A Genetically Encoded Fluorescent Heme Sensor Detects Free Heme in Plants.

Heme is produced in plants via a plastid-localized metabolic pathway and is subsequently distributed to all cellular compartments. In addition to covalently and non-covalently bound heme, a comparatively small amount of free heme that is not associated with protein is available for incorporation into heme-dependent proteins in all subcellular compartments and for regulatory purposes. This "labile" fraction may also be toxic. To date, the distribution of the free heme pool in plant cells remains poorly understood. Several fluorescence-based methods for the quantification of intracellular free heme have been described. For this study, we used the previously described genetically encoded heme sensor 1 (HS1) to measure the relative amounts of heme in different plant subcellular compartments. In a proof of concept, we manipulated heme content using a range of biochemical and genetic approaches and verified the utility of HS1 in different cellular compartments of Arabidopsis (Arabidopsis thaliana) and tobacco (N. tabacum and N. benthamiana) plants transformed either transiently or stably with HS1 and HS1(M7A), a variant with lower affinity for heme. This approach makes it possible to trace the distribution and dynamics of free heme and provides relevant information about its mobilization. The application of these heme sensors will create opportunities to explore and validate the importance of free heme in plant cells and to identify mutants that alter the subcellular allocation of free heme.

Changes in morphology, chlorophyll fluorescence performance and Rubisco activity of soybean in response to foliar application of ionic titanium under normal light and shade environment.

Titanium (Ti) is considered an essential element for plant growth; however, its role in crop performance through stimulating the activities of certain enzymes, enhancing chlorophyll content and photosynthesis, and improving crop morphology and growth requires more study. We therefore conducted a laboratory experiments to study the effects of ionic Ti application on morphology, growth, biomass distribution, chlorophyll fluorescence performance and Rubisco activity of soybean (Glycine max L.) under normal light (NL) and shade conditions (SC). In this study, we sprayed soybean plants with five different levels of ionic Ti (T1 = 0, T2 = 1.25, T3 = 2.5, T4 = 5 and T5 = 10 mg Ti Plant-1) through foliar application method. Our results show that with increasing moderate (2.5 mg Ti Plant-1) Ti concentration, the chlorophyll pigments (chlorophyll [Chl] a, b, carotenoid [Car]), plant biomass, photochemical efficiency of photosystem II (Fv/Fm), and electron transport rate (ETR) of soybean increased, but higher levels (5-10 mg Ti Plant-1), resulted in leaf anatomical and chloroplast structural disruptions under both NL and SC. Soybean plants showed maximum biomass, leaf area, leaf thickness, Chl a, b, Car, Rubisco activity, Fv/Fm and ETR for T3 at 2.5 mg Ti Plant-1; however, declined significantly for T5 at high concentration of 10 mg Plant-1. In NL, the application of 2.5 mg Ti Plant-1 (T3) increased the Chl a, b, and total Chl contents 40, 20, and 27% as compared to control treatment (T1). In SC, the application of 1.25 mg Ti mg Plant-1 (T2) increased the Chl a, b, and total Chl contents 38, 19, and 14% as compared to control treatment. In NL, the Fv/Fm, qP, PSII, and ETR were higher in the T3 treatment over the T1 (control) by 7, 0.3, 16, and 16%, respectively. In SC, the Fv/Fm, qP, PSII, and ETR were higher in the T3 treatment over the T1 (control) by 5, 5, 19, and 19%, respectively. Moreover, Rubisco activity was at peak (55 and 6% increase under NL and SC) at 2.5 mg Ti Plant-1and decreased with increasing Ti concentration, reaching the lowest at 10 mg Ti Plant-1, which indicates that leaf cells were damaged as observed in the leaf anatomy. We concluded that ionic Ti expresses a hormesis effect: at lower concentrations, promoting soybean growth, however, at higher concentrations, suppressing soybean growth both under NL and SC. We therefore suggest that under different light stress conditions, Ti application could serve to mitigate abiotic stresses, especially in intercropping systems.