684 BOTANY: EPSTEIN, RAINS, AND ELZAM PROC. N. A. S.

As long as ZiveqI << 1/4, the equilibrium remains close to the "valley state." Ac-cording to equations (16) and (21), this requires fairly low temperatures (,A >> 1).Specifically, in experiments on persistent currents where v (the magnetic flux quan-tum number) is often a very large number (-.105, say), the condition is T <K eT,where 1/e, for InI v| >> 1,8 may be written in the form of an expansion:

1/e = 0.57[Inj vI + 1/2 In (327r lnf v| + ...)] (24)

As the temperature is raised toward the value T, = eTca Oveq approaches the criticalvalue - 1/4 (if v > 0); then any initial state (_ 1/4 < SP < 1/4) necessarily decaysin the direction of decreasing current (d In (P,)/dt < 0).

Finally, we raise T above T1. The free eheingy would then certainly be loweredif the system were allowed to "drop into the neighboring valley" (P Pv- 1/2, ifv > 0). But this would involve, as Bohr and Mottelson' say, a "macroscopic modi-fication" of the system: a large number of Cooper pairs would have to change theirwavefunction. Before doing so, the system presumably prefers to let the backflowgrow (aS < - 1/4) while v remains unaltered: the pairing forces remain effective inthe same k' space. The metastable state finally reached is again determined byequations (21-23), in accordance with the two-fluid model (but now bveq < 1/4)-Clearly, in the temperature interval T, < T < To, the half-integral nature of v is notdirectly relevant for the metastability of the terminal state, although it provides amore convincing reason for the constancy of v during the decay process.

* This work was supported in part by the U.S. Atomic Energy Commission.1 Bohr, A., and B. R. Mottelson, Phys. Reu., !25, 495 (1962).2 Bardeen, J., Phys. Rev. Letters, 1, 399 (1958). I am grateful to Professor Bardeen for corre-

spondence on the two-fluid model.3 See, e.g., Byers, N., and C. N. Yang, Phys. T cv. Letters, 7, 46 (1961).4 Parikh, J., Phys. Rev., 128, 1530 (1962).6 Wentzel, G., Phys. Rev., 120, 1572 (1960); earlier references are listed there. Also in the

transport problem, in Momentum Transfer, Hre. is negligible; see Parikh, J.,4 Appendix B.6 Allowing A to depend on w2 [see Bardeen, J., Rev. Mod. Phys., 34, 667 (1962), Appendix BI

does not affect the main part of our argument, concerned with terms linear in w.7 Note that, contrary to our notation, the momentum of a quasiparticle of "type 1" (spin-)

is called -k in ref. 4.8 The condition owkF << 1 in equation (16) sets an upper bound to Jv| but still admits fairly

large values of In 1vi (.10, say).

RESOLUTION OF DUAL MECHANISMS OF POTASSIUM ABSORPTIONBY BARLEY ROOTS*

BY EMANUEL EPSTEIN, D. W. RAINS, AND 0. E. ELZAM

DEPARTMENT OF SOILS AND PLANT NUTRITION, UNIVERSITY OF CALIFORNIA (DAVIS)

Communicated by G. Ledyard Stebbins, March 21, 1963

The concept that ions are transported into plant cells through transient attach-ment to carriers, initially based on a variety of experiments and considerations,1has more recently received specific support from the findings that the kinetics ofabsorption are in close agreement with classical enzyme kinetics.2 Among the

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chief advantages gained through this approach is the fact that the enzyme-kineticanalysis of transport provides a rationale for selectivity. Selective transport isinterpreted in terms of binding of the ions to ion-specific or ion group-specificsites of carriers. Competition among ions, on equal or unequal terms, and otherkinds of interaction become amenable to quantitative treatment and to interpre-tation in terms of differential affinities of carrier sites for different ions. Researchalong this line" I has led to the conclusion that a multiplicity of carrier sites isinvolved in the transport of the many ionic species to which roots and other tissuesare exposed and which they selectively absorb from the inorganic substrates.

