Sulfur in Plant Physiology
  Uptake, Transport, and Assimilation of Sulfate
    - Foliar Uptake and Metabolism of Sulfurous Gases
  Major Organic Sulfur Compounds
  Secondary Sulfur Compounds
  Interactions between Sulfur and Other Minerals
    - Nitrogen–Sulfur Interactions
    - Interactions between Sulfur and Micronutrients
Sulfur in Plant Nutrition
  Diagnosis of Sulfur Nutritional Status
    - Symptomatology of Single Plants
    - Symptomatology of Monocots
    - Sulfur Deficiency Symptoms on a Field Scale
Soil Analysis
Plant Analysis
  Analytical Methods
  Assessment of Critical Nutrient Values
  Sulfur Status and Plant Health
Sulfur Fertilization

The uptake and transport of sulfate in plants is mediated by sulfate transporter proteins and is energy-dependent (driven by a proton gradient generated by ATPases) through a proton-sulfate (presumably 3H+/SO4 2-) co-transport (14). Several sulfate transporters have been isolated and their genes have been identified. Two classes of sulfate transporters have been identified: the so-called 'high- and low-affinity sulfate transporters,' which operate ideally at sulfate concentrations <0.1 mM and ≥0.1mM, respectively. According to their cellular and subcellular expression, and possible functioning, the sulfate transporter gene family has been classified into as many as five different groups (15,22-24). Some groups are expressed exclusively in the roots or shoots, or in both plant parts. Group 1 transporters are high-affinity sulfate transporters and are involved in the uptake of sulfate by the roots. Group 2 are vascular transporters and are low-affinity sulfate transporters. Group 3 is the so-called 'leaf group;' however, still little is known about the characteristics of this group. Group 4 transporters may be involved in the transport of sulfate into the plastids prior to its reduction, whereas the function of Group 5 sulfate transporters is not yet known. Regulation and expression of the majority of sulfate transporters are controlled by the sulfur nutritional status of the plants. A rapid decrease in root sulfate content upon sulfur deprivation is regularly accompanied by a strongly enhanced expression of most sulfate transporter genes (up to 100-fold), accompanied by a substantial enhanced sulfate uptake capacity. It is still questionable whether, and to what extent, sulfate itself or metabolic products of sulfur assimilation (viz O-acetylserine, cysteine, glutathione) act as signals in the regulation of sulfate uptake by the root and its transport to the shoot, and in the expression of the sulfate tranporters involved (15,22-24).

The major proportion of the sulfate taken up by the roots is reduced to sulfide and subsequently incorporated into cysteine, the precursor and the reduced sulfur donor for the synthesis of most other organic sulfur compounds in plants (16,17,25–27). Even though root plastids contain all sulfate reduction enzymes, reduction predominantly takes place in the chloroplasts of the shoot. The reduction of sulfate to sulfide occurs in three steps (Figure 7.1). First, sulfate is activated to adenosine 5'-phosphosulfate (APS) prior to its reduction, a reaction catalyzed by ATP sulfurylase. The affinity of this enzyme for sulfate is rather low (Km ~1 mM) and the in situ sulfate concentration in the chloroplast may be ratelimiting for sulfur reduction (7). Second, the activated sulfate (APS) is reduced by APS reductase to sulfite, a reaction where glutathione (RSH; Figure 7.1) most likely functions as reductant (17,26). Third, sulfite is reduced to sulfide by sulfite reductase with reduced ferredoxin as reductant. Sulfide is subsequently incorporated into cysteine, catalyzed by O-acetylserine(thiol)lyase, with O-acetylserine as substrate (Figure 7.1).

The formation of O-acetylserine is catalyzed by serine acetyltransferase, and together with O-acetylserine(thiol)lyase it is associated as an enzyme complex named cysteine synthase (28,29). The synthesis of cysteine is a major reaction in the direct coupling between sulfur and nitrogen metabolism in the plant (6,9).

Sulfate reduction and assimilation in plants
FIGURE 7.1 Sulfate reduction and assimilation in plants.

Sulfur reduction is highly regulated by the sulfur status of the plant. Adenosine phosphosulfate reductase is the primary regulation point in the sulfate reduction pathway, since its activity is generally the lowest of the enzymes of the assimilatory sulfate reduction pathway and this enzyme has a fast turnover rate (16,17,26,27). Regulation may occur both by allosteric inhibition and by metabolite activation or repression of expression of the genes encoding the APS reductase. Both the expression and activity of APS reductase change rapidly in response to sulfur starvation or exposure to reduced sulfur compounds. Sulfide, O-acetylserine, cysteine, or glutathione are likely regulators of APS reductase (9,16,17,26). The remaining sulfate in plant tissue is predominantly present in the vacuole, since the cytoplasmatic concentration of sulfate is kept rather constant. In general, the remobilization and redistribution of the vacuolar sulfate reserves is a rather slow process. Under temporary sulfur-limitation stress it may be even too low to keep pace with the growth of the plant, and therefore sulfur-deficient plants may still contain detectable levels of sulfate (13,15,22).

