Mechanism Research On The Nucleo-cytoplasmic Trafficking And Salt Tolerance Of OsTZF7

Tandem CCCH zinc finger protein(TZF)is a subfamily of CCCH zinc finger proteins.It is an important component of processing bodies(PBs)and stress granules(SGs),which is important for the growth,development and the stress response in animals and plants by post-transcriptional regulation of target mRNA.Therefore,analyzing the mechanism of TZF protein is of the greatest importance for the enhancement of plants' stress tolerance,the promotion of crops' growth and yield.To characterize the OsTZF7 function in salt stress,we performed salt stress tolerance assay.The results revealed that OsTZF7OE lines showed significant salt tolerance,whereas OsTZF7Cas9 lines showed a hypersensitive phenotype,when processed by 120 mM NaCl for 3 days.After re-watering with 1/2 MS liquid medium for 4 days,the survival rate of OsTZF7OE lines was considerably higher than wild type lines,on the contrary,OsTZF7Cas9 lines were fewer to be survived than wild type lines.Furthermore the H2O2 which induced by salt stress,was accumulated in OsTZF7Cas9 lines but not in OsTZF70E lines.Finally,by checking the chlorophyll contents,we found that there were higher chlorophyll contents in OsTZF70E lines and less in OsTZF7Cas9 lines compared to wild type lines.These results indicate that OsTZF7 positively regulates salt tolerance in rice.Subcellular localization analysis found that OsTZF7 localized in the cytoplasm and cytoplasmic foci,and also co-localized with PBs and SGs markers.Sequence analysis showed that OsTZF7 contained both nuclear localization signal(NLS)and nuclear export signal(NES).After truncating the NES,OsTZF7 was localized in the nuclei.Furthermore,when treated with LMB,OsTZF7 was accumulated in the nuclei of the OsTZF7 transiently transformed tobacco,which indicating that the nuclear export of OsTZF7 is depended on XPO1/CRMl protein and OsTZF7 may function in both cytoplasm and nucleus.In addition,OsTZF7AC protein which was removed the NES,was used for yeast library screening.Three candidate interacting proteins,Os02g0700000,Os04g0599100,and Os01g0558500 were all belong to the PWWP family.BiFC and Pull down experiments were carried out in vitro and in vivo,respectively,to prove that these three proteins can interact with OsTZF7.In summary,OsTZF7 is mainly located in the cytoplasm and cytoplasmic foci which formed by PBs and SGs,and it shuttle between nuclear and cytoplasm via XPO1/CRM1.OsTZF7 positively regulates salt response in rice,overexpression of OsTZF7 dramatically improves salt tolerance of rice.Moreover,OsTZF7 interacted with PWWP family proteins.It is possible that OsTZF7 interacted with PWWP proteins and regulate stress-related genes expression to improve the salt tolerance in rice.