Chemically related ions may have virtually equal affinities for common carriersites, as evidenced by their mutually competing on almost equal terms. Thepair K-Rb is a case in point.2 4 More surprising was the finding that a single ionicspecies, Rb, was found to be transported by two discrete mechanisms, one operatingat low external Rb concentrations and very indifferent to Na, the other cominginto play at high Rb concentrations and possessing pronounced affinity for Na aswell.2 Since recent research4 has revealed Ca ions to be indispensable to theunimpaired functioning of selective K-Rb transport, the work presented in thispaper was undertaken in the expectation that inclusion of Ca in the experimentalsolutions, together with improved experimental technique,5 would permit a moreprecise resolution of the two K-Rb transporting mechanisms than has been achievedso far, as well as a clear definition of the effect of Na on the two mechanisms.This expectation was realized.Methods.-Culture of seedlings: Seeds of barley, Hordeum vulgare var. Arivat, were germinated

and seedlings grown in the dark in aerated solutions of 2 X 10-4M CaSO4, as described earlier.4The absorption experiments: Samples of roots weighing 1.00 gm (fresh weight) were suspended

for about 30 min in an aerated solution of 0.5 mM CaCl2 at 300C prior to the absorption period.Just before the absorption period, each sample was rinsed for 1 min with fresh 0.5 mM CaCl2 at300C, excess solution was spun out, and the sample was transferred to the aerated experimentalsolution at 300C.Except in the experiments with sulfate as the anion, each solution contained, in addition to the

chloride of the monovalent cations under consideration, CaCl2 at 0.5 mM. The pH of the experi-mental solutions (unbuffered) was 5.9, and seldom varied by more than 0.2 pH units during theexperimental runs. Differences in pH of this magnitude do not measurably affect absorption ratesat any of the concentrations used. The volumes of the experimental solutions ranged from 2,000ml at the lowest substrate ion concentration used (0.002 mM) to 250 ml at the highest concentra-tions. Volumes were chosen in such a manner that depletion of the solutions due to absorption bythe roots did not lower the concentration of the substrate ion by more than 5-6% during the 10min experimental run.

Substrate ions were radioactively labeled, K with K42 (subsequently called K*) and Rb withRb"4' (Rb*). After absorption from solutions in the higher concentration range used (0.5 to 50mM K* or Rb*), the roots contain a fraction of readily exchangeable K* or Rb* ions held by super-ficial exchange sites probably associated with the cell wall.'. 6 These readily exchangeable K* orRb* ions were desorbed at the end of the absorption period by a 30 min exposure of the tissue to.an aerated, cold (50C) solution of 5.0 mM KCI and 0.5 mM CaCl2.5 Details of the experimentaltechnique have been described.5 Modification of the procedure in experiments in which the anionwas sulfate instead of chloride will be described in connection with those experiments. Rates, v,throughout are reported in terms of ,umole/g/hr, and concentrations of substrate salts in terms ofmM for chlorides and meq/l for sulfates, so that equal numbers refer to equalmM concentrationsof the monovalent substrate cations. Rates were constant for at least ;1 hr at all concentra-tions.5,7

Resgults.-Figure 1 presents the results of an experiment on K absorption from

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25,24

n- ~~~~~~~~~~~~~~~~~02221-

20

19-

is K

16 /

15-

13-

°E 12 ----------------------------------

3*

0.02 004 006 006 0.90 0* OS 0X Of00S.5 5 10 ff 20 25 S0 M6 40 45 50

(S).mM

FIG. 1.-Rate of K absorption as a function of the concentration of KCl. CaCl2,0.5 mM throughout. Temperature, 300C. The circles represent experimental data;the solid line represents equation (1), with the following parameters: KM 1 0.021 mMN,Vmax 1 11.9 ,umole/g/hr, KM 2 11.4, Vmax 2 13.2. The dashed line represents the firstterm only of equation (1).

solutions ranging in concentration from 0.002 mM KCl to 50 mM. In order toaccommodate the data over this wide range of concentrations, the horizontal scaleonly is changed between 0.2 and 0.5 mM. The points, as in all figures, representthe data from a single experiment, unreplicated. Quantitative analysis of theabsorption kinetics and the final equation arrived at were based, first of all, on theobvious conclusion that we are dealing with two absorption mechanisms. Over the0.002 to 0.2 mM concentration range of K, a single Michaelis-Menten term de-scribes the relation between concentration and rate of absorption. The mechanismdefined by this term operates almost at saturation level at 0.2 mM K. The equa-tion for this term was obtained by making a Lineweaver-Burk plot,2 i/v versus 1()of the data. This plot yielded a straight line which was determined by the leastsquare method. The calculation yields the values for slope and intercept of theLineweaver-Burk plot, from which the values for Vmax and KM were calculated.These values were inserted into the Michaclis-Menten equation, first term of (1),yielding the theoretical absorption rates, v, represented by the solid line at the lowconcentrations, continued by the dashed line at the higher couceutrations. (Fig. 1).

