The rice Os06g0683400 was reported as OsCCD1  in 2016 by researchers from Chinese Academy of Sciences. OsCCD1 is a small Ca2+-binding protein with one EF-hand motif in rice. This protein was asmall protein with M.W. 13.9 kDa, pI 5.08.
- OsCCD1 is a small calcium-binding protein with one EF-handmotif.
- OsCCD1 gene enhanced the responsiveness of rice seedlings toexogenous ABA application, and conferred the tolerances of riceseedlings to osmotic and salt stress.
- Semi-quantitative RT-PCR analysis revealed that OsDREB2B, OsAPX1 and OsP5CS genes are involved in the rice tolerance to osmotic and salt stresses. In sum, OsCCD1 gene probably affects the DREB2B and its downstream genes to positively regulate osmotic and salt tolerance in rice seedlings.
- To identify the functions of OsCCD1 gene in the osmoticand salt tolerance, scientists generated three homozygous OsCCD1-overexpressing transgenic rice lines (OD1-1, OD1-2 and OD1-3) in the ZH11 background, and identified twohomozygous T-DNA mutant lines (TD1-1 and TD1-2) in the HY background.
- To detect the tolerance of rice seedlings to osmotic stress,OsCCD1-overexpressing lines, DNA mutant lines and their respec-tive wild type plants, were hydroponically cultured in the 1/4 MSliquid medium to 10 days, followed by the osmotic stress treatment with 20% PEG-6000 for 7 days and then recovery for 7 days. After the osmotic stress treatments, OsCCD1-overexpressing lines showed the significantly higher survival rates (84.6%, 83.2%, and 82.3%)than the wild type ZH11 (77.0%) (p < 0.05), the T-DNA mutant lines showed the significantly lower survival rates (57.8% and 58.3%)than the wild type HY (93.1%) (p < 0.01) (Figure 1).
- In addition,OsCCD1-overexpressing lines had significantly higher Proline (Pro),soluble sugar contents and significantly lower MDA contents in theshoots of rice seedlings than the wild type ZH11 (p < 0.05). However, T-DNA mutant lines had significantly lower levels of solublesugars and Pro but significantly higher MDA contents in the shootsof rice seedlings than the wild type HY (p < 0.05, p < 0.01) (Figure 1), Before the osmotic stress treatment, the chlorophyll contents ofthe shoot tissues did not show significant difference between thethree overexpressing transgenic lines and the wild type ZH11 orbetween two T-DNA insertional mutant lines and the wild type HY. However, osmotic stress treatments decreased the chlorophyll con-tents of all these rice lines. There were significant differences in thechlorophyll contents between the overexpressing lines or T-DNAinsertional mutant lines and their respective wild types (p < 0.01)(Figure 1), suggesting that OsCCD1 genes enhanced the osmotic stresstolerance of rice seedlings and the physiological metabolism.
- When the ten-day-old rice seedlings were treated with 150 mM NaCl for 3 days followed by recovery for 7 days, OsCCD1-overexpressing lines showed significantly higher survival rates(77.4%, 78.2% and 81.1%) than the wild type ZH11 (62.3%) (p < 0.05),the T-DNA mutant lines showed significantly lower survival rates(44.2% and 45.5%) than the wild type HY (64.9%) (p < 0.01) (Figure 2). In addition, OsCCD1-overexpressing lines had significantlybeen higher soluble in sugar and Proline (Pro) contents and signif-icantly higher MDA contents in the shoots of rice seedlings thanthe wild type ZH11 (p < 0.01). However, T-DNA mutant lines hadextravagantly lower contents of soluble sugars and Pro but sig-nificantly higher contents of MDA in the shoots of rice seedlingsthan the wild type HY (p < 0.05, p < 0.01) (Figure 2). Before the saltstress treatments, the chlorophyll contents of the shoot tissues didnot show significant difference between the three overexpressingtransgenic lines and the wild type ZH11 or between two T-DNAinsertional mutant lines and the wild type HY. However, comparedwith their respective wild type rice lines, chlorophyll contentsshowed significant difference between the overexpressing linesor T-DNA insertional mutant lines and their respective wild types(p < 0.05, p < 0.01) (Figure 2), suggesting that OsCCD1 genes enhancedthe salt stress tolerance of rice seedlings and the physiological metabolism.
- OsCCD1 gene was induced by osmotic and salt stress viacalcium-mediated ABA signal, and Ca2+ played as a mediatorin the transduction of the ABA signal in rice seedlings.
- Transient expression ofgreen fluorescent protein (GFP)-tagged OsCCD1 in rice protoplasts showed that OsCCD1 was localized inthe nucleus and cytosol of rice cells in the rice seedlings(Figure 3).
- Phylogenetic and EF-hand motif analysis showed that, basedon the degree of amino acid sequence similarity of different plantspecies, the CCD1 homologues in plants can be categorized intothree groups (Groups I, II and III), containing nine, four and onemembers, respectively. Six homologous CCD1 proteins from mono-cot cereal crops clustered together in the Group I. In addition, OsCCD1 manifested the highest sequence identity with TaCCD1 and BdPBP1, and the conserved one-EF-hand; Group II consisted of genesequences from dicot species, and the Arabidopsis AtKIC was clas-sified as Group III (Figs. S1 and S2). Notably, all of these 14 proteinspossessed one EF-hand domain with the conserved Ca2+-binding loop DxDGDGALxxx, implying that they are conservedin the calcium signal transduction in the plant growth and stressresponses.
- Calcium ion, Ca2+, is adopted as a macroelement in the growthand development process, and a ubiquitous secondary messengerinvolved in the signal transduction of the development and stressresponse in plants. These Ca2+ signaturesare decoded by several types of Ca2+ sensor proteins that contain a high-affinity Ca2+-binding helix-loop-helix structure, known as the EF-hand motif. The binding of Ca2+-binding proteins(CaBPs) triggered a change in conformation and enzymatic activ-ity, followed by the participation of these activated Ca2+/Ca2+ sensor complexes in the induction of appropriate physiological responses, including ion transport, metabolism, post-translational protein modifications and gene expressions.
- Based on the Ca2+-binding affinities and mode of actions, CaBPs can be classified into two groups: (1)Ca2+ sensors, translate thesignal to various responses; (2) Ca2+ buffers, control the level offree Ca2+ ions in the cytoplasm.
Labs working on this gene
- MOA Key Laboratory of Crop Ecophysiology and Farming System in the Middle Reaches of the Yangtze River
- College of Plant Science and Technology
- Huazhong Agricultural University, Wuhan 430070, PR China
- Jing P, Zou J, Kong L, Hu S, Wang B, Yang J, Xie G. OsCCD1, a novel small calcium-binding protein with one EF-hand motif, positively regulates osmotic and salt tolerance in rice. Plant Sci. 2016 Jun;247:104-14. doi: 10.1016/j.plantsci.2016.03.011. Epub 2016 Mar 24. PubMed PMID: 27095404.
- I.S. Day, V.S. Reddy, G. Shad Ali, A.S. Reddy, Analysis of EF-hand-containingproteins in arabidopsis, Genome Biol. 3 (2002), RESEARCH0056.