J.PlantPhysiol. Vol. 134.pp.113-114{1989} Short C011101Unication

Refixation of Photorespiratory CO2 and NH3 by the Leaf Slices of Parthenium hysterophorus L.

P. A. KUMAR and Y. P. ABROL

Nuclear Research Laboratory and Division of Plant Physiology, Indian Agricultural Research Institute, New Delhi 110012, India

Received May 1, 1988 . Accepted September 30,1988

Summary

The capacity to reassimilate carbon dioxide originating from 14C-Iabelled photorespiratory substrates has been investigated in Parthenium hysterophorus, a C3 -C4 intermediate species in comparison with wheat (C3) and sorghum (C4). Dark/light ratios of 14C02 evolution from either labelled glycolate or glycine by the leaf slices of P. hysterophorus were intermediate to those of wheat and sorghum, indicating that P. hysterophorus possesses an enhanced capacity for internal refixation of 14C02 produced during photorespiration. The rate of ammonium accumulation caused by methionine sulfoximine (MSO) in the leaves of P. hysterophorus was also intermediate to those observed with the leaves of C3 and C4 plants.

Key words: Parthenium hysterophorus; photorespiration; C3 - C4 intermediate.

Abbreviations: INH, isonicotinyl hydrazide; MSO, methionine sulfoximine.

Introduction

Parthenium hysterophorus L. a weed belonging to the family Asteraceae, has been identified as a species exhibiting low photorespiration (Rajendrudu and Das, 1981). Recently, Ku et aI. (1985) reported that P. hysterophorus possesses a dis­tinct Kranz leaf anatomy and low CO2 compensation point. However, the activities of several key enzymes of C4 path­way of photosynthesis are very low. The primary products of photosynthesis after an 8 second pulse have been iden­tified as 3-phosphoglycerate and sugar phosphates. Thus, P. hysterophorus has been classified as a C3 - C4 intermediate species. One of the intrinsic features of C4 photosynthesis is the very high capacity for the internal refixation of CO2

originating from photorespiratory sources. It has been shown that C3 - C4 intermediate species like Moricandia ar­vensis, Panicum milioides and Flaveria pubiscens possess an in­creased reassimilation capacity (Holbrook et al., 1985; Bauwe et al., 1987). In this communication, we report that refixation of photorespiratory CO2 occurs in the leaves of P. hysterophorus which may be responsible for the low rates of apparent photorespiration in this species. We have also ob-

© 1989 by Gustav Fischer Verlag, Stuttgart

served that the rate of ammonia accumulation induced by MSO, an inhibitor of glutamine synthetase, is intermediate to those observed in wheat and sorghum.

Materials and Methods

P. hysterophorus L. growing wildly in our Institute campus was used for the study. Mature leaves were cut under water and placed in darkness for 30 minutes. The cut ends were immersed in water. Leaf slices (1.5 mm; 0.3 gm) were floated on 2 ml MES-KOH buffer (0.3 M, pH 5.5) containing 0.3 M sorbitol, 1 mM KH2 P04 and 1 mM Mg Cb in the outer well of the Warburg Flask. 0.3 ml of 20 per cent KOH was placed in the central well. Decarboxylation as­says were initiated by adding 0.2 ml of 5 mM photorespiratory sub­strate (18.5 KBq). Reactions were run either in light (800 ~mol m -2S-1) or in dark at 30 cC. After 1 h of incubation, 0.1 ml H 2S04

(3N) was tipped into the outer well from the side arm and flasks al­lowed to stand for 15 min. 0.1 ml KOH was removed from the cen­tral well and the radioactivity was determined by scintillation spec­troscopy. Specific activities of (l.l4C)-labelled glycolate and glycine were 370MBq/m mol and 740MBq/m mol, respectively. Similar assays of decarboxylation were performed with the leaf slices of

114 P. A. KUMAR and Y. P. ABROL

Table 1: Decarboxylation of (1)4C)-glycolate and (1)4C)-glycine by leaf slices of P. hysterophorus, T. aestivum and S. bieolor.

Species

P. hysterophorus T. aestivum S. bieolor

14C02 evolution {dpm/flask} (1 )4C)-glycolate {1 )4C)-glycine

Dark Light D/L Dark Light D/L

23633 3664 6.45 54679 5540 9.87 54170 40437 1.37 60085 28342 2.12 12120 1182 10.25 36715 2691 13.64

wheat {Triticum aestivum L. cv. Shera} and sorghum {Sorghum bi­color L. Moench cv. 296B}. Ammonium accumulation in the leaves of all the three species as induced by MSO and inhibited by isonico­tinyl hydrazide {INH}, an inhibitor of glycine decarboxylation, was estimated according to Kumar et al. {1984}. The experiments were repeated thrice. Representative data are presented.

