title: Impairing both HMA4 homeologs is required for cadmium reduction in tobacco.
abstract: In tobacco, the heavy metal P1B-ATPases HMA4.1 and HMA4.2 function in root-to-shoot zinc and cadmium transport. We present greenhouse and field data that dissect the possibilities to impact the two homeologous genes in order to define the best strategy for leaf cadmium reduction. In a first step, both genes were silenced using an RNAi approach leading to >90% reduction of leaf cadmium content. To modulate HMA4 function more precisely, mutant HMA4.1 and HMA4.2 alleles of a Targeting Induced Local Lesions IN Genomes (TILLING) population were combined. As observed with RNAi plants, knockout of both homeologs decreased cadmium root-to-shoot transfer by >90%. Analysis of plants with segregating null and wild-type alleles of both homeologs showed that one functional HMA4 allele is sufficient to maintain wild-type cadmium levels. Plant development was affected in HMA4 RNAi and double knockout plants that included retarded growth, necrotic lesions, altered leaf morphology and increased water content. The combination of complete functional loss (nonsense mutation) in one homeologous HMA4 gene and the functional reduction in the other HMA4 gene (missense mutation) is proposed as strategy to limit cadmium leaf accumulation without developmental effects.
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Verena
authors.identifier: 0000-0003-2120-9594
authors.initials: V
authors.lastname: Liedschulte
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Hélène
authors.initials: H
authors.lastname: Laparra
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: James Nicolas Duncan
authors.initials: JN
authors.lastname: Battey
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Joanne Deborah
authors.initials: JD
authors.lastname: Schwaar
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Hervé
authors.initials: H
authors.lastname: Broye
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Régis
authors.initials: R
authors.lastname: Mark
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Markus
authors.initials: M
authors.lastname: Klein
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Simon
authors.initials: S
authors.lastname: Goepfert
authors.affiliation: PMI R&D, Philip Morris Products SA (part of Philip Morris International group of companies), Neuchâtel, Switzerland.
authors.forename: Lucien
authors.initials: L
authors.lastname: Bovet
mesh_terms: D029681:Arabidopsis Proteins / Q000737:chemistry; D002104:Cadmium / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D008954:Models, Biological; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D018515:Plant Leaves / Q000378:metabolism; D010940:Plant Proteins / Q000378:metabolism*; D030821:Plants, Genetically Modified; D034622:RNA Interference; D017386:Sequence Homology, Amino Acid; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism
chemical_list: D029681:Arabidopsis Proteins; D010940:Plant Proteins; D002104:Cadmium; D015032:Zinc
keywords: NtHMA4; TILLING population; field study; heavy metal ATPase; zinc deficiency