Pergamon Adv. Space Res. Vol. 14, No. 10, pp. (10)33-(10)39, 1994

Copyright © 1994 COSPAR Printed in Cn'eat Britain. All rights reserved.

0273-1177/94 $7.00 + 0.00

MUTATIONAL EFFECTS OF SPACE FLIGHT ON ZEA MAYS SEEDS

M. Mei,* Y. Qiu,* Y. He,* H. Bucker** and C. H. Yang***

* South China Agricultural University, Guangzhou~ P.R. China ** Institute of Aerospace Medicine, Cologne, Germany *** NASA Johnson Space Center, Houston, TX, U.S.A.

ABSTRACT

The growth and development of more than 500 Zea g~.= seeds flown on LDEF were studied. Somatic mutations, including white-yellow stripes on leaves, dwarfing, change of leaf sheath color or seedling color were observed in plants developed from these seeds. When the fre- quency of white-yellow formation was used as the endpoint and compared with data from ground based studies, the dose to which maize seeds might be exposed during the flight was estimated to be equivalent to 635 cGy of gamma rays. Seeds from one particular holder gave a high mutation frequency and a wide mutation spectrum. White-yellow stripes on leaves were also found in some of the inbred progenies from plants displayed somatic mutation. Electron microscopy studies showed that the damage of chroloplast development in the white-yellow stripe on leaves was similar between seeds flown on LDEF and that irradiated by accelerated heavy ions on ground.

INTRODUCTION

As the age of long-term space exploration comes, a deeper understanding of the biological ef- fects of space radiations in various organisms is needed. For examining the potential biologi- cal effects of ionizing radiation in space, maize seeds from special genetic stock were flown, re- covered, and planted; and, some developmental abnormalities were observed /1,2/. Peterson et al reported the finding of white-yellow stripes on leaves of a plant developed from one of the maize seeds flown on ASTP mission/3/. These maize seeds contained a recessive lemon white gene.

This report gives the results of our studies on the bioeffects of space radiations on ZeaKlay.~ seeds from similar genetic stocks. These seeds were flown to an altitude of 257 NM (476.27 km) in the Long Duration Exposure Facility (LDEF) as a part of the Biostack experiment.

METHODOLOGY

Dry seeds of Zea mays, heterozygous for LWl/IW 1 alleles, were obtained from two genetic stocks, namely Group I and Group I1. Group I was from Ratio Seeds Company in Indiana, USA,

(10)33

(10)34 M. Mei et al.

and Group II from Dr. M. Freeling of University of California, Berkeley, USA. The mutation of Lw 1 in embryonic cells for leaf development, induced by ionizing radiation, will cause white-yel-

low stripe formation on leaves 141. All seeds were kept in the plastic holders stacked in alu- minum containers in Tray A1 and A2 on LDEF during the space flight from April 7, 1984 to January 12, 1990 at the orbit of 28.5 °, 257 nm (476.27 km). Part of the seeds from the same genetic stocks were kept on ground in similar environmental conditions as that for flown seeds and used as control.

After retrieval, flown seeds kept in 11 holders and control ones were planted in a greenhouse at South China Agricultural University in August 1990 or August 1991. The growth and devel- opment of these seeds, including germination, plant growth, occurrence of white-yellow stripes on leaves, and variances in other morphological char-acters of plant were studied. Inbred progenies of plants with mutated characters were also investigated. In comparing with cells of normal green leaf, the structure of chloroplasts in mesophyll cells of white or yellow stripes were examined by transmission electron microscope.

RESULTS AND DISCUSSION

Germination and Growth Studies

Most of flown seeds were able to germinate normally in August 1990. However, when seeds were stored on ground for a long period of time after recovery from LDEF flight, the germina- tion rate declined significantly, as shown in Table 1. It appears that the environmental condition in the aluminum Biostack container in space had no obvious effects on the germination of seeds. Most of the germinated seeds from LDEF flight grew into plants normally, but with aver- age height shorter than that of control. Some germinated seeds failed to grow into plants due to infestation of ants and were not included in the data analysis.

