Calcium (Ca2+) is one of the essential mineral nutrients for plant growth and development. However, the effects of long-term Ca2+ deficiency in orphan crops such as Tef [(Eragrostis tef) (Zucc.) Trotter], which accumulate high levels of Ca in the grains, remained unknown. Tef is a staple crop for nearly 70 million people in East Africa, particularly in Ethiopia and Eritrea. It is one of the most nutrient-dense grains, and is also more resistant to marginal soils and climatic conditions than main cereals like corn, wheat, and rice. In this study, tef plants were grown in a hydroponic solution containing optimum (1 mM) or low (0.01 mM) Ca2+, and plant growth parameters and whole-genome transcriptome were analyzed. Ca+2-deficient plants exhibited leaf necrosis, leaf curling, and growth stunting symptoms. Ca2+ deficiency significantly decreased root and shoot Ca, potassium (K), and copper content in both root and shoots. At the same time, it greatly increased root iron (Fe) content, suggesting the role of Ca2+ in the uptake and/or translocation of these minerals. Transcriptomic analysis using RNA-seq revealed that members of Ca2+ channels, including the cyclic nucleotide-gated channels and glutamate receptor-like channels, Ca2+-transporters, Ca2+-binding proteins and Ca2+-dependent protein kinases were differentially regulated by Ca+2 treatment. Moreover, several Fe/metal transporters, including members of vacuolar Fe transporters, yellow stripe-like, natural resistance-associated macrophage protein, and oligo-peptide transporters, were differentially regulated between shoot and root in response to Ca2+ treatment. Taken together, our findings suggest that Ca2+ deficiency affects plant growth and mineral accumulation by regulating the transcriptomes of several transporters and signaling genes.

MicroRNAs (miRNAs) play an important role in growth, development, stress resilience, and epigenetic modifications of plants. However, the effect of calcium (Ca 2+ ) deficiency on miRNA expression in the orphan crop tef ( Eragrostis tef ) remains unknown. In this study, we analyzed expression of miRNAs in roots and shoots of tef in response to Ca 2+ treatment. miRNA‐seq followed by bioinformatic analysis allowed us to identify a large number of small RNAs (sRNAs) ranging from 17 to 35 nt in length. A total of 1380 miRNAs were identified in tef experiencing long‐term Ca 2+ deficiency while 1495 miRNAs were detected in control plants. Among the miRNAs identified in this study, 161 miRNAs were similar with those previously characterized in other plant species and 348 miRNAs were novel, while the remaining miRNAs were uncharacterized. Putative target genes and their functions were predicted for all the known and novel miRNAs that we identified. Based on gene ontology (GO) analysis, the predicted target genes are known to have various biological and molecular functions including calcium uptake and transport. Pairwise comparison of differentially expressed miRNAs revealed that some miRNAs were specifically enriched in roots or shoots of low Ca 2+ ‐treated plants. Further characterization of the miRNAs and their targets identified in this study may help in understanding Ca 2+ deficiency responses in tef and related orphan crops.

Calcium (Ca2+) is one of the essential mineral nutrients for plant growth and development. However, the effects of long-term Ca2+ deficiency in orphan crops such as tef (Zucc.) Trotter], which accumulate high levels of Fe in the grains, remained unknown. Tef is a staple crop for nearly 70 million people in East Africa, particularly in Ethiopia and Eritrea. It is rich in mineral nutrients (Ca, Fe, Zn, and Mn), vitamins, and essential amino acids. In this study, tef plants were grown in a hydroponic solution containing optimum (1 mM) or low (0.01 mM) Ca2+, and plant growth parameters and whole-genome transcriptome were analyzed. Ca+2-deficient plants exhibited leaf necrosis, leaf curling, and growth stunting symptoms. Ca2+ deficiency significantly decreased root and shoot Ca, potassium (K), and copper content in both root and shoots. At the same time, it greatly increased root iron (Fe2+) content, suggesting the role of Ca2+ in the uptake and/or translocation of these minerals. Transcriptomic analysis using RNA-seq revealed that members of Ca2+ channels, including the cyclic nucleotide-gated channels (CNGC) and glutamate receptor-like channels (GLRs), Ca2+-transporters, Ca2+-binding proteins (CBP) and Ca2+-dependent protein kinases (CDPK) were differentially regulated by Ca+2 treatment. Moreover, several Fe/metal transporters, including members of vacuolar Fe transporters (VIT), yellow stripe-like (YSL), natural resistance-associated macrophage protein (NRAMP), and oligo-peptide transporters (OPT), were differentially regulated between shoot and root in response to Ca2+ treatment. Taken together, our findings suggest that Ca2+ deficiency affects plant growth and mineral accumulation by regulating the transcriptomes of several transporters and signaling genes.