Received 8 Sept. 2003 Accepted 7 Jan. 2004Supported by the National Natural Science Foundation of China (30200021).

* Author for correspondence. E-mail: <bsqiu@public.wh.hb.cn>.

http://www.chineseplantscience.com

Acta Botanica Sinica植 物 学 报 2004, 46 (5): 552-559

Diurnal Photosynthesis and Photoinhibition of Rice Leaves with Chlorophyll Fluorescence

LIU Ji-Yong1, QIU Bao-Sheng1, 2* , LIU Zhi-Li2, YANG Wan-Nian1

(1. College of Life Sciences, Central China Normal University, Wuhan 430079, China;2. College of Life Sciences, Nanjing University, Nanjing 210093, China)

Abstract : The diurnal photosynthesis and photoinhibition in the photoper iod-sensit ive genicmale-sterile rice (Oryza sativa L.), Nongken 58S (NK58S), were investigated. From 06:00 to 09:00, noremarkable photoinhibition occurred, and the down-regulation of photosynthesis might be due to therunning of xanthophylls cycle. From 10:00 to 12:00, the specific energy flux for dissipation (DIo/RC) andthe net rate of reaction centers (RCs) closure (dV/dto) were increased, while the probability of electrontransport at the acceptor side (ψo) and the density of active RCs (Do) were decreased. These indicatedthat the photoinhibition of NK58S was exacerbated with the inactivation of PSⅡ RCs. Fluorescence darkrelaxation analysis and inhibitor treatment suggested that all of state transition, xanthophyll cycle andinactivation of PSⅡ RCs could contribute to protect NK58S against photodamage. Compared with theinactivation of PSⅡRCs, xanthophyll cycle had an immediate response to high light stress, wh ichfunctioned mainly in the period of relatively low light intensity. However, the inactivation of PSⅡ RCsplayed an important role in protecting the remaining active RCs when xanthophyll cycle was saturated.Key words: photoperiod-sensitive genic male-sterile rice; Nongken 58S; JIP test; photoinhibition;

reaction center inactivation; D1 protein turnover; xanthophylls cycle

Higher plants are usually encountered with light stressin their habitats. Strong irradiance will inevitably inducephotoinhibit ion with the decrease of photochemical effi-ciency when the absorbed energy is much higher than thatutilized by the photosynthetic organs (Long et al., 1994).The photoinhibition will occur much frequently when strongirradiance is combined with high temperature, drought oro ther s t ress es in s ummer midday . Furthermore, thephotoinhibition could result in photooxidative damage, pig-ment bleaching and even irreversible damage to the photo-syn thetic apparatus. Numerous p rotective mechanismshave been developed to minimize the sunlight absorptionin long-term evolu tionary history, such as alterat ion inleaves ang le, chloroplast movement and reflective cuticlecovering. Alternatively, state transition, xanthophylls cycleand PSⅡ repair cycle contribute internally to avoiding thedamage to the photosynthetic apparatus (Long et al., 1994;Horton et al., 1996). However, various species might differin their protective mechanisms for photoinhibition.

Photoinhibition is characterized by the decline of pho-tos ynthet ic quan tum efficiency and pho tochemicalefficiency, and Fv/Fm value is widely used as an index toevaluate the extent o f photoinhibit ion (Krause and Weis,1991; Govindjee, 1995; Maxwell and Johnson, 2000).Nevertheles s, many processes includ ing PSⅡ reaction

center repair cycle and thermal dissipation associated withxan thophylls cycle could lead to the decrease o f Fv/Fm(Long et a l ., 1994). In add it ion , the locat ion o fpho to inh ib it ion varies with the s pecific inh ib itorymechanisms, donor side or acceptor side of PSⅡ reactioncenter (Long et al., 1994; Andersson and Aro, 2001). Thesecould not be distinguished just by evaluating the changesof Fv/Fm. Then, a thorough investigation on the primaryphotochemical reactions as well as the whole electron trans-port chain by the fluorescence techniques is required. Thisis the story of “JIP test”.

Fast fluorescence rise of dark-adapted leaves recordedwith high time-resolution fluorimeters such as PEA showsa polyphasic O-J-I-P transient. The polyphasic O-J-I-P risereflects the kinetic process of PSⅡ closure and has alsobeen found to be very sensit ive to s tres ses caused bychanges in different environmental conditions, such as lightintensity, temperature, drought, atmospheric CO2 or ozoneelevation and chemical in fluences (Strasser and Tsimilli-Michael, 2001). But it is hard to evaluate the PSⅡ pho to-chemical behav iors just from its shape change. The JIP-tes t has been in troduced and further developed for thequantitative analysis of O-J-I-P transient (St rass er andStrasser, 1995). It translates the shape changes of O-J-I-Ptransient to quantitative changes of various parameters and

Acta Botanica Sinica 植物学报 Vol.46 No.5 2004558

down-regulation o f photosynthetic activity in NK58S.Nevertheless, each mechanism plays different roles to dis-sipate excess excitation energy and protects NK58S againstphotodamage. It has been suggested that state transitionis much important only in low light (Krause and Weis, 1991).Thus, we have not paid much attention to it in this paper.The xanthophyll cycle was initiated immediately in the firsthour when NK58S was exposed to strong irradiance in-doors (Fig.6). Thereafter, certain amounts of PSⅡ RC wereinactivated. The inactivation of PSⅡ RC is inevitable whenthe fully operation of xanthophyll cycle could not dissipateexcess excitation energy effectively. This is consistent withthe suggestion that xanthophyll cycle is the firs t line ofdefense against PSⅡ over-excitat ion (Long et a l., 1994).The JIP test has also shown that the photoinhibition ofNK58S is not marked from 06:00 to 09:00 and the down-regulation of photosynthetic activity is mainly related withthe xanthophyll cycle (Fig .4). The inactivation of PS Ⅱ

RC occurs when ligh t intensity is increased to 1 200µmol·m-2·s-1 at 10:00, and both Do and ψo values started todecrease (Fig.4). This is also consistent with Quick andStitt (1989). They had suggested that the fast componentof fluorescence quenching was not well suited to dissipatelight energy at super-saturating intensities.

Our resu lts sugges t that the damage-repair cycle ofPSⅡ RCs could contribute to protecting NK58S againstphotodamage. The inactivation of PSⅡ RC has ever beenregarded simply as damage to plants. Its protective effectsagainst photodamage have been known gradually. Theaccumulated inactivated RCs could act as potent quenchers,convert absorbed excitation energy to heat, and protectthe remaining centers against further damage (Long et al.,1994). However, the exact mechanism for inactivated RCsto dissipate excess excitation energy is still uncertain. Therepair of photodamaged PSⅡ requires a number of p ro-cesses including the dephosphorylation of D1 protein, itsdegradation, de novo synthesis, and reassembly into func-tional PSⅡ complexes (Zhang et al., 2000; Andersson andAro, 2001). The damaged D1 protein is t riggered to bedegraded via a multistep proteolytic reaction requiring GTPand ATP and catalyzed by proteases Deg P2 and FtsH(Goldberg, 1992; Spetea et a l., 1999; Andersson and Aro,2001). It is uncertain whether the s peedup of ATP con-sumption in the damage-repair cycle of PSⅡ RC could con-tribute NK58S to dissipating excess excitation energy.

It has been s uggested that the net degradation of D1protein may res ult from the strong irradiance used indoorwhich the field grown p lants seldom encounter in theirhabitats. The indoor treatment of strong irradiance is 1 200

µmol·m-2·s -1 in this study and is far below the maximallight intensity in the field (2 000 µmol·m-2·s-1). This sug-gests that the inactivation of PSⅡ RCs in NK58S is not dueto the difference of light intensities indoor and outdoor.

In this study, we have investigated the photoinhibitionof field grown NK58S with JIP test, from which the conclu-sion is well in accordance with that of laboratory experi-ment with the utilization of inhibitor. The JIP-test has beenproved to be a very useful tool for the in vivo investigationof the adaptive behavior of the photosynthetic apparatusand, especially, of PSⅡ to a wide variety and combinationof stressors owing to its convenient, rapid and non-inva-sive features (Lu and Vonshak, 1999; Strasser and Tsimilli-Michael, 2001). However, the recorded fluorescence tran-sients carry much more information than those used by theJIP test. The more powerful methods of numerical simula-tions utilizing the rich in formation are expected. This isdependent on our profound understanding to the fluores-cence signal and the appearance of more powerful fluorim-eters in the future.

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(Managing editor: HE Ping)