq (12). When jIRT1-synth becomes larger than jIRT1-degr iron levels rise and the IRT1 degradation rate jIRT1-degr goes into saturation (t = 100 to t = 150). The negative feedback loop is broken and iron homeostasis lost. (b) Demonstration of iron-independent degradation of IRT1 when jIRT1-synth > jIRT1-degr. The overexpression rate (k3) is kept constant at 1 × 103 while external iron concentrations Feext are changing. For each Feext value (5.0, 0.5, and 0.01) the IRT1 degradation rate is at its maximum value and independent of the cytosolic iron concentration. Rate constants, except k3, are as in Fig 3c. (c) Calculated IRT1 expression levels shown as “dot-blots”. (d) Corresponding experimental results by Barberon et al. (Fig 1D in [23]).
Fig 5.
Model for iron uptake including IRT1 and FIT.
(a) Reaction scheme of the model. See S3 Text for rate equations. (b) Overview over the feedback structure of the model at the protein and cytosolic iron levels. The inhibition outlined in blue defines an additional auxiliary negative feedback which does not influence the set-point of cytosolic iron, but accelerates the adaptation kinetics of the controller (see (d)). (c) Regulation of IRT1- and FIT-mRNA and protein levels and response kinetics of the iron uptake system at low (-Fe) and high (+Fe) external iron concentrations and in the absence of the auxiliary feedback (). The lower part of the panel shows the IRT1- and FIT-mRNA and protein levels in a blot-like representation, where high levels under -Fe conditions have a gray scale of 100%, while +Fe levels have a reduced gray scale in relation to their reduced numerical values. (d) Same as in (c), but now in the presence of the auxiliary feedback (
). Note the improvement in the adaptation kinetics of the system. Rate constants for (c) and (d): k1 = 1.0, k2 = 2.0, k3 = 1 × 102, k4 = 1.0, k6 = 4 × 102, k8 = 1 × 102,
, k11 = 1 × 103, k12 = 1 × 103,
, k18 = 1 × 102, k19 = 1 × 101, k20 = 1 × 104, k21 = 2 × 104,
, k25 = 4.0,
,
. Initial concentrations for (c): Fecyt0 = 1.0, IRT10 = 4.0,
, FIT0 = 381.0, TF0 = 0.5, FIT⋅TF0 = 1905.0,
. Initial concentrations for (d): Fecyt0 = 1.0, IRT10 = 4.0,
, FIT0 = 5.6, TF0 = 0.5, FIT⋅TF0 = 28.0,
. (e) Experimental results of IRT1 and FIT mRNA and protein levels in wild-type Arabidopsis roots under iron sufficient (“+Fe”) and iron deficient (“-Fe”) conditions. The IRT1-protein and mRNA results as well as the FIT-mRNA blot are reproduced with permission from Fig 1 of Ref. [12]. The FIT-protein Western blot is reproduced with permission from Fig 4A of Ref. [47].
Fig 6.
Model of plant iron homeostasis integrating uptake, storage, assimilation/transport and remobilization from the store.
(a) The model combines a low-affinity iron uptake based on an iron-dependent derepression mechanism of inhibitor S [53], which leads to iron storage (outlined in blue), an R-based iron remobilization mechanism from the store (outlined in ochre), the FIT-based high-affinity iron uptake mechanism from Fig 5a (outlined in green) and a lumped expression for the iron assimilation and transport flux to other parts of the plant (outlined in red). Note the renumbering for some of the rate constants in comparison with Fig 5a. See S4 Text for rate equations. (b) Calculation showing cytosolic and external iron concentrations during the low- and high-affinity uptake of iron and iron remobilization from the vacuole. (c) Same calculation as in (b), but showing the different iron fluxes. Rate constants and initial concentrations for (b) and (c): k1 = 1.0, k2 = 2.0, k3 = 1 × 102, k4 = 1.0, k6 = 4 × 102, k7 = 1 × 102, k8 = 1 × 102, k9 = 15.0, ,
,
, k13 = 0.5, k14 = 0.8,
,
, k19 = 0.5, k22 = 5 × 10−4,
, k24 = 4.0,
,
, k27 = 1 × 103, k28 = 1 × 103, k29 = 1 × 104; k30 = 2 × 104, k32 = 1 × 102, k33 = 10.0,
,
. Feext,0 = 10.0, all other initial concentrations are zero.
Fig 7.
Iron-based derepression mechanism of iron storage is not active at low external iron concentrations.
The figure shows that the iron-based derepression mechanism for iron storage (Fig 6a) cannot activate iron storage at low external iron concentrations when the IRT1 high-affinity uptake system is active. In this calculation the external iron concentration Feext is kept constant at 1.0. Initial concentrations and rate constants for this calculation are as in Fig 6b and 6c, except for k22 (= 0.1) which accounts for a symplastic removal of iron out of the vacuole and leads to a decrease in Festore. The FIT and IRT1-based high affinity uptake is able to keep the cytosolic iron at its set-point , while the S-based outflow controller (Fig 6a), responsible for iron storage, is inactive.
Fig 8.
Iron storage during high-affinity uptake.
(a) The model is an extension of that in Fig 6a containing in addition inflow controller molecule I inside the vacuole which activates a transporter located in the vacuolar membrane. Controller molecule I is subject to an iron-dependent degradation. The rate equation for I is: (b) Calculation showing the increase of iron in the vacuole, Festore, as a function of time. The iron set-point inside the vacuole is given by Eq (31) and set to 700.0. The flux of iron entering the vacuole due to controller I is given as:
. The additional parameter values are: k17 = 1 × 10−3, k18 = 700.0,
,
. Other parameter values and initial concentrations as in Fig 6. The initial concentration of controller I is zero.