V= Vmaxi1(S) + VmaX2(S)(1KMl + (S) I(M2 ± (S)

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Over the high concentration range of 0.5 to 50 mM K, absorption rates consistof two terms: the contribution from the low-range mechanism 1 discussed above,operating virtually at maximal velocity, and a contribution from a second mech-anism with much lower affinity for K. The contribution by this mechanism 2 wascalculated separately. First, the theoretical contribution to the observed velocityfrom the low-range mechanism 1 was calculated for each high-range concentration(dashed line) and subtracted from the experimental value. The resulting v2values, representing velocities due to the upper-range mechanism 2 only, were usedto construct a Lineweaver-Burk plot which gave a straight line whose y interceptand slope were calculated as indicated above, yielding values for Vmax 2 and KM2for this mechanism 2 only. The actual velocity, v, represents the sum of the con-tributions to the total velocity from both mechanisms (equation 1).

Equation (1) with the constants given in the legend of Figure 1 describes therelation between the K concentration in the solution and the rate of K absorptionover the 25,000-fold range of concentrations explored (the solid line of Fig. 1).

Identity of K and Rb transporting mechanisms in this tissue has been demon-strated before.2'4 Figure 2 gives the results of an experiment in which theabsorption of Rb instead of that of K was investigated. The relation betweenconcentration and rate of absorption of Rb (from RbCl) closely resemblesthat for K. For the low concentration (high affinity) mechanism 1, the M/lichaelis

20

19

16_

5 - O: from Cl

14 Rb

13 a: from SO412 -

A~~~~~~~~~~~~~~~~~

0 10-E

7/

0O 004QO0.0 0J0 0.12 0 0J6 0JS 2 0.5 5 10 IS 2025 30 3 40 45 SO

CS) . me/

FIG. 2.-Rate of Rb absorption as a function of the concentration of RbCl, over the range 0.01to 50 mM, with CaCl, at 0.5 mM, and as a function of the concentration of Rb2SO4, over the range3 to 50 meq/l, with CaSO4 at 0.5 mM. Circles and triangles represent experimental data; thesolid line over the low range, continued by the dashed line, represents the first term of equation(1), with these parameters: Km 1 0.017 mM, V,,1 10.0 j.mole/g/hr. The solid lines over theupper range represent equation (1) with the above parameters for the first term and, for bKlI,KM 2 14.5, VMan 2 11.8; and for Rb2SO4, KM 2 37.9 meq/l, V.. 2.3.

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constant Km for K is slightly higher than that for Rb, whereas the reverse is thecase for the high concentration (low affinity) mechanism 2. Maximal velocities areslightly higher for K than Rb absorption by both mechanisms, suggesting that ratesof metabolic turnover in the presence of K are somewhat greater than with Rb.The high affinity mechanism 1 of K-Rb uptake is very indifferent to Na ions.

Figure 3 shows a Lineweaver-Burk (1/v versus 1/ (S)) plot of K absorption over the

0.5 H

0.4K/

FIG. 3.-Lineweaver-10.3 _K-NEBurk plot of K absorp-tion over the range 0.005to 0.2 mM KC1, in the

0.2 - absence of NaCi and itspresence at 0.5 mM.Other conditions as for

.1 ffiFig. 1.

50 loo lso 200

low concentration range (0.005 to 0.2 mM), in the absence of Na and its presence at0.5 mM. Competition by Na is slight, the Km for K being 0.026 mM, whereasthe K, for Na is 1.25. Maximal velocities V.,, were 11.9 pmole/g/hr in the absence,and 12.2 in the presence, of Na, evidence that the interference by Na, though weak,is competitive in nature.While Na competes ineffectively in the high-affinity mechanism 1 of K-Rb ab-

sorption, it is a potent competitor in the low affinity mechanism 2 which comes intoplay in the upper concentration range. Figure 4 presents the findings for K ab-

1.0

FIG. 4.-Lineweaver-0.8 ' - Burk plot of K absorption

over the range 3 'to 50mM KC1, in the absence

v. °6~ofNaCl and its presenceat 2.0 mM. Other condi-itions as for Fig. 1.