Cysteine is used as the reduced sulfur donor for the synthesis of methionine, the other major sulfur-containing amino acid present in plants, via the so-called trans-sulfurylation pathway (30,31). Cysteine is also the direct precursor for the synthesis of various other compounds such as glutathione, phytochelatins, and secondary sulfur compounds (12,32). The sulfide residue of the cysteine moiety in proteins is furthermore of great importance in substrate binding of enzymes, in metal-sulfur clusters in proteins (e.g., ferredoxins), and in regulatory proteins (e.g., thioredoxins).

Foliar Uptake and Metabolism of Sulfurous Gases

In rural areas the atmosphere generally contains only trace levels of sulfur gases. In areas with volcanic activity and in the vicinity of industry or bioindustry, high levels of sulfurous air pollutants may occur. Sulfur dioxide (SO2) is, in quantity and abundance, by far the most predominant sulfurous air pollutant, but locally the atmosphere may also be polluted with high levels of hydrogen sulfide (18,19,21). Occasionally the air may also be polluted with enhanced levels of organic sulfur gases, viz carbonyl sulfide, methyl mercaptan, carbon disulfide, and dimethyl sulfide (DMS). The impact of sulfurous air pollutants on crop plants appears to be ambiguous. Upon their foliar uptake, SO2 and H2S may be directly metabolized, and despite their potential toxicity used as a sulfur source for growth (18-21). However, there is no clear-cut transition in the level or rate of metabolism of the absorbed sulfur gases and their phytotoxicity, and the physiological basis for the wide variation in susceptibility between plants species and cultivars to atmospheric sulfur gases is still largely unclear (18-21). These paradoxical effects of atmospheric sulfur gases complicate the establishment of cause-effect relationships of these air pollutants and their acceptable atmospheric concentrations in agro-ecosystems.

The uptake of sulfurous gases predominantly proceeds via the stomata, since the cuticle is hardly permeable to these gases (33). The rate of uptake depends on the stomatal and the leaf interior (mesophyll) conductance toward these gases and their atmospheric concentration, and may be described by Fick's law for diffusion

Jgas (pmol cm-2 s-1) = ggas (cm s-1) x Δgas (pmol cm-3)

where Jgas represents the gas uptake rate, ggas the diffusive conductance of the foliage representing the resultant of the stomatal and mesophyll conductance to the gas, and Δgas the gas concentration gradient between the atmosphere and leaf interior (18,20,34). Over a wide range, there is a nearly linear relationship between the uptake of SO2 and the atmospheric concentration. Stomatal conductance is generally the limiting factor for uptake of SO2 by the foliage, whereas the mesophyll conductance toward SO2 is very high (18,20,35). This high mesophyll conductance is mainly determined by chemical/physical factors, since the gas is highly soluble in the water of the mesophyll cells (in either apoplast or cytoplasm). Furthermore, the dissolved SO2 is rapidly hydrated and dissociated, yielding bisulfite and sulfite (SO2 + H2O → H+ + HSO3?2H+ + SO3 2-) (18,20). The latter compounds either directly enter the assimilatory sulfur reduction pathway (in the chloroplast) or are enzymatically or nonenzymatically oxidized to sulfate in either apoplast or cytoplasm (18,20). The sulfate formed may be reduced and subsequently assimilated or it is transferred to the vacuole. Even at relatively low atmospheric levels, SO2 exposure may result in enhanced sulfur content of the foliage (18,20). The liberation of free H+ ions upon hydration of SO2 or the sulfate formed from its oxidation is the basis of a possible acidification of the water of the mesophyll cells, in case the buffering capacity is not sufficient. Definitely, the physical- biochemical background of the phytotoxicity of SO2 can be ascribed to the negative consequences of acidification of tissue/cells upon the dissociation of the SO2 in the aqueous phase of the mesophyll cells or the direct reaction of the (bi)sulfite formed with cellular constituents and metabolites (18,20).

The foliar uptake of H2S even appears to be directly dependent on the rate of its metabolism into cysteine and subsequently into other sulfur compounds, a reaction catalyzed by O-acetylserine (thiol)lyase (19,21). The basis for the phytotoxicity of H2S can be ascribed to a direct reaction of sulfide with cellular components; for instance, metallo-enzymes appear to be particularly susceptible to sulfide, in a reaction similar to that of cyanide (18,19,36).

The foliage of plants exposed to SO2 and H2S generally contains enhanced thiol levels, the accumulation of which depends on the atmospheric level, though it is generally higher upon exposure to H2S than exposure to SO2 at equal concentrations.

Changes in the size and composition of the thiol pool are likely the reflection of a slight overload of a reduced sulfur supply to the foliage. Apparently, the direct absorption of gaseous sulfur compounds bypasses the regulation of the uptake of sulfate by the root and its assimilation in the shoot so that the size and composition of the pool of thiol compounds is no longer strictly regulated.