Abstract
Increasing photosynthetic ability as a whole is essential for acquiring higher crop yields. Nonleaf green organs (NLGOs) make important contributions to photosynthate formation, especially under stress conditions. However, there is little information on the pod wall in legume forage related to seed development and yield. This experiment is designed for alfalfa (Medicago sativa) under drought stress to explore the photosynthetic responses of pod walls after 5, 10, 15, and 20 days of pollination (DAP5, DAP10, DAP15, and DAP20) based on ultrastructural, physiological and proteomic analyses. Stomata were evidently observed on the outer epidermis of the pod wall. Chloroplasts had intact structures arranged alongside the cell wall, which on DAP5 were already capable of producing photosynthate. The pod wall at the late stage (DAP20) still had photosynthetic ability under well-watered (WW) treatments, while under water-stress (WS), the structure of the chloroplast membrane was damaged and the grana lamella of thylakoids were blurry. The chlorophyll a and chlorophyll b concentrations both decreased with the development of pod walls, and drought stress impeded the synthesis of photosynthetic pigments. Although the activity of ribulose-1,5-bisphosphate carboxylase (RuBisCo) decreased in the pod wall under drought stress, the activity of phosphoenolpyruvate carboxylase (PEPC) increased higher than that of RuBisCo. The proteomic analysis showed that the absorption of light is limited due to the suppression of the synthesis of chlorophyll a/b binding proteins by drought stress. Moreover, proteins involved in photosystem I and photosystem II were downregulated under WW compared with WS. Although the expression of some proteins participating in the regeneration period of RuBisCo was suppressed in the pod wall subjected to drought stress, the synthesis of PEPC was induced. In addition, some proteins, which were involved in the reduction period of RuBisCo, carbohydrate metabolism, and energy metabolism, and related to resistance, including chitinase, heat shock protein 81-2 (Hsp81-2), and lipoxygenases (LOXs), were highly expressed for the protective response to drought stress. It could be suggested that the pod wall in alfalfa is capable of operating photosynthesis and reducing the photosynthetic loss from drought stress through the promotion of the C4 pathway, ATP synthesis, and resistance ability.
Highlights
Photosynthesis is considered as the most important chemical reaction and provides over 90% of dry matter for crop yield formation [1,2]
Stoma acted in respiration and transpiration and allowed CO2 to enter for operating photosynthesis as well
Previous research has shown that stoma was found on other nonleaf green organs (NLGOs), such as the exposed peduncles of wheat [22], the silique shell of oilseed rape [4], and the capsule wall of castor [5]
Summary
Photosynthesis is considered as the most important chemical reaction and provides over 90% of dry matter for crop yield formation [1,2]. Little is known on the physiological response and the photosynthetic contribution of the pod wall in alfalfa under drought stress. Physiological, ultrastructural, and proteomic analyses were carried out to (1) investigate the photosynthetic characteristics of the pod wall in alfalfa, and (2) research the response mechanism of photosynthesis in the pod wall to drought stress. Chloroplasts on DAP5 existed with the intact structure and were arranged close to the cell wall (Figure 2A). Lots of starch grains could still be observed on DAP15, while the evident changes occurred in the structure of chloroplasts, i.e., the membrane was partly broken, and the grana lamellae of thylakoids became blurry (Figure 2G). Changes of Chlorophyll Concentration in Pod Wall under Drought Stress Except for chloroplasts, the structure of other organelle or tissues changed under drought stress as weWlli.thThtheecednetvreallovpamcuenotleowf tahsebpiogdgewr ainll,ththeecceollnucenndterratWioWn o(fFcihgluorreop2Dhy),llwah, cilhelothroepghaypllbbe,twanedentotthael cchenlotrroapl hvayclludoelecraenadsetdheuncedlel rwbaoltlhbeWcoWmeanwdidWerS utrnedaetmr WenSts(,Faingdurteh2eHco).nTcehnetmraetimonbroafncehsltorruocptuhryelloaf wthaes mhiigtohcehrotnhdanriothnawt oafscihnltoarcotpahnydll cbl,eraersopnecDtivAePly5. uFnudrtehreWrmSo(rFe,igthuereco2nEc),enwthrailteioint owfacshblorrookpehnyallnad, cbhlulorrroepdhoynll Db,AaPn2d0to(Ftaigl uchrelo2rHop).hyll in the treatment of WS decreased significantly (p < 0.05) compared with WW (Table 1)
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