Abstract: 【Objective】Mitogen-activated protein kinases (MAPKs) play an important role in stress signal transduction process in plant. To provide data for exploring the functional mechanism of TaMAPK2, a wheat cDNA library was constructed and its interacting proteins were screened by yeast two-hybrid system.【Method】The wheat cDNA was used as the template for amplifying the TaMAPK2 gene, and the bait plasmid pGBKT7-TaMAPK2 was constructed. The mixture of recombinant plasmid pGBKT7- TaMAPK2, pGADT7 with wheat cDNA library was introduced into yeast cell AH109. Transformed cells were incubated on SD/-Trp/ -Leu/-His/-Ade plate at 30℃ for 3-5 days before selection of clones and further incubated on SD/Raf/Gal/x-gal for screening blue clones. The candidate proteins that interacted with TaMAPK2 were obtained by sequencing and analyzing the blue clones. Enzyme digestion and ligation method was used to construct the expression vectors of pGADT7-HSP90, pSPYNE-TaMAPK2 and pSPYCE-HSP90. The recombinant plasmids pGADT7-HSP90 and pGBKT7-TaMAPK2 were transformed into yeast competent cells AH109, and the yeast two-hybrid system was used to analyze the interaction between TaMAPK2 and HSP90. The expression vectors pSPYNE-TaMAPK2 and pSPYCE-HSP90 were transformed into wheat protoplasts by using PEG-Ca2+ method. At last the interactions of TaMAPK2 and HSP90 were observed using confocal laser scanning microscope. A real-time PCR testing system was constructed to quantify HSP90 in wheat by the specific primer, designed on the conservative sequences of HSP90 complete genome, and a testing mode based on SYBR Green I technology．The expression patterns of HSP90 gene in response to different abiotic stresses and in different organs were analyzed by quantitative real-time PCR.【Result】 The proteins that interacted with TaMAPK2 were obtained, and most of the candidate proteins involved in signal transduction and immune process. This result suggests that TaMAPK2 possibly plays significant roles in stress signal transduction, transcription regulation of downstream genes and translation process in stress environments. Yeast two-hybrid and BiFc showed that HSP90 protein interacted with TaMAPK2. Subcellular localization analysis revealed that HSP90 is located on the plasma membrane, the cytoplasm and nucleus. The QRT-PCR showed that HSP90 expressed in roots, stems and leaves. The transcripts of HSP90 were more abundant in roots. Real-time PCR showed that the expression of HSP90 was up-regulated by the imposition of high-temperature, low-temperature, high-salt and ABA stresses.【Conclusion】The HSP90 is a widespread family of molecular chaperones. HSP90 can contribute to maintain cellular integrity by folding damaged proteins and maintaining protein conformation. Abiotic stress conditions cause the accumulation of aggregates containing damaged and abnormal proteins in plant. HSP90 is required for the degradation of damaged and misfolded proteins, and mediates expression of transcription factors. This result suggests that TaMAPK2 transmits signals by interacting with other proteins in plant abiotic stress responses.

Key words: common wheat , yeast two-hybrid system , mitogen-activated protein kinases (MAPK) , protein interaction , signal transduction