Results and Discussion

Table 1 shows the rates of decarboxylation of exogenously supplied (V4C)-glycolate and (1- 14C)-glycine by the leaf slices of the three species studied in the dark and the light. Upon illumination the rates of 14C02 evolution from either of the photorespiratory substrates decreased in all the three species. The magnitude of this decrease was higher in sorghum, lower in wheat and intermediate in P. hyste­rophorus. This indicated that the internal refixation of 14C02 released from either glycolate or glycine is enhanced in P. hysterophorus relative to that in a C3 plant, wheat. The darkl light ratios of 14C02 evolution express the extent to which the process of reassimilation occurs and it is clear from Table 4 that 14C02 refixation takes place to a considerable extent in P. hysterophorus. Such an internal recycling of CO2 can ac­count for the low rates of photorespiration observed in P. hysterophorus (Rajendrudu and Das, 1981; Ku et al., 1985). Since this plant exhibits very low activities of the enzymes of C pathway (Ku et al., 1985), it is likely that the refixation of photorespiratory CO2 occurs via an efficient ribulose bis­phosphate carboxylase as observed in Panieum milioides and Moricandia arvensis (Holbrook et al., 1985).

During glycine decarboxylation, ammonia and CO2 are released in a stoichiometric manner. The ammonia released is reassimilated by glutamine synthetase (Keys et al., 1978). Berger and Fock (1983) employed MSO, an inhibitor of glut­amine synthetase, to estimate the rate of ammonia accumula­tion. They proposed that the rate of ammonia accumulation can be considered as a minimum estimate of the rate of

Table 2: Ammonium accumulation in the leaves of P. hysterophorus, T. aestivum and S. bicolor as affected by MSO and INH.

Treatment

1. Control {H20} 2. MSO {2.5 mM} 3. INH {35 mM} 4. MSO {2.5 mM} +

INH {35 mM} PRNH3 {2-4}

NHt {/-Imol g-l fr.wt. h- 1} P. hysterophorus T. aestivum S. bieolor

1.72±0.09 2.37±0.12 1.10±0.08 5.50±0.17 6.35±0.23 3.51±0.13 1.82±0.07 1.93±0.07 1.50±0.05 3.37±0.15 3.56±0.17 2.90±0.12

2.13 2.79 0.61

PRNH3 = protorespiratory ammonia.

glycine decarboxylation because glutamine synthetase was not completely inhibited in the beginning of the experiment. However, alternative sources of ammonia other than photo­respiration can also influence the rate of ammonia accumula­tion (Singh et al., 1985). In the present study, we used INH, an inhibitor of glycine decarboxylation, together with MSO (Table 2). The difference between the rates of ammonia accu­mulation caused by MSO and MSO plus INH was taken as the minimum estimate of the rate of glycine decarboxyla­tion. Wheat, sorghum and P. hysterophorus exhibited min­imal rates of 2.8 jlmol, 0.61 jlmol and 2.13 jlmol per gram fresh weight per hour, respectively. These results showed that the rate of photorespiratory ammonia (PRNH3) accu­mulation in P. hysterophorus was intermediate to those of C3

and C4 plants, thus confirming the intermediate character of this plant. It is also evident that photorespiratory nitrogen cycle is operative in P. hysterophorus.

References

BAUWE, H., O. KEERBORG, R. BASSUNER, T. PARNIK, and B. BASSUNER: Planta 172, 214-218 {1987}.

BERGER, M. G. and H. P. FOCK: Aust. J. Plant Physiol. 10, 187 -194 {1983}.

HOLBROOK, G. P., D. B. JORDAN, and R. CHOLLET: Plant Physiol. 77, 578-583 {1985}.

KEYS, A. J., J. F. BIRD, M. J. CORNELIUS, P. J. LEA, R. M. WALLS­GROVE, and B. J. MIFLIN: Nature 275,741-743 {1978}.

Ku, M. J. B., V. R. FRANCESHI, B. D. MOORE, and S. H. CHENG: Plant Physiol. 78, S-288 {1985}.

KUMAR, P. A., T. V. R. NAIR, and Y. P. ABROL: Plant Sci. Lett. 33, 303-307 (1984).

RAJENDRUDU, G. and V. S. R. DAS: Curro Sci. 50, 592-593 (1981). SINGH, P., P. A. KUMAR, Y. P. ABROL, and M. S. NAIK: Physiol.

Plant. 66, 169-176 (1985).