Qccurance of White-Yellow StriDes on Leaves

White-yellow stripes displayed on leaves of some plants developed from flown seeds (Table 2). No plant in the control group showed any similar effect. Most of the stripes were found on the fourth to the sixth leaf, and only one stripe was observed in each plant studied. The length of stripes varied from 10.0 to 39.5 cm with the width about 0.1 cm. Some large stripes extended into the leaf sheath. A high percentage of plants with stripes on leaves, 6.5 - 13.6%, was found when the number of plants with stripes and the total number of plants developed from seeds from the same holder were used for the calculation (Table 2, F). The white-yellow muta- tion frequency was about 0.46 - 1.1% for holder 7-8 and 4-4 when the number of leaves show- ing stripes and the total number of leaves observed were used for the calculation (Table 2, L.F.). A comparison of these data with ground based experimental results, obtained from ir-ra- diated maize seeds of same genetic stock as Group I, can give an estimation of dose to which the seeds exposed in space. Figure 1 shows experimental results obtained earlier 151. Based on these dose-response curves, the estimated dose equivalent to gamma rays was about 13.1 Gy, a value much higher than that measured in LDEF, for inducing leaf stripe formation at a fre- quency of 0.46%. The L.F. value calculated from all survival plants from flown seeds of Group I

was 2.2xl 0 -3, which was equal to the ef-fect induced by gamma rays at a dose of 635 cGy in ground experiments (Figure 1). These estimated doses suggest that in addition to low-LET radiations the flown seeds might exposed to certain charged heavy ions from galactic cosmic rays, such as iron particles, which can induce mutation very effectively, as found from ground based studies (Figure 1). Information on fluences, charges, LETs, and number of particle hits in the embryo region from radiation dosimetry of Biostack Experiment on LDEF are needed to give a better explanation of such high frequency of stripe formation in some specific holders.

Effect of Space Right on Zea Mays (10)35

0.03

0°02

0.0046 0.0022

; I i

56Fe (600MeV/u )

I/ 4OAr (400MeV/u)

2ONe (A251]eV/u)

60Co aroma r a y s

6.35 10 13.1 20 30 DOSE (Gy)

Figure 1. Dose-response curves of white-yellow stripe formation in plant leaves after maize seeds exposed to various radiations (from/5/).

Table 1. Germination Rate of Maize Seeds

Flown Control

Group Ih

A. (Planted in 1990)

Flown Control

B. (Planted in 1991)

Flown Control

No. of Seed ~ of Seeds g.Er.mJJ~t~ Rate Plants

208 203 97.5% 151 74 67 92.5% 52

176 161 91.4% 112 62 57 91.9% 49

145 80 55.0% 74 19 9 47.3% 8

(10)36 M. Meiet al.

Other Somatic Mutations Observed

Plants developed from flown seeds of Group I were examined during the entire growth period, and several types of mutation were observed. Table 3 shows a summary of these observa- tions. Dwarf plants with height less than 50% of that of control were found in plants developed from seeds from every holder studied. Inducing dwarfing in plants is a common phenomenon for ionizing radiation. The high frequency of dwarf-ing plants observed in this study suggests that the radiation in space has similar ef-fects on plant development as on ground.

Other variation of morphological characters, such as the change of leaf sheath color from red- purple (control group) to green and the change of color of seedling from nor-mal green to yel- low-green, were also observed, as indicated in Table 3. Plants devel-oped from seeds in one holder (LBL I.D. #7-8), showed a high mutation frequency and wide mutation spectrum. A measurement of space radiation hitting this holder is required for a better understanding of the role of charged particles in inducing these mu-ations.

Table 2. White-yellow Stripes Observed in Plants Developed from Flown Seeds

Source of I.D. of Holder ~eeds LBL# DLR#

Occurrence of White-yellow No. of SurvivingStdDes in Leaves Rants No. plants R%~* L.F.**

Group I: Flown: Ratio 7-3 EB-125 47 seeds 7-4 EB-130 13

7-7 $7-158 46 7-8 $7-159 45

Total: 151

Control: 8-6 & 8-8

Flown: 1-2 $2-243 6-3 E5-59 6-4 E5-60 2-2 3-2 4-4 5-4

Total:

Control: 1-4 & 6-6

Group Ih M. Freeling seeds

1 4.2 0.0014 0 0.0 0.0000 1 2.1 0.0014 3 6.5 0.0046

5 3.3 0.0022

52 0 0.0 0.0000

37 46 29 8 14 22 30

186

2 5.4 0.0037 3 6,5 0.0049 1 3.4 0,0026 0 0.0 0.0000 0 0.0 0.0000 3 13,6 0.0110 1 3.3 0.0027

10 5.4 0.0040

49 0 0.0 0.0000

*F (frequency) = (No. plants with striped leaf)/(No, surviving plants) **L. F. (white-yellow striped leaves frequency) =

(No. leaves with white-yellow stripes)/(Total No. of leaves in plants)

Serf-bred progenies of mutants with white-yellow stripes on leaf or dwarfing character were grown and studied. White or yellow stripes were found on leaves of 2 - 5% of progenies from

Effect of Space Right on Zea Mays

each M 1 plant. And all progenies of dwarfing plant were dwarf.

(10)37

Figure 2. Electron micrograph of cross section of chloroplasts in mesophyll cells from (A) nor- mal green part of a leaf ('11,000x), (B) a yellow stripe of leaf of corn plant developed from a seed exposed to argon ions ('11,000x), (C) a white stripe of leaf, containing no chloroplast, of com plant developed from a seed exposed to argon ions, (D) normal green part of leaf of con- trol plant (*5,000x), (E) a yellow stripe of leaf of com plant developed from a seed flown on LDEF, and (F) a white stripe of leaf of corn plant developed from a seed flown on LDEF ('11,000x). *Film magnification.

(10)38 M. Meiet al.

Alteration of Chloroplast Structure in MesoDhvll Cells of White-Yellow Strioe

Figure 2 shows transmission electron microscope pictures of cross sections of meso-phyll cells from the stripes of leave of plants developed from seeds irradiated on ground or from flown seeds. For ground based experiments, the number of chloroplasts in the cell and the number of lamella per grana in chloroplasts from yellow stripes are smaller than that from normal parts of the green leaf. No chloroplast was observed in cells from a white stripe (Figure 2, B, and C). For the space flight experiment, the similar alteration was found (Figure 2, E and F). However, chloroplasts were still observed in the cell from a white stripe (Figure 2, F) although with few lamella in each grana. These results indicate that the mutation of Lw 1 gene induced by ioniz-

ing radiation may inhibit the transformation of protochloroplast to chloroplast, which alters the structure of the mature chloroplast.

Table 3. Other Somatic Mutation Observed in Plants Developed from Flown Seeds of Group I.

Mutation Observed Yellow-green Green Leaf

She h I. D. of No. Surviving Holder Plants ~ Plants F(%~* Plants F(%!* Plants R%~*

Flown:

7-3 47 4 8.5 2 4.2 0 0.0 7-4 13 1 7.7 0 0.0 0 0.0 7-7 46 6 13.0 3 6.5 0 0.0 7-8 45 12 26.7 1 2.2 2 4.3

Control:

8-6 & 8-8 52 0 0.0 0 0.0 0 0.0

*F (frequency) = (No. plants with mutated character)/(No, surviving plants) ** Plant height < 50% of the average plant height of the control group

ACKNOWLEDGEM ENTS

The valuable assistances from W. Lu, H. Wu of South China Agriculatural University, and Ms. L. Craise of Lawrence Berkeley Laboratory, University of California were greatly appreciated. We also wish to acknowledge Mr. Q. Zhang and Mr. X. Lun, Laboratory of Electron Microscope, South China Agricultural University, for electron microscope operation.

REFERENCES

1. J. V. Slater and C. A. Tobias, Effects of cosmic radiation on seeds differentiation and devel- opment, Radiat. Res. 19, 218 (1963)

2. H. J. Curtis and H. H. Smith, Corn seeds affected by heavy cosmic ray particles, .~,%7,J9/1~ 141,158-160 (1963)

3. D. D. Peterson, E. V. Benton, M. Tran, T. Yang, M. Freeling, L. Craise, and C. A. Tobias,

Effect of Space Hight on Zea Mays (10)39

Biological effects of high-LET particles on corn seed embryos in the ApoUo-Soyuz test project- Biostack III experiment. Life Science and z , ~ Research XV,151-155 (1977)

4. Y. Qiu, M. Mei, Y. He and Y. Lu, Mutagenic effects of accelerated heavy ion irradiation on Zea mays. , J . S o u t h C h i n a ~ . U B j ~ , 12, 48-54 (1991)

5. M. Mei, H. Deng, Y. Lu, C. Zhuang, Z. Liu, Q. Qiu, Y. Qiu and T. C. Yang, Mutagenic effects of heavy ion radiation in plant, Adv. Space Res. (1992) (this issue)

JASR 14:10-E