Planta 142, 187-193 (1978)

Planta 9 by Springer-Verlag t978

The Relationship between Phytoehrome Photoequilibrium and Development in Light Grown Chenopodium album L.* D.C. Morgan and H. Smith Department of Physiologyand Environmental Studies, University of Nottingham, School of Agriculture, Sutton Bonington, Loughborough LE12 5RD, U.K.

Abstract. Chenopodium album seedlings were grown

in light environments in which supplementary far-red light was mixed with white fluorescent light during various parts of the photoperiod. Both the logarithmic rate constant of stem extension and the leaf dry weight:stem dry weight ratio were linearly related to estimated phytochrome photoequilibrium (~b) in each treatment regime. These data are taken to be indicative of a functional link between phytochrome and development in the green plant. A layer of chlorophyllous tissue only affected the linearity between calculated q5 and the logarithmic stem extension rate at high chlorophyll concentrations, whilst even low concentrations - equivalent to the levels found in stem t i s s u e - c a u s e d a significant shift in measured qS. Endof-day supplementary far-red (FR) light induced between 0-35 per cent of the response elicited by all-day supplementary FR, whilst daytime supplementary FR (with a white fluorescent light end-of-day treatment) induced approximately 90 per cent. The ecological significance of this difference is discussed with respect to shade detection. Key words:

equilibrium

Far-red PhotoPhytochrome -- Stem extension.

Chenopodium

Introduction

Phytochrome, the red/far-red (R/FR) photoreversible plant photoreceptor, has been rigorously investigated in dark grown tissue, and its role in seedling de* Paper 7 in the series ~176 function of phytochrome in the natural environment" [for paper 6 see McLaren, J.S., Smith, H., Plant, Cell and Environment 1, 61 67, 1978]

etiolation well documented. Its function in green plants is less clear. The previous papers in this series have expounded the hypothesis that in nature phytochrome mediates the developmental responses to shade light quality (Holmes and Smith, 1977a, b, c; Smith and Holmes, 1977; Tasker and Smith, 1 9 7 7 ) - t h a t is the change in R / F R ratio (~) due to attenuation of daylight by the shading vegetation. Most striking is the increased internode extension (Morgan and Smith, 1976), which can be explained ecologically as an adaptation for avoiding shade. In their previous paper the authors briefly reported a linear relationship between phytochrome photoequilibrium (~b), estimated from etiolated tissue, and stem extension rate in green Chenopodium album. These experiments are fully reported here, together with two related investigations. Firstly, chlorophyll screening is known to shift phytochrome absorption maxima (Grill, 1972), and unless the photoreceptor is located in the upper epidermis this could well affect the response linearity. Therefore, the effect of a layer of chlorophylous tissue on calculated and measured ~b, and their relationship with stem extension, was investigated. Secondly, several workers have proposed that in nature the end-ofday spectrum determines the phytochrome-controlled response to shade (Kasperbauer, 1971; Shropshire, 1973; Vince-Prue, 1977), mainly because in the controlled environment plants readily react to a F R light treatment at the end of the daily white light period. Two points make this hypothesis less plausible than daytime detection, these being: (a) daytime detection offers more complete information; and (b) the non-vegetational change in ~ at dusk (Shropshire, 1973; Holmes and Smith, 1977a) would complicate end-of-day shade detection. Therefore, t h e relative responses of C. album to daytime and end-of-day supplementary F R were investigated.

0032-0935/78/0187/$01.40

188

D.C. Morgan and H. Smith: Phytochrome Photoequilibrium and Development in Light-grown Plants

Materials and Methods Light Measurements and Phytochrome Determinations Light quality was measured using a Gamma Scientific (San Diego) spectroradiometer, and light quantity, defined as the spectral photon flux of the 400-700 nm waveband (photosynthetically active radiation; McCree, 1972), was measured with a Lambda Instruments (Lincoln, Nebraska) quantum meter. Both instruments have been fully described before (Holmes and Smith, 1977a)_ Phytochrome photoequilibrium (q~) was determined by two indirect methods, both of which gave estimates relating to etiolated tissue. The first, estimated (~ (4~), is determined from ~ (the 660 nm:730 nm ratio of spectral photon fluence rate) by using Smith and Holmes' (1977) plot of the ~b/~relationship. The second, calculated q~ (4~o), is calculated from the spectral photon distribution of the 400-800 nm waveband using an expansion of Hartmann's (1966) formula, which will be described in detail by Tasker and Smith later on. Direct determination, which was used in the screening experiments, is described below.

Plant Material and Light Sources C. album seed was collected from Field 12, on the Experimental Farm, Sutton Bonington. Seedlings were grown up to the third leaf pair stage in 6.35 cm pots in John Innes potting compost No. 1, and then batches of between 10 and 15 seedlings, depending on uniformity, were transferred to each of the treatment environments. The pretreatment environment was lit by a bank of 1525 mm 80 W Phillips "Warm White" fluorescent tubes (4=9.0; P A R = 180 + 11 gmol m - 2 s- 1). The photo- and thermo-periods were 16 h: 20_+1 ~ C day and 8 h: 15_+1 ~ night throughout. The light sources of the two controlled environment treatment cabinets (floor area 0.6 m 2) could be adjusted to provide a ~"range of between 3.8 and 0.16 at a PAR of 100+4 gmol in -2 s -1. White light was provided by a bank (rtormalty six) of 600 mm 40W Osram-GEC "Colour Matching 55" fluorescent tubes (4=3.8). FR was provided in two ways: a small addition was achieved by adding two pearled 60 W tungsten filament Iamps to the fluorescent source; for larger additions the light from up to 40 clear 150 W water-cooled tungsten filament lamps was filtered through one layer of 3.15 mm thick green 600 and one layer of 3.15 mm thick red 400 Perspex (ICI, Welwyn Garden City). Two cm of water below the lamps acted as au efficient heat (infra-red) filter, so that the maximum temperature difference between the treatments was 2 ~ C. The highest temperature invariably occurred in the high-intermediate ( values (i.e. the White plus two unfiltered 60 W tungsten filament lamps).

Chlorophyll Extraction and Screening Techniques Chlorophyll was extracted by grinding the tissue with 80% acetone in a mortar and pestle. The volume was made up to 10 ml, and spun at 2000 rpm for 10 min in an MSE Mistral 6L. The absorption of the supernatant at 663 and 645 nm was measured on a Pye Unicam SP 1800 spectrophotometer. All operations were conducted at 0 ~ C. Chlorophyll concentration was estimated from the formulae of MaeKinney (1941). For shielding experiments two plastic cuvettes-the front loaded with a particular chlorophyll solution, the rear packed with 5 d old etiolated pea epicotyl tissue, were placed in an SP 1800 euvette holder, and irradiated for 1 rain. The cuvette holder was covered so that light could only enter via the 8 • 24 mm entrance

slit. The resulting ~b in the pea epicotyl tissue was measured using a Perkin-EImer model 156 dual-wavelength spectrophotometer. Between treatments the etiolated tissue was kept on ice, and the Perkin-Elmer irradiation chamber was water cooled. Two light sources were used to irradiate the shielded material. White light was provided by a single 150 W clear tungsten filament lamp, held in a reflective aluminium housing. FR was supplied by a 108 W platinum ribbon filament lamp focused through a FR filter of the type described above.

Results (i) Effect o f Added F R on S t e m Extension Rate B a t c h e s o f 15 p l a n t s w e r e g r o w n i n l o w a n d h i g h ~, the treatment being given over the whole of the photop e r i o d ( ' a l l d a y ' ; Fig. 1), f o r 21 d. T h e i m m e d i a t e and large developmental response to FR irradiance is r e a d i l y a p p a r e n t (Fig. 2). I t s h o u l d b e n o t e d t h a t no attempt was made to equalise the photosynthetically active radiation as t h e s u p p l e m e n t a r y - F R t r e a t e d p l a n t s g r e w taller. B y d a y 9 t h e d i f f e r e n c e b e t w e e n t h e t o p s o f t h e t w o sets o f p l a n t s in t e r m s of photosynthetically active radiation and temperature was 7 gmol m- 2 s- t and 0 ~ C respectively. These differences are small and considered unimportant. H o w e v e r , b y d a y 21 t h e d i f f e r e n t i a l s w e r e 49 g m o l m - 2 s - x a n d 2 ~ C, a n d t h e s e will h a v e b e e n r e s p o n s i b l e f o r s o m e o f t h e o b s e r v e d i n c r e a s e in h e i g h t .

(ii) E f f e c t o f a Range o f ~ on Extension Rate, and its Relationship to B a t c h e s o f b e t w e e n 10 a n d 15 s e e d l i n g s w e r e g r o w n i n a r a n g e o f l i g h t e n v i r o n m e n t s , d i f f e r i n g o n l y in their FR irradiance, for 9 d (the period over which s t e m e x t e n s i o n is e x p o n e n t i a l (Fig. 2) a n d d u r i n g which differences in photosynthetically active radiation and temperature at the plant apices are insignificant). The light treatment was given over the whole d a y ( ' a l l d a y ' ; Fig. 1). O v e r t h e 9 d p e r i o d t h e e x t e n s i o n r a t e w a s g r a d e d a c c o r d i n g t o ~, a n d s i n c e it w a s e x p o n e n t i a l it c o u l d b e a c c u r a t e l y q u a n t i f i e d as t h e l o g a r i t h m i c r a t e c o n s t a n t , w h i c h is r e f e r r e d t o as t h e l o g a r i t h m i c s t e m e x t e n s i o n r a t e (Fig. 3). A n a l ysis o f v a r i a n c e i n d i c a t e s a s i g n i f i c a n t d i f f e r e n c e ( P < 0.001) b e t w e e n t h e e x t e n s i o n r a t e s o f t h e d i f f e r e n t t r e a t m e n t s . T h e r e l a t i o n s h i p s b e t w e e n qS~ a n d t h e loga r i t h m i c s t e m e x t e n s i o n r a t e , a n d ~bo a n d t h e l e a f d r y w e i g h t : s t e m d r y w e i g h t r a t i o a r e l i n e a r (Fig. 4). That both of these extension related parameters are l i n e a r l y r e l a t e d t o ~be is t h o u g h t t o b e s t r o n g , a l t h o u g h circumstantial, evidence for a functional link between phytochrome and development in the green plant.

D.C. Morgan and H. Smith: Phytochrome Photoequilibrium and Development in Light-grown Plants TREATMENT

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Fig. 3. The relationship between loglo mean height and time for C. album seedlings grown under four different spectral photon distributions. ~ is shown in parenthesis. Each of the twelve relationships, including the four shown here, were significantly different from zero slope (t***) and linear (r 0.99). Note the slope of the relationship is termed the logarithmic stem extension rate

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