Abstract
Graminaceous plants take up iron through YS1 (yellow stripe 1) and YS1-like (YSL) transporters using iron-chelating compounds known as mugineic acid family phytosiderophores. We examined the expression of 18 rice (Oryza sativa L.) YSL genes (OsYSL1-18) in the epidermis/exodermis, cortex, and stele of rice roots. Expression of OsYSL15 in root epidermis and stele was induced by iron deficiency and showed daily fluctuation. OsYSL15 restored a yeast mutant defective in iron uptake when supplied with iron(III)-deoxymugineic acid and transported iron(III)-deoxymugineic acid in Xenopus laevis oocytes. An OsYSL15-green fluorescent protein fusion was localized to the plasma membrane when transiently expressed in onion epidermal cells. OsYSL15 promoter-beta-glucuronidase analysis revealed that OsYSL15 expression in roots was dominant in the epidermis/exodermis and phloem cells under conditions of iron deficiency and was detected only in phloem under iron sufficiency. These results strongly suggest that OsYSL15 is the dominant iron(III)-deoxymugineic acid transporter responsible for iron uptake from the rhizosphere and is also responsible for phloem transport of iron. OsYSL15 was also expressed in flowers, developing seeds, and in the embryonic scutellar epithelial cells during seed germination. OsYSL15 knockdown seedlings showed severe arrest in germination and early growth and were rescued by high iron supply. These results demonstrate that rice OsYSL15 plays a crucial role in iron homeostasis during the early stages of growth.
Highlights
Ϳ50% of all anemia cases can be attributed to iron deficiency [1]
Plants grown on calcareous soils often exhibit severe chlorosis because of iron deficiency, which is a major agricultural problem resulting in reduced crop yields [2]
To estimate possible functions of the 18 OsYSL genes, we performed RT-PCR in the epidermis/exodermis, cortex, and stele of iron-sufficient and -deficient roots (Fig. 1)
Summary
Plant Materials—Nontransgenic and transgenic rice seeds were germinated on Murashige and Skoog (MS) medium and transferred to nutrient solution [11, 22] in a greenhouse with 30 °C light/25 °C dark periods under natural light conditions. Iron-sufficient and iron-deficient roots were fixed by a 10-min infiltration of 3:1 ethanol:acetic acid into the tissues under vacuum on ice. The vials containing the samples in the fixative were gently mixed on a rotator at 4 °C for 1 h. A 2019-bp fragment of OsYSL15 full-length cDNA was amplified by PCR using a cDNA pool prepared from iron-deficient rice roots and primers 5Ј-TCGTGGGAATTCTCGAGCAGCTAAGCGAGATCGACGC-3Ј and 5Ј-TTTATTTCTAGAATCCTCCACCCATGAAATTAAACAC-3Ј. A probe specific for OsYSL15 was amplified by PCR using a cDNA pool prepared from iron-deficient rice roots and the same primers as those used for the RT-PCR analysis of microdissected tissues. Quantitative RT-PCR Analysis—Quantitative RT-PCR analysis of roots and seedlings was carried out as described previously [21] using the specific primers shown in supplemental Table S1. For quantitative RT-PCR analysis of OsYSL15, seeds were first germinated on MS medium with 10% (v/v) of Tetsuriki-aqua for 6 days and transplanted to iron-free MS medium for 6 days
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