plants under climatic stress - plant physiology

Plant Physiol. (1971) 47, 713-718

Plants under Climatic StressI. LOW TEMPERATURE, HIGH LIGHT EFFECTS ON PHOTOSYNTHESIS

Received for publication September 24, 1970

A. 0. TAYLOR AD J. A. RowLEYPlant Physiology Division, Department ofScientifc and Industrial Research, Palmerston North, New Zealand

ABSTRACr

Photosynthetic rates of both C4- and Crpathway plants grownat 25 C were measured before and during a period of chillingstress at 10 C, and then again at 25 C following various periodsat 10 C. When temperatures are first lowered photosyntheticrates drop immediately, then undergo a further reductionwhich is quite rapid in species such as Sorghum, maize, andPennisetum; slower in soybean; and very slow in Paspalum andryegrass. Visible light causes progressive permanent damage tothe photosynthetic capacity of leaves during this period oflowered photosynthesis. The extent of damage increases withlight intensity and the length of time leaves are held at 10 Cbut varies greatly between species, being roughly correlatedwith the extent to which chilling initially and subsequentlylowers photosynthesis. Three days of chilling (10 C) at 170w'm'2 reduces the photosynthetic capacity of youngest-maturePaspalum leaves only 30 to 40% while Sorghum leaves are es-sentially inoperative when returned to 25 C after the samestress. Root temperature has a substantial rapid effect on photo-synthesis of soybean and little immediate effect on Sorghum.Photosynthesis of stress-intolerant species (Sorghum) is re-duced only slightly more than that of semitolerant species(Paspalum) when temperatures are lowered at mid-photo-period, but to a far greater extent if temperatures are reducedat the commencement of a photoperiod.

Agronomists (6, 26) have emphasized that maize varietiessuitable for cool temperate climates not only should survivelow temperatures but should be able to grow at low tempera-tures during the early part of the growing season. It is welldocumented (7, 10) that photosynthesis of maize and othertropical grasses is very low at temperatures below 15 C andthat chilling temperatures will cause visible lesions on theleaves of these plants (22); yet we are still unsure of the pri-mary sites of temperature sensitivity.

Papers in this series will attempt to (a) define the primarysites of low temperature inhibition of photosynthesis in theC4-pathway (9) tropical grasses; (b) define factors causingpermanent damage to the photosynthetic system when theseplants are held at chilling temperature; (c) compare the chillingsensitivities and mechanisms of chilling damage in C,- and C,pathway species; and, ultimately, (d) assess the variation inability of tropical grass species and varieties to withstand chill-ing conditions.

This first paper investigates the effects of light and time onphotosynthetic rates and photosynthetic pigments, during andafter chilling stress.

MATERIAIS AND METHODSPlant Species. C-pathway type: Sorghum, hybrid NK 145;

maize, hybrid Wisc. 575; Paspalum dilatatum Poir.; Pen-nisetum typhoides (Burm.) S. & H. 23DA. C.-pathway type:ryegrass, Lolium multiflorum (L) var. Grasslands TamaWesterwolds; soybean, Glycine max (L) Merr. cv. Merit.Growing Conditions. Seed of all varieties was germinated in

a glasshouse, and seedlings were subsequently repotted into 4: 1pumice-peat. Young plants were transferred to controlled en-vironment cabinets 10 to 14 days prior to use and were fedadequate Hoagland's nutrient solution. Light in the cabinets(170 w*m', 400-700 nm) was supplied by a mixture of mer-cury vapor lamps (Philips HPLR, 2800 w total), tungsten fila-ment lamps (Mazda reflector floods, 450 w total), and bluefluorescent tubes (Philips type 126221, 450 w total) passingthrough a 2-cm deep flowing water screen. Photoperiod was 12hr with ambient CO. and uncontrolled humidity.