0.4i Rates v represent the con-tribution of the secondterm of equation (1) only--see text.

0.05 00 o05 0.20 025 0.30 0.35

(S)

sorption rates over the 3-50 mM range of K concentrations, in the absence of Naand in its presence at 2.0 mM. The data are presented in the form of Lineweaver-Burk plots The rates, v2, the reciprocals of which are plotted on the y axis, repret

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sent the contribution from the low affinity mechanism 2 only. The contributionmade by the high affinity (low concentration range) mechanism 1 was independentlydetermined and subtracted from the total rates obtained over the 3-50 mM rangeof concentrations, as described above. The Michaelis constant for K, KM, is 17.3mM, and the constant, K, for Na is 0.84 mM. The interference by Na with K ab-sorption is competitive. In a similar experiment on Rb absorption over the sameconcentration range, Na at 10mM virtually eliminated Rb absorption by the upper-range mechanism 2.

In the above experiments, the anion was chloride throughout. Since chlorideitself is absorbed at appreciable rates by this tissue, K-Rb absorption was studiedin the presence of an anion whose rate of absorption is very much less. Sulfate issuch an anion.8 In the experiments with SO4 the roots, grown initially in CaSO4solution as always, were kept before the absorption period proper in an aerated solu-tion 0.5 mM with respect to CaSO4 instead of CaCl2, were rinsed just prior to theabsorption period with 0.5 mM CaSO4 solution, and were then transferred to theexperimental solutions which contained Rb2*SO4 and in addition, 0.5 mM CaSO4.That is, treatment of the tissue with respect to Ca was precisely as in the other ex-periments, but SO4 was substituted for Cl throughout. (The original, high specificactivity Rb*Cl solution used to tag the Rb2SO4 stock solution contributed Cl to the-extent of I//36m of the SO4 present, on an equivalent basis.)

Figure 5 shows the results of an experiment on Rb absorption over the low range

10 Hofrom S04

FIG. 5.-Rate of Rb absorp- 7 " 0

tion as a function of the concen- X

tration of Rb2SO4. CaSO4, 0.5 t 6mM throughout. Other condi- Z 5tions as in Fig. 1. The circles Erepresent experimental data; ;the line represents the first term 3of equation (1), with these pa- 2rameters: KM 10.016 Ivmax 1I9.1 /hmole/g/hr.

0.02 004 0.06 0.08 0.10 0.12 Q14 0.16 0.16 0.20

Ms), mWe

of concentrations of Rb2SO4 (0.002 to 0.2 meq/l). The solid line represents therelation described by equation (1) with Vmax and KM values given in the legend ofthe figure. These values were calculated by the least square method from a Line-weaver-Burk plot of the data. Results were virtually identical with those obtainedwhen the anion was chloride, over the same range of Rb concentrations (Fig. 2).The same indifference to the identity of the anion is not characteristic of the ab-

sorption mechanism operating at the higher concentrations (mechanism 2). Figure2 shows the result of an experiment in which absorption of Rb from RbCl and Rb2-SO4 was compared, at the high concentration range (3-50 meq/l). Analysis of thekinetics was as described above. The contribution to the velocity from the lowconcentration range mechanism 1 was determined using RbCl, since there is nosignificant difference in the Vmax 1 obtained for this range, whether Cl (Fig. 2) or

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S04 (Fig. 5) is the counterion. Over the high concentration range the results ofthe Cl run in this experiment closely resemble those obtained in other experimentswith Rb and K. With SO4 as the anion, on the other hand, Rb absorption ratesare very much lower. Whereas for the Cl run, the KM2 for Rb was 14.5, it was37.9 with SO4. Maximal velocities Vmax 2 were 11.8 and 2.3 gmole/g/hr with Cland SO4 anions, respectively.

Discussion.-Kinetics: A single equation consisting of the sum of two Michaelis-Menten terms (equation 1) describes with impressive precision the relation betweenthe external concentration of K or Rb and the rate of absorption of these ions bybarley root tissue, over a 25,000-fold range of external concentrations (0.002 to 50mM). These findings and the approximately 500- to 1,000-fold difference in the Kand Rb KM values for the two Michaelis-Menten terms are evidence for the dualityof the K-Rb transport mechanisms. Sodium competes in both mechanisms, butits affinity for the low concentration range mechanism 1 is on the order of 1 per centof the affinity of K-Rb for this mechanism, as judged by the Km and KI values,whