Proteomic Study Of Thylakoid Membrane From A Chlorophyll B Less Rice Mutant

Two-dimensional blue native-SDS-PAGE and isoelectric focusing-SDS-PAGE have been used to compare the thylakoid proteome between the low chlorophyll b mutant(Oryza sativa L. var.Zhenhui 249Y) and the wild type (Oryza sativa L. var. Zhenhui 249W), which was followed by MALDI-TOF and LC-MS/MS identification. Some of the proteins were verified by immuno detection of whole chloroplast proteins. The results are as followed:Totally,52 proteins were identified with MS identification from two-dimensional blue native-SDS-PAGE and isoelectric focusing-SDS-PAGE separation. Comparative study indicated that the LHC-IIb in the mutant was about 40% less than that in wild type. However, both LHC-Ia and LHC-Ib decreased more greatly in this mutant, only 20% remaining. PSI subunits are nearly the same as wild type. PSII subunits Dl, D2, Cyt b559, and OEC 33KD protein are increased in the mutant. Moreover, the mutant showed increased amount of ATP synthase in chloroplast. IEF/SDS-PAGE shows that H+-transporting ATP synthase, lipocalin, fructose-bisphosphate aldolase, prenyltransferase and PSII assembly/stability factor HCF136 increased in the mutant, while PsaE and 8.7 kDa Fe-S protein decreased in the mutant compare to the wild type. Taken together, the results indicated that in this novel low chlorophyll b rice mutant, moderately decreased LHCII has no effects on PSII stability, and increased stiochiometries of PSII:PSI compensates for the reduction of LHCII. At the same time, higher level of ATP synthase might facilitate the turnover of Dl protein, which increased PSII stability under high light stress. In our results, the mutant showed increased HCF136 compared with the wild type, which helps to form a tighter structure of PSII core monomer. High expression of lipocalin and prenyltransferase increased antioxidation ability of the mutant. More phosphorylated LHCII can balance the light distribution between two photosystem under light conditions and increase PSI light absorbtion.We also studied the thylakoid proteome change under high temperature. Under 40℃heat stress, the supercomplexes dissolved very quickly, the PSII are the most sensitive protein complex to the heat stress, LHCII and OEC subunits dissolved first.In contrast, PSI are more stabilized. High expression of PsaE subunit increase cyclic electron flow around PSI which can help photosynthesis tolerate and recover from heat stress. ATP synthase increased under heat stress, which produce more ATP and decrease transthylakoid proton motive force help the plant to tolerate high temperature. Ferredoxin--NADP reductase and fructose-bisphosphate aldolase expression increased, which may also participate in heat stress response. In contrast, the mutant are more sensitive to heat stress. The supercomplexes dissolved more quickly and PSI subunits are less stable in the mutant than the wild type. PsaE expression decreased under 50℃treatment, which means the mutant lacks the cyclic electron flow around PSI to protect the photosystem under extremely high temperature.