Indian 1. Plant Physiol., Vol XXXVll, No.3 pp. 164-168 (September, 1994)

PROTEINS, NUCLEIC ACIDS AND SOME ENZYME ACTIVITIES IN MAIZE PLANTS AS AFFECTED BY PRES OWING SEED TREATMENT WITH

R. DOGRA AND A.K. TIlUKRAL

Department of Botanical Sciences, Guru Nanak: Dev University, Amritsar 143 005.

Received on 26 March, 1994

SUMMARY TIle effect of pre-sowing seed treatment with some hormones (estrone, testosterone, and ) and cholesterol was studied on protein, DNA and RNA contents, peroxidase. catalase and acid andaikaUne phosphatase activities inthe leavesofZeamaysL. cv. Ganga­ S. Lower concentrations (10-8 and 10-6 M) were found tobe more effective than the higher one (1 ()-4 M). Estrone at 10-8 M concentration caused maximum enhancement in protein and nucleic acid contents. Peroxidase and catalase activities were influenced most by 10-8 M pregnenolone acetate and lo-c' M testosterone respectively. while acid and alkaline phosphatase activities were found to be enhanced maximum by 10-8 M testosterone and 10-6 M pregnenolone acetate.

INRODUCTION sterilizedwithO.Ol % ofmercuric chloride for one minute. After washing in double distilled water two-three times, The disco-very of steriod honnone's existence in seeds were soaked in different honnone concentrations higher plants by Dohrnetal. (1926) raised a vital question (l0'8, 10-6 and 10-4 M) prepared in Hoagland's nutrient about its role. These steroids isolated from various plant medium (Hoagland and Amon, 1939). Since the experi­ parts and different species, when tested on animals, have ment was also accompanied by laboratory studies on the been reported to produce effects similar to those of germination of seeds in Hoagland's nutrient medium animal's own steriod honnones (Geuns, 1978). When mended with the honnones, the same medium was chosen applied exogenously on plants, they are reported to affect for field studies too. Since maximum imbition occurred cell division and elongation, flowering, sex expression, during 3 hours, the seeds were soaked for this duration vegetative growth, synthesis of endogenous honnones prior to sowing in experimental plot. and by synthesizing more nucleic acids, proteins and certain enzymes (Cerana et al., 1985). Re­ The biochemical analysis for proteins, nucleic acids cently, several workers havy reported the endogenous and enzymes was done after 45 days of sowing, just prior presence of , , and to the initiation of flowering and fruiting. The youngest progestagens. Since maize is an important cereal crop, it leaves from the nodes no. 1 to 5, top downwards, of was envisaged to study the comparati ve effect ofsteroids different treatments were collected along with the control (estrone, testosterone, pregnenolone, hydrocortisone and plants for biochemical estimations. The protein content cholesterol) and plant growth regulators with an objective was determined following the method of Lowry et al. to produce baseline data to understand the biochemical (1951). Quantitative estimations of DNA and RNA were effects of honnones in maize. done by the method proposed by Schneider (1957). The activities of peroxidase and catalase were detennined MATERIALS AND MEnIODS following Kar and Mishra (1976) while the activities of The experiment was conducted under field condi­ acid and alkaline phosphatases were measured following tions. Healthy and unifonn-sizett-seeds were surface McIntyre (1971).

I f , ~ I

PRE-SOWING SEED TREAlMENT WITH STEROIDS IN MAIZE 165

The biochemical differences between the treatment nenolone acetate in comparison with the controL Simi­ and control were tested through ANOVA and Tukey's larly, the RNA content was found to be enhanced most by multiple comparison test (Meyers and Grossen, 1974). 10-8 M estrone (Table II). RESULTS AND DISCUSSION There was a significant enhancement in enzyme activities atlowerconcentrations ofthe hormones. 10-8 M The pre-soaking treatment of seeds caused a signifi­ pregnenolone acetate caused maximum rise in peroxidase cant enhancement in proteins, nucleic acids and enzymes. " activity as compared to the control (Table III). Catalase The highest protein content in the leafsamples was found activity increased most by 10-6 M testosterone followed at 10-8 M concentration of estrone treated ones with by 10-6 M pregnenolone acetate in comparison with the respect to control (Table I). Amongst nucleic acids, : control (Table fiI). 10-8 M testosterone was found to be the I maximum DNA content ofthe leaves was observed at 10­ most effective treatment in enhancing acid phosphatase 8 M concentration of estrone followed by 10-6 M preg­ activity while alkaline phosphatase activity was increased maximum by 10-6 M pregnenolone acetate (Table IV). TableI: Water soluble protein content (mg g-! f. w.)inthe The steroid pre-treatment to seeds enhanced the final leaves of maize plants. as affected by steroids and plant productivity and seed number and weight significantly in hormones maize (Dogra and Thukral, 1969). The time taken for the emergence ofcobs was much reduced in the plants when Hormone Concentration (M) HSD value atP

Table n:DNA and RNA content (mg g-1 f. w.) inthe leaves ofmaize plants as affected by~ and plant hormones

Hormone Concentration (M) HSD value atP < 0.05 0 10"8 10-4 10-4 DNA RNA DNA RNA DNA RNA DNA RNA DNA RNA.