Photosynthetic Rates. Photosynthetic rates of single attachedleaves were measured by 1'CO2 depletion from a closed system.Leaves were clamped in a half-liter Perspex chamber betweensoft rubber seals, and the internal CO(2 concentration was raisedby 6 pt/liter with the addition of 1"CO.2 (50 mc/mmole). Photo-synthetic rates at 170 w-m-2 (400-700 nm) were determinedfrom the slope of a tangent to the 1"CO2 depletion curve at 300p4/liter CO, and expressed on a dry weight basis. Leaf tem-perature measured by abaxially attached thermocouples wasregulated to ±0.5 C. The rest of the plant was also maintainedat the desired temperature (± 1 C) by placing the Perspex cham-ber and plant in a controlled environment chamber.Whole plant photosynthetic rates were measured in both

open and closed type IR gas analysis systems and based onleaf dry weight corrected for each day from measured totalC(0 fluxes into the leaves. Shoots and roots were sealed intoseparate compartments of the plant chamber using siliconrubber sealant. Leaf temperature measured as above was main-tained at +0.5 C of the desired temperature and took 26 min toadjust from 25 C to 10 C. Root temperature took approxi-mately 4.5 hr to reach 10 to 11 C. Main illumination wasprovided by Sylvania Metalarc or Philips HPLR mercuryvapor lamps with small tungsten filament supplementaries.Light was passed through a 2-cm flowing water screen belowthe lamps to remove most infrared radiation and through neu-tral density screens to achieve the required intensity. In theopen system, humidity was uncontrolled and CO, was close toambient levels. Humidity was controlled in the closed systemwith CO(2 maintained at 300 pl/liter.

Relative Leaf Water Content. Relative water contents ofleaves were determined before and during stress according toBarrs and Weatherley (3). Groups of discs (1.3 cm) fromyoungest-mature leaves were floated on water (15 C) for 1.5 hrunder laboratory lighting to regain turgor fully.

ChIoroplast Pigments. Chlorophylls extracted in methanol713

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TAYLOR AND ROWLEY

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FIG. 1. Photosynthesis of youngest-mature leaves of some tropi-cal grasses at 25 C and then during 3 days of chilling stress at 10C. Note differences in scale. Rates were measured by incorporationof "4CO2 at 170 w.m2. Leaves of comparable age held in compara-ble orientation to the light source undergo very similar changes inphotosynthesis.

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FIG. 2. Photosynthesis recovery of youngest-mature leaves ofsome tropical grasses after their return to 25 C following 1.5 or2.5 days of chilling stress at 10 C. Prestress rates are shown on left-hand side for comparison. All photosynthetic rates were measuredby "4CO2 incorporation at 25 C and 170 wm-'.

were measured according to Holden (11). Carotenoids wereestimated spectrophotometrically from Goodwin's data (11)after separation by thin layer chromatography on MN 300cellulose in methanol-dichloromethane-water (100: 18:20, v/v)(21).

RESULTS

Visual Observations. Transfer of the C4-pathway tropicalgrasses from 25 C to 10 C while held under moderate light

intensities may ultimately cause necrosis of all leaf areas fullyexposed to the light. The upper or lower surface of the leafmay be affected first depending on leaf orientation. Visiblelesions take much longer to develop if areas of the leaf are bentaway significantly from the horizontal (400 or more) or areshaded by other leaves. One of the most sensitive species was

Sorghum, the leaves of which showed pronounced lesions after2 days' stress. Even if plants were returned to 25 C followingthis short period at 10 C, areas which appeared bleached coulddouble in size over a subsequent 2- to 3-hr period. Red pig-mentation also developed in the leaves around these visiblydamaged areas. This pigmentation could not be solubilized ina range of polar or nonpolar solvents and was presumed to beanthocyanins oxidatively polymerized in damaged cells.

Least affected of these tropical grasses under chilling con-ditions were leaves of Paspalum which showed only traces ofwhiteness after 3 to 4 days at 10 C. If temperatures were raisedto 25 C after this period of stress, Paspalum leaves did not be-come necrotic.On the basis of rate and degree of visible damage, the C,-

pathway plants could be placed in the tentative tolerancesequence: Paspalum > maize > Sorghum = Pennisetum.

Lowering the night temperature only for up to 8 days didnot cause visible stress lesions on the leaves of Sorghum, al-though growth appeared to be reduced. Lowering the daytemperature only caused slightly more rapid onset of stresslesions than if both day and night temperatures had been re-duced to 10 C.

Photosynthesis at Low Temperature. Changing patterns ofphotosynthesis in youngest-mature leaves of three C4-pathwayspecies following a change in temperature from 25 C to 10 Care shown in Figure 1. Photosynthesis fell immediately whenthe temperature was lowered, then showed some recoveryfollowed by a slow decline in rate in Paspalum or slight re-

covery followed by a rapid decline in rate in the less tolerantSorghum and maize.

The photosynthetic response of whole plants measured withan open IR gas analysis system (Table I) showed a trend simi-lar to that seen in the single leaf. Young Pennisetum andSorghum plants were essentially inoperative photosyntheticallyafter 3 days of chilling stress and showed negligible recoveryon return to 25 C. Photosynthesis in maize was markedlyaffected also, but the plants did show some recovery when thetemperature was raised after 3 days of chilling. With Paspalum,the immediate drop in photosynthesis which occurs when thetemperature is lowered was not proportionately as great as inthe other tropical grasses. Plants also retained this rate reason-

ably well and recovered better than any of the other tropicalgrasses when the temperature was raised. Damage is time-dependent, however, and even Paspalum is more severely dam-aged during longer periods of stress.The two C3-pathway species also showed very different re-

sponses to this chilling stress. Photosynthesis of Tama ryegrassplants was reduced less than 35% when the temperature was

lowered initially to 10 C, though plants did not rapidly re-

cover their prestress rates on return to 25 C after 3 days ofchilling. Photosynthesis of soybean held at low temperaturecontinued to fall quite rapidly over the 3-day period andshowed much lower recovery on returning the plants to 25 C.This response of soybean superficially resembles that of thestress-sensitive tropical grasses.

Recovery of Photosynthetic Rates. Data in Table I showingthe extent to which these plants recovered their photosyntheticcapacity on return to 25 C after 3 days of chilling stress havebeen noted above. These assessments were complicated, how-ever, by mutual leaf shading and varying leaf ages (see latersections). The extent of photosynthetic recovery in some tropi-

714 Plant Physiol. Vol. 47, 1971



PLANTS UNDER CLIMATIC STRESS. I

cal grasses is shown unambiguously in Figure 2 where singleleaves of uniform age were used. If Sorghum leaves are re-turned to 25 C after 1.5 days at 10 C (170 w m'), their photo-synthetic rate reaches only half its original level during the re-maining 6 hr of that photoperiod and during the next photo-period shows a marked decrease, apparently through somelatent stress sensitization. Sorghum leaves after 2.5 days ofstress were virtually inactive on return to 25 C and appearedfaintly white on the upper surface. After the same period ofchilling, maize leaves regained approximately one-third of theiroriginal rate, while Paspalum leaves recovered to more thantwo-thirds their original rate.An examination was also made of the effect of leaf age on

the extent of photosynthetic recovery. In all three tropicalgrass species it was found that second-mature leaves re-covered to roughly twice the extent of half-expanded leaves.Time of Day and Extent of Photosynthetic Drop. In previ-

ous work, temperatures were lowered from 25 C to 10 C atthe start of a photoperiod. Results shown in Table II demon-strate that photosynthesis of tropical grasses is not reduced tothe same extent when they are transferred from 25 C to 10 Ctoward the middle of a photoperiod as when the temperaturereduction occurs at the start of a photoperiod. Photosyntheticrates were very low immediately after a temperature drop at thestart of a photoperiod but recovered slowly to a maximum after2 to 3 hr in the light. Percentages shown in Table II were calcu-lated, therefore, from photosynthetic rates measured 3 hr afterthe temperature reduction.

Influence of Light Intensity. The marked dependence ofleaf damage on light intensity was seen clearly in cabinet ex-penments where light came predominantly from one directionand leaf movement was minimal. Areas of Sorghum leavesbent away approximately 40° from the horizontal appeared un-

Table I. CO2 Uptake of Plants prior to, during, and Immediatelyafter Chilling Stress

Plants conditioned at 25 C were given a 3-day chilling period at10 C and then returned to 25 C for 1 day. Temperature changeswere made at the start of a photoperiod, with light intensity main-tained at 170 w * m7' (400-700 nm). Photosynthetic rates were meas-ured in an open IR gas analysis system and are presented as meandaily rates in mg CO2/g leaf dry wt.-hr. Figures in parenthesesare absolute rates during and after stress expressed as a percentageof prestress rates. Data are from individual experiments but aretypical of those obtained using plants of 0.4 to 0.8 g dry weightof leaf.

CO Uptake

Pre- During chilling stress at 10 C Afterstress stressat 25 C 1st day 2nd day 3rd day at 25 C

mg CO2/g leaf dry wt-hr

Pennisetum ty- 41.2 4.4 1.4 <0.5 <0.5phoides (10.6%) (3.5%)

Sorghium NK 145 48.2 5.5 2.9 1.2 1.5(11.4%) (6.0%o) (2.5%) (3.2%)

Maize Wisc. 575 63.5 6.9 3.4 0.6 14.2(10.8%) (5.3%) (0.9%) (22.4%)

Paspalum dilatatum 33.6 11.1 8.9 7.3 15.6(33.1%) (26.5%) (21.7%) (46.3%)

Soybean "Merit" 23.2 5.2 3.1 1.6 6.4(22.5%) (13.5%) (6.8%) (27.8%)

Ryegrass "Tama" 18.6 12.4 12.7 10.0 9.5(66.5%) (68.8%) (53.5%) (51.0%)

Table II. Time of Temperature Drop and Extent ofPhotosynthetic Reduction

Photosynthetic rates of some C4-pathway tropical grasses weredetermined before and after a temperature drop from 25 to 10 C.Measurements were made at 170 w m-2 and expressed as in TableI. Temperatures were lowered at the start or the middle of a photo-period, and rates were determined 3 hr after the temperaturedrop. Relative humidity was maintained at 70%0. Data are meansof 5 to 10 determinations on plants of 0.4 to 0.8 g dry weight leaf.

C02 Uptake

Rate at 10 CBefore

temperaturedrop at 25 C Start of Mid-

photoperiod photoperiod

mg CO2/g leaf dry wt-krSorghum NK 145 48.2 8.8 ± 0.6 20.1 i 0.8

(18.2%) (41.7%)Paspalum dilatatum 33.6 14.6 i 1.2 17.2 i 0.8

(43.3%) (51.3%)Maize Wisc. 575 56.0 12.8 i 2.1 21.3 2.5

(22.9%) (38.0%o)

Table III. Light Intensity Effects on CO2 Uptake of SorghumNK 145 at Chilling Temperatures

All plants were conditioned at 25 C and 170 w_m72. Photosyn-thetic rates prior to, during, and after chilling were measured atthree light intensities and expressed as in Table I.

CO2 Uptake

LightIntensity During chilling stress at 10 C After(400-700 nm) Prestress stress

a lst day 2nd day 3rd day at25C

W-m-2 mg CO2/g Icaf dry wt - .r

215 50.1 3.0 0.4 <0.4 <0.4(5.9%) (0.7%)

170 48.2 5.5 2.9 1.2 1.5(11.4%) (6.0%o) (2.5%) (3.2%)

50 22.4 3.0 1.2 0.7 9.2(13.3%) (5.4%) (3.2%) (41.0%0)

damaged after 3 days of stress, while more horizontal regionsdeveloped marked visible lesions.

Table III demonstrates this effect more quantitatively. At215 w*m- Sorghum plants were essentially dead after 2 daysat 10 C; at 170 w m' damage was very pronounced after 3days at 10 C and recovery was almost negligible at 25 C; whileat 50 w *m2 damage was only slight and the plant was virtuallyback to normal after 1 day at 25 C. The total energies ofvisible light obtained at intensities of 215, 170, and 50 w*m'over 12-hr photoperiods are roughly equivalent to those ona sunny day, light cloudy day, and dull overcast day, re-spectively.Root Temperature and Leaf Temperature. The light porous

potting mixture used in this work allows root temperaturesto reach 10 to 11 C within 4.5 hr of the drop in chamber tem-perature, and so both leaf and root temperatures were essen-tially at 10 C for the total stress period. Lowering root tem-perature only or leaf temperature only had proportionatelydifferent effects on photosynthesis of Sorghum and soybean(Table IV).

Keeping the temperature of Sorghum roots at 25 C allowed

Plant Physiol. Vol. 47, 1971 715



TAYLOR AND ROWLEY

Table IV. Lowered Root or Lowered Leaf Temperature Effects on CO2 Uptake of Sorghum and SoybeanPlants were conditioned and photosynthetic rates were measured and expressed as in Table I. Root and shoot temperatures were

controlled at the level of the potting medium surface. Stem and leaf meristems of the young Sorghum plants would have been at thetemperature of the roots.

Temperature of C02 Uptake

During chilling stress at 10 CRoot Shoot ~~~~~~Prestress ____________________________ After stressRoot Shoot at 25 C iat 25 C

1st day 2nd day 3rd day

C mg CO/g leaf dry wi -hr

SorghumNK 145 10 10 48.2 5.5 2.9 1.2 1.5(11.4%) (6. 0%) (2.5%) (3.2%)

Sorghum NK 145 25 10 49.8 5.0 1.0 <0.5 <0.5(10.11%) (1 .8c%o)

Sorghum NK 145 10 25 46.7 34.5 17.8 12.1 11.4(73.8% ) (38.1%) (26.1% ) (24.5%)

Soybean Merit 10 10 23.2 5.2 3.1 1.6 6.4(22.5%) (13 .5%) (6.8%) (27.8%)

Soybean Merit 25 10 24.2 8.4 7.7 6.1 12.2(34.9%) (31.7%) (25.0%)O (50.6%)

Table V. Effect of Chtilling Stress on Leaf Relative Water ConltentTechnique for measuring relative water content is described in

the text. Discs were punched from youngest-mature leaves of arange of species before and during chilling stress, 10 C at 170w-mn2. Humidity was not controlled in the environment cabinetsand varied between 60 and 70%0 relative humidity at 25 C and 75and 82% relative humidity at 10 C.

Relative Water Content of Leaves

During chilling stress at 10 CPrestressat 25 C

Ist day 2nd day 3rd day

Sorghum NK 145 94.2 4 0.7 93.6 + 0.9 94.7 ± 1.3 92.7 i 1.1Maize Wisc. 575 94.6 i 0.4 94.6 ak 0.6 97.2 4 0.7 95.5 ± 0.4Paspalum dilatatum 94.0 0.6 94.4 4 0.5 96.7 ±t 0.3 94.4 4- 0.7Soybean "Merit" 90.1 ± 0.4 87.4 ± 0.6 87.4 i 0.8 92.3 ± 0.4Ryegrass "Tama" 92.1 4 0.4 91.2 1.1 92.8 ± 0.5 91.5 ± 0.8

Table VI. Acclimationt of Sorghum NK 145 and Subsequenit Effectsof Chilling Stress oni C02 Uptake

Plants were acclimated at 17 C for 3 or 8 days at 50 or 170 w m-2.Photosynthetic rates were then measured and expressed as inTable I. Chlorophyll was measured in youngest-mature leaves atleast partially developed under the acclimation conditions andexpressed as a percentage of that in leaves developed at 25 C.

C02 Uptake

Acclimation Chloro- Post- During chilling stress at 10 Cphyll accli- _________________ Aftermation stress

ate25C 1st day 2nd day 3rd day at 25 C

% mg C02/g leaf dry wt-ilr

14 days 25 C 100 48.2 5.5 2.9 1.2 1.5(170 wm72) (11.4%O) (6.0%) (2.5%) (3.2%)3 days 17 C 50 45.6 5.3 3.2 2.1 6.6(170 wm-2) (11.7%) (7.1%) (4.7%) (14.5%)8 days 17 C 23 23.8 4.7 2.0 1.4 4.9(170 wm-2) (19.6%) (8.3%) (5.7%) (20.6)8 days 17 C 70 32.5 4.4 1.2 0.9 2.1(50 wm-2) (13.4%7,) (3.7%) (2.8) (6.6%)

leaf photosynthesis at 10 C to drop at least as rapidly as whenroots also were lowered to 10 C. Lowering the root tempera-ture only reduced photosynthesis just 26% during the 1st dayof stress, though photosynthesis does fall further during pro-longed root chilling.

Lowering only leaf temperature of soybean has a reasonablyrapid and pronounced effect on leaf photosynthesis, althoughnot as great as the combined effects of lowered root and leaftemperature. These observations could mean that lowered roottemperatures cause water stress in the leaves of soybean morerapidly than in Sorghum.The relative water content of soybean leaves (Table V) was

low even before chilling and fell further during initial stages ofstress, while that of the tropical grasses was significantly higherthan that of soybean before stress and generally rose slightlywhen the temperature was lowered.

Acclimation to Stress. Attempts were made to acclimateSorghum to tolerate low temperature, high light stress. Plantswere placed at 17 C for varying lengths of time under differentlight intensities. Some typical results are shown in Table VI.Photosynthetic rates on return to 25 C for 1 day were markedlydifferent following the various acclimation treatments.Lowered chlorophyll content of the leaves and longer periodsat 17 C both correlated with reduction in these rates. Loweredchlorophyll content in the leaves also appeared to protectplants somewhat from more pronounced stress as evidencedby their higher photosynthetic recovery.

Although these general patterns seem reasonably concise,many parts of the plant probably respond differently from oneanother. Tissue formed prior to acclimation retained itschlorophyll during acclimation, while that formed (18) duringacclimation had increasingly lower levels of chlorophyll. Thiscaused marked gradients in pigmentation to arise along de-veloping leaves. Small areas of individual leaves may need tobe used to get more precise data on the effects of acclimation,but interactions with the whole plant may still be difficult tounravel.

Chloroplast Pigments during Stress. Effects of low tempera-ture, high light stress on the level of chloroplastic pigmentswere investigated in youngest-mature leaves of several tropicalgrasses. Pigment changes seen in Sorghum leaves (Fig. 3) weresimilar, though more marked, than those seen in other species.Chlorophyll levels were essentially unaltered by 2.5 days ofstress. The main change was an increase in lutein matched by

716 Plant Physiol. Vol. 47, 1971



PLANTS UNDER'CLIMATIC STRESS. I

a fall in the xanthophyll epoxides (violaxanthin and neoxan-thin) and fl-carotene. Levels rapidly returned to normal on in-creasing the temperature if only limited permanent photo-synthetic damage had occurred. Since we were primarilyconcerned with events leading to inactivation of the photo-synthetic apparatus by light rather than the ultimate conse-quences of necrosis, regions showing distinct visible lesionswere discarded from samples taken for analysis. Once lesionsdevelop, electron microscopy has revealed (25) that muchof the upper palisade is necrotic, and the level of all chloro-plast pigments dropped rapidly in these dying cells.

DISCUSSION

When the temperature of the leaf, shoot, and root environsof a plant were lowered from 25 C to chilling levels (10 C),photosynthesis of the leaves dropped immediately. Species ofboth C4-pathway tropical grasses and C-pathway plants dif-fered widely in the extent of this reduction in photosyntheticrates. If the temperature was maintained at 10 C (Table I),photosynthesis of some species (Paspalum, ryegrass) con-tinued to fall only slowly while photosynthesis of others(Pennisetum, maize, Sorghum) rapidly reduced to negligiblerates after 2 to 3 days. The rate of this second slower decline inphotosynthetic rates was generally correlated with the extentof the immediate reduction in photosynthesis which occurredwhen temperatures were initially lowered.

There were at least two causes of the initial temperature-induced reduction in photosynthesis: first, a direct effect on theleaf and, second, an indirect one acting through reduced wateruptake by the roots (2), which then lowers leaf water content.With soybean, these effects were roughly equal. Lowering bothroot and leaf temperatures doubled the decline in photo-synthetic rate caused by lowered leaf temperature alone, andleaf water content fell (Table V) to levels documented as re-ducing photosynthesis (23). Sorghum responded somewhatdifferently. Lowering only leaf temperatures had the sameeffect as lowering both leaf and root temperatures. Slower re-ductions in photosynthesis which develop when the roots arechilled may be caused by reduced cation uptake (27). A rapidtransient drop in the water content of maize leaves caused bylowering the roots only, to 5 C, has been reported using ,B-gauge techniques (16), but no similar effect was detected usingthe leaf disc assay when the temperature of whole plants (roots,shoots, and leaves) of maize or any other tropical grass was re-duced to IO C.When some plants are exposed to chilling temperatures,

light causes a time-dependent destruction of the photosyntheticapparatus (Table III). Solarization (12), a term coined for thedamaging effects of very high light intensities on starch pro-duction in the leaves of many plants; high light, low tempera-ture inhibitions of algal growth (24); and the photoinactivationof isolated chloroplasts (13, 14) may be phenomena related toeffects described in this paper. Reducing photosynthesis bylowering the temperature puts little increased heat-dissipatingload on the chloroplast, since 75 to 85% of visible light ab-sorbed by the chloroplastic pigments is lost as heat under evenoptimal photosynthetic conditions, yet this light under thesechilling conditions causes a rapid change in the ultrastructureof mature chloroplasts (25). The effect of these ultrastructuralchanges on the photosynthetic capacity of leaves seen on re-turning leaves to 25 C are profound (Fig. 2). The maximal re-covered rate after stress may not even stay constant, andSorghum especially shows a pronounced latent stress sensitiza-tion. Another complication when using whole plants is thatnone of the leaves is horizontal over its entire length so thatmarked gradients in light interception and hence photo-

