A heavy metal P-type ATPase OsHMA4 prevents copper accumulation in rice grain

Authors: HUANG, XIN-YUAN - University Of Aberdeen; DENG, FENGLIN - Okayama University; Pinson, Shannon; GUERINOT, MARY LOU - Dartmouth College; SALT, DAVID - University Of Aberdeen; MA, JIAN FENG - Okayama University

Submitted to: Nature Communications
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/10/2016
Publication Date: 7/8/2016
Citation: Huang, X., Deng, F., Pinson, S.R., Guerinot, M., Salt, D.E., Ma, J. 2016. A heavy metal P-type ATPase OsHMA4 prevents copper accumulation in rice grain. Nature Communications. 7:121138. doi10.138/ncomms12138.

Interpretive Summary: The objective of this comprehensive study was to identify the gene underlying a QTL on rice chromosome 2 associated with relatively large differences in grain copper concentrations. Rice is one of the most important staple crops in the world, not only providing about one fifth of daily calories for more than half of the world’s population but also providing a critical source of essential mineral nutrients. Mineral nutrients such as calcium, iron, zinc and copper in rice grain are a critical source of these nutrients for humans, but the genetic basis underlying the accumulation of these beneficial elements in rice grain remains largely unknown. Here, we identified a copper transporter that transports copper into root vacuoles, and we showed that natural variation in the function of this transporter (that is to say, different gene alleles that already exist naturally in rice accessions from around the world) affects how much copper gets transferred to and accumulated in the grains of field cultivated rice. While other copper transporters were previously shown to impact other transfer points (such as xylem loading or phloem loading), this is the first copper transporter documented to impact the rate of copper sequestration by transporting it into vacuoles. While other copper transporters had been studied in rice, previous studies all involved singe-gene mutations that caused extreme, generally detrimental changes in plant phenotype. In contrast, this study identified natural variation in the gene from among two high-yielding varieties. This means that the alleles that increase copper concentrations in both the grains are not so extreme as to cause copper toxicity in the plants themselves. The identification of such natural, non-lethal variation among rice genes offers rice breeders the opportunity to fine-tune the copper concentrations in rice grain to match both the level of copper in the soil and the nutritional needs of the people the rice sustains. The study was based on the fact that some elements are known to share uptake and regulatory mechanisms such that a change in the concentration of one element can be predictive of a change in the concentration of the other. Examples of element pairs known to share uptake and regulatory mechanisms are calcium (Ca) and strontium, and rubidium and potassium. This study first evaluated the concentrations of 15 elements in addition to As to determine which individual elements (e.g., [Ca]) or 2-element combinations (e.g. [Ca]/[Cu]) were most strongly associated with grain [As]. Knowledge on how those associated elements are mobilized in the plant was then interpreted relative as to how those mechanisms might also be affecting [As].

Technical Abstract: As one of the most important staple crops, rice not only provides more than one fifth of daily calories for half of the world’s human population but is also a major source of mineral nutrients. However, little is known about the genetic basis of mineral nutrient accumulation in rice grain such as copper (Cu), an essential element for humans but is harmful if consumed in excess. Here, we identified the gene responsible for the quantitative trait locus (QTL) qGCu2 that controls Cu accumulation in the grain of rice. Results showed that qGCu2 encodes a heavy metal P1B-type ATPase OsHMA4 which localizes to the vacuolar membrane where it impacts rate of Cu transport into the vacuoles. OsHMA4 is expressed highly in roots, mainly in the vascular tissues. Expression of OsHMA4 is induced by excess Cu and suppressed by Cu deficiency. Loss of function of OsHMA4 decreased sequestration of Cu in the root vacuoles, leading to increased Cu translocation to the shoots and increased accumulation of Cu in the grain. The naturally occurring alternative alleles of OsHMA4, differed for a single base change and caused a single Valine to Alanine amino acid substitution in the protein, resulting in distinct Cu transporting activities. The function of this gene was further proven by genetically engineering the two rice alleles into Arabidopsis thaliana and yeast where they again cause a difference in the rate of Cu transport into vacuoles. When alleles for this gene were evaluated in a diverse world-wide collection of more than 1200 rice accessions, the Valine to Alanine mutation was found to 8% of the variation observed for grain Cu, and while other nucleotide differences were identified, only the base change found to cause the Valine to Alanine amino acid change in the protein was found associated with difference in grain Cu concentration. The identification of natural allelic variation in OsHMA4 and the functional polymorphism provides an opportunity for the development of rice varieties with grain Cu concentrations tuned to both the concentration of Cu in the soil and dietary needs.