CRISPR-mediated promoter editing of a cis-regulatory element of OsNAS2 increases Zn uptake/translocation and plant yield in rice

Yvonne Ludwig; Conrado Dueñas Jr; Erwin Arcillas; Reena Jesusa Macalalad-Cabral; Ajay Kohli; Russell Reinke; Inez H Slamet-Loedin

doi:10.3389/fgeed.2023.1308228

CRISPR-mediated promoter editing of a cis-regulatory element of OsNAS2 increases Zn uptake/translocation and plant yield in rice

Front Genome Ed. 2024 Jan 23:5:1308228. doi: 10.3389/fgeed.2023.1308228. eCollection 2023.

Authors

Yvonne Ludwig¹, Conrado Dueñas Jr², Erwin Arcillas¹, Reena Jesusa Macalalad-Cabral¹, Ajay Kohli¹, Russell Reinke¹, Inez H Slamet-Loedin¹

Affiliations

¹ International Rice Research Institute, Rice Genetic Design and Validation Unit, Rice Breeding Innovations, Los Baños, Philippines.
² Department of Biology and Biotechnology "L. Spallanzani", University of Pavia, Pavia, Italy.

Abstract

Developing nutritious rice with a higher yield is one approach to alleviating the problem of micronutrient deficiency in developing countries, especially human malnutrition involving zinc and iron (Fe) deficiency, and achieving better adoption. The transport of micronutrients such as Fe and Zn is mainly regulated via the nicotianamine synthase (OsNAS) gene family, whereas yield is a complex trait that involves multiple loci. Genome editing via CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9, focusing on the OsNAS2 promoter, particularly the deletion of the cis-regulatory element ARR1AT at position -933, was conducted for an enhanced accumulation of Zn in the grain and per plant. The results showed that our promoter editing increased Zn concentration per plant. Evidence also showed that an improved spikelet number per main panicle led to increased grain per plant. The traits were inherited in "transgene-free" and homozygous plant progenies. Further investigation needs to be conducted to validate trait performance under field conditions and elucidate the cause of the spikelet increase.

Keywords: CRISPR-Cas9; Fe; OsNAS2 promoter; Zn; biofortification; promoter editing; rice.

Grants and funding

The author(s) declare that financial support was received for the research, authorship, and/or publication of this article. This research was funded by HarvestPlus: Developing high-iron and high-zinc transgenic rice to alleviate iron deficiency in Bangladesh and Southeast Asia (Agreement No. 2015H5315) and the United States Agency for International Development under the contribution agreement CGIAR (MTO No. 069033) designated to the IRRI OneRice project—Healthier Rice.