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FIG. 3. Level of major chlorophylls, carotenoids, and xantho-phyils (ug cm-' leaf) in youngest-mature leaves of Sorghum NK 145at 25 C, then during 2.5 days of chilling stress (10 C at 170 w.m-'),and finally on return to 25 C. Areas of leaves with visible stresslesions were not used.

synthetic disruption occur along the leaves. This is especiallyso in environmental cabinet work where light comes largelyfrom one direction. These gradients, coupled with mutual leafshading and the greater sensitivity of growing leaves overmature leaves, probably mean that no one area of the leaf orleaves responds exactly the same. Nevertheless, the trends areobvious. Reducing the light intensity from 170 to 50 w mprotects Sorghum plants significantly from damage caused bychilling temperatures that otherwise would have resulted inalmost complete necrosis (Table III). Photosynthesis at 50w *m'2 still declines over the 3 days at 10 C, but this cannot bedue to permanent photosynthetic disruption. An apparentlyrelated effect of light intensity on chilling damage has beennoted by Amin (1), who chilled cotton plants under constantconditions (72 hr at 3 C; 200 ft-c) and subsequently found aneffect of light intensity on expression of the damage.

Changes in the level of chlorophylls a and b were not de-tected in leaves of Sorghum (Fig. 3) or other tropical grasses,until some permanent photosynthetic damage had occurred,although these assays would not detect possible changes in lowlevels of other forms of chlorophyll (5). Similar observationshave been made using Chlorella vulgaris (19) held underhigh light intensities, and in isolated chloroplasts (13, 14) visi-ble light causes the loss of all chloroplast activities at 50 timesthe rate of chlorophyll and carotenoid bleaching. Under chill-ing stress, reduced levels of the xanthophyll epoxides andviolaxanthin and increased levels of the reduced xanthophylllutein develop in Sorghum leaves. This increase in the level ofreduced xanthophylls does not support the concept of a photo-oxidative destruction of the photosynthetic apparatus (15, 19)with xanthophyll oxidations having a protective function (17).Hager has reported (8) that a decrease in the oxidation level ofthe xanthophylls can be caused by a decrease in pH of thethylakoid intraspaces.

In this work (Table II) it has been demonstrated that photo-synthesis of tropical grasses is lowered less if temperaturesare reduced in mid-photoperiod rather than at the start of a

photoperiod. Photosynthetic rates at 10 C measured 2 hr aftera temperature drop in mid-photoperiod were very similar atbetween 16 and 21 mg of CO2 per g of leaf dry weight per hr

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717Plant -Physiol. Vol. 47, 1971



TAYLOR AND ROWLEY

for the tropical grasses and the C3-pathway ryegrass. Thesedata are more consistent with the concepts of Bjorkman et al.(4) of some mutual site of photosynthetic temperature sensitiv-ity in both C4- and C3-pathway species. Greater reductionsin photosynthesis seen in stress-sensitive tropical grasses suchas Sorghum when temperatures are lowered at the start of a

photoperiod are suggestive of a light-temperature-dependentinduction or activation of some enzyme or enhanced low tem-perature sensitivity of stomatal opening mechanisms (20).

Interesting features demonstrated in this work have been(a) the rapidity with which permanent photosynthetic damageoccurs during simulated short periods of chilling temperatures;(b) the crucial involvement of light within intensity ranges

commonly experienced in the field; (c) marked differences intolerance to chilling stress shown by some species of both C,-and C3-pathway species; and (d) basically similar patterns ofphotosynthetic disruption in chilling-sensitive species of bothC4- and C.-pathway plants. These findings also demonstratethe care required in the experimental control of temperature,light, and time in selection programs for chilling tolerance.

Acknowledgments-Thanks are due to N. M. Jepsen for pigment assays andC. G. Tunnicliffe for constructing and maintaining much of the equipment. Penni-setum typhoides (Burm.) S. & H. 23 DA seeds were supplied by Coastal PlainsExperiment Station, Georgia, U.S.A.

LITERATURE CITED

1. AMiN, J. V. 1969. Growth and development of cold-injured cotton plants. PlantSoil 31: 365-373.

2. ARNDT, C. H. 1937. Water absorption in the cotton plant as affected by soiland water temperatures. Plant Physiol. 12: 703-720.

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plants under climatic stress - plant physiology

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