. Esttone 37.2 37.7 71.2' 75.0­ 65.2" 66.9­ 61.2­ 62.0­ 6.26 6.34 Testosterone 37.2 37.7 64.5" 65.3­ 58.5­ 60.0­ 51.9­ 52.5" 5.73 6.24

. Pregnenolone acetate~ 37.2 37.7 60.5­ 61.3­ 67.8­ 68.7­ 52.5­ 55.2­ 6.05 6.24 Hydrocortisone 37.2 37.7 61.2­ 61.3­ 60.5­ 62.0­ 52.5­ 53.2' 7.02 7.11 Cholesterol 37.2 37.7 45.9' 50.5­ 5l.2' 53.2­ 41.9 45.8­ 6.16 6.44 Indole-3-acetic acid 37.2 37.7. 47.9­ 57.3' 54.5" 51.9" 51.9" 52.5' 7.62 7.02 Gibberellic acid 37.2 37.7 51.9' 52.5" 58.5­ 59.3" 50.5­ 51.2­ 5.62 5.04 63.8­ 64.7­ 57.9" 59.3­ 44.5­ 5.62 5.62 5.00 Kinetin , 37.2 37.7 a denotes the level of significance at 5%P

"'" 166 R. IX)(}RA AND A.K. THUKRAL

Table m:Peroxidase (Ilmol5 min-!) and Catalase activity (mM II.02 decomposed g-! oftissue 5 min-!) in maize plants as affected by steroids and plants hormones

Peroxidase Catalase

Hormone Concentration (M) HSD value Concentration (M) HSD value at p

Estrone 0.251 0.450' Q457' 0.405" 0.100 0.047 0.086' 0.108' 0.046 0.006 Testosterone 0.251 0.476" 0.495' 0.416' 0.059 0.047 0.099' 0.118' 0.061' 0.004 Pregnenolone acetate 0.251 0.694' 0.656' 0.489' 0.042 0.047 0.112" 0.114" 0.061" 0.004 Hydrocortisone 0.251 0.311 0.435" 0.274 0.096 0.047 0.103' 0.113' 0.053' 0.004 Cholesterol 0.251 0.469' 0.540' 0.379' 0.056 0.047 0.105' 0.111' 0.060' 0.004 Indole-3-acetic acid 0.251 0.240 0.476' 0.270 0.074 0.047 0.090' 0.089' 0.049 0.004 Gibberellic acid 0.251 0.259 0.390' 0.203 0.063 0.047 0.101' 0.092' 0.053" 0.004 Kinetin 0.251 0.364' 0.499­ 0.255 0.105 0.047 0.089' 0.096" 0.052' 0.004 a denotes the level of significance at 5%P

Table IV : Acid and alkaline phosphatase (ACP and ALP) activity (mM) in maize plants as affected by steroids and plant hormones

Hormone Concentration (M) F HSD value value atp

ACP ALP ACP ALP ACP ALP ACP ALP ACP . ALP ACP ALP

Estrone 0.178 0.254 ,0.225' 0.391* 0.248' 0.461' 0.166 0.263 42.3' 342.7' 0.028 0.026 Testosterone 0.178 0.254 0.365' 0.436' 0.321' 0.467' 0.193 0.315' 150.1' 167.0' 0.036 0.035 Pregnenolone acetate 0.178 0.254 0.301' 0.359' 0.359' 0.508' 0.272' 0.353' 205.9' 293.9' 0.025 0.029 Hydrocortisone 0.178 0.254 0.266' 0.420' 0.304' 0.450' 0.291 0.388' 49.6' 200.3' 0.039 0.029 Cholesterol 0.178 0.254 0.229' 0.465' 0.301' 0.453' 0.156 0.363' 42.3' 233.7' 0.028 0.032 Indole-3-acetic acid 0·t78 0.254 0.215' 0.388' 0.228' 0.312' 0;158 0.266 42.3' 96.8' 0.028 0.030 Gibberellic acid 0.178 0.254 0.362' 0.365' 0.300' 0.315' 1).189 0.283' 150.1' 93.4' 0.036 0.026 Kinetin 0.178 0.254 0.309' 0.391' 0.351' 0.461' 0.279' 0.263 205.9' 342.7' 0.025 0.026

a denotes the level of significance at 5%P

revealed that as in animals, these steroids seems to be the the plant growthhorrnones (Harborne, 1977). Some ofthe integral part of the plants and control their growth, important biochemical processes in animals which are flowering and sex expression at different phases of the induced by the application of steroids" include stimula­ development. Their function is of course still open to tion of DNA and RNA precursors, and their synthesis, question. However it is also suggested that they may ,­ nucleotide metabolism, polysome abundance, protein control different physical and physiological responses of and phospholipid synthesis and increflSe in mitotic activ- I' PRE-SOWlNG SEED TREAIMENT wrrn STEROIDS IN MAIZE 167

Table V : Emergence ofcobs on maize plants as affected ity, etc. (O'Malley and Means, 1974). Similar results by steroids and plant hormones were obtained when they were applied to plants. The Growth regulator Concentration Percentage emergence after earlier studies with the seedlings of maize plants also (M) 55 days 60 days 65 days showed enhanced rate ofmetabolism. There was a signifi­ cant increase in the vegetative growth, biochemical con­ Control 0 0.0 30.5 75.6 stituents (proteins, DNA and RNA) and enzymes of the Estrone 10-8 24.5' 48.9' 89.2 10-6 23.2' 45.1 90.3 10-day old maize seedlings when cultured in different 10-4 20.1 40.2 80.1 hormone concentrations and lower concentrations were 8 Testosterone lOasdfjk1;rgh- 27.1' 57.1' 92.5 found to be more effecti ve than the higher one. A number 10-6 22.1 46.2' 90.1 10-4 18.0 35.1 85.2 of reports are available describing the effects of steroids Pregnenolone acetate 10-8 25.0' 55.2' 93.5 on vegetative andreproducti ve parameters ofa number of 10-6 27.2' 52.3' 88.2 plants, but no information is yet available in maize, an 10-4 21.0 40.6 77.5 -Hydrocortisone 10-8 27.0' 55.0' 96.2 important crop of India. 10.6 30.1' 65.1' 98.5 The enhanced vegetative growth of the maize plants 10-4 19.3 43.5 90.1 Cholesterol 10-8 26.1' 54.2' 95.1 under the iilfluence of steroids finds support from the 10.6 25.2' 50.1' 97.2 earlier studies by Geuns (1978) in mung bean plants. An 10-4 20.5 45.2 82.2 enhanced RNA synthesis was noted under treat­ Indole-3-acetic acid 10-8 25.1' 49.2' 87.2 10-6 24.2' 45.3 90.3 ment in mung bean roots. Similarly, brassinosteroids 10-4 20.1 43.2 81.2 were reported to enhance proteins, nucleic acids and DNA 10.8 Gibberellic acid 26.4' 55.1' 95.2 and RNA polymerases in several plants (Mandava et al. 10-6 23.2' 55.2' 82.3 10-4 19.3 43.5 78.5 1987). Inthe present study, increased contents ofproteins Kinetin 10.8 26.0' 52.9' 94.2 and nucleic acids and the activities of enzymes might 10-6 25.0' 50.0 93.5 represent an enhanced metabolic rate and vegetative 10-4 20.0 47.1 85.6 growth of the plants. *Significantly different from the control at p

Table VI : Number ofcobs per plant, number of seeds per cob and lOO-seed weight of maize as affected by steroids and plant hormones

Growth regulator Number of cobs per plant Number of seeds per cob l00-seed weight Concentration (M) HSD Concentration (M) HSD Concentration (M) HSD value value value 10-8 10-6 10-4 at 10-8 10-6 10-4 at 10-8 10-6 10-4 at p<0.05 p<0.5 p<0.05

Estrone 5.D' 4.9" 4.0" O.~ 405.P 401.5' 364.3' 20.1 32.0 31.9 30.0 Testosterone 4.8" 4.5" 4.2' 0.7 399.2' 407.1' 345.5 15.6 32.1 32.0 33.2 Pregnenolone acetate 5.2' 5.1" 3.8 0.6 410.0" 399.2' 365.1' 27.9' 31.6 31.6 31.8 Hydrocortisone 5.D' 5.D' 4.8' 0.4 390.0' 369.1' 341.3' 10.2 31.3 31.9 31.6 Cholesterol 5.D' 5.l" 4.8" 0.5 387.1' 370.2" 310.3 11.6 31.9 31.4 30.8 Indole-3-acetic acid 4.7' 4.8' 4.0 0.7 359.D' 357.2' 319.5 13.9 31.9 31.9 31.4 Gibberellic acid 5.D' 4.7' 4.1' 0.7 360.1' 364.D' 328.0 25.1 31.6 32.0 31.7 Kinetin 4.9' 4.6' 4.0 0.7 358.D' 360.1' 318.2 15.1 31.6 32.0 31.2 '.' Control 3.4 330.0 30.6 ." -; Not significant at p<0.05. Asterisks represent the level of significance for ANOVA p

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