Study On The Function Of OsCBL5Gene In Rice

Calcium as a second messenger in cells plays an important role in the stress-signaling pathways in plants. The spatial and temporal Ca+signal in plant cells, induced by the different stresses and decoded by many Ca2+-sensors, will activate downstream cascades such as phosphorylation leading to leasing expression of stress-related genes. Calcinerurin B-like proteins (CBLs) are plant-specific calcium banding proteins. CBL-interacting protein kinases (CIPKs) are Ca2+-dependent serine/threonine kinases with a high conserved SNF kinase domain and a NAF motif. CBL/CIPK signaling system has an important function in the response of plants to stresses. Rice is a model monocot plant and also one of the best important crops. The researches on CBL in rice will help us not only better understand molecular mechanisms of CBL/CIPK signal transduction under stresses in plants, but also genetically improve the resistance and productivity of rice or other crops.Transgenic plants (T3) overexpressing sense OsCBL5and OsCBL6gene have been obtained from previous studies. Here, expression patterns of exogenous gene and tolerance to the abiotic stresses including salt and drought were further analyzed in these transgenic lines. Semi-quantitative RT-PCR showed that the expression of OsCBL5and OsCBL6extremely increased in many sense OsCBL5and OsCBL6ransgenic seedlings, respectively. To (?)vestigate the function of OsCBL5and OsCBL6in response to salt and mannitol stress, the seeds of wiio type (WT) and transgenic lines were germinated and grown in medium plus NaCl and mannitol for14days. The results showed that OsCBL5-overexpressing plant showed reduced tolerance to salt and mannitol stresses, and there was no visible difference between wild type and OsCBL6-overexpressing plant in two stresses. Under salt stresses, OsCBL5-overexpressing transgenic plants accumulated significantly higher contents of proline than the wild type. Putative proline synthetase and transporter genes had significantly higher expression level in the transgenic plants than in the wild type. OsCIPK24exhibited a significant interaction with OsCBL4, OsCBL5and OsCBL10in yeast two hybrid interaction analyses. We observed a ring-like distribution of OsCBL5::GFP at the plasma membrane and nucleus. OsCBL5may interact with OsCIPK24that also interact with OsCBL4and OsCBL10. Such an interaction network may cause antagonistic effects on the function of OsCBL4and OsCBL10. These studies suggest that OsCBL5functions as a negative regulator of salt and osmosis responses in plants.To explore the function of OsCBL3under stresses, its open reading frames (ORFs) was cloned into the vector pCABMIA1300S and transformed into the calli of rice variety Nipponbare by Agrobacterium tumefaciens-mediated method. The transgenic regenerated seedlings screened by Hygromycin have been confirmed through PCR assay, and their functional identification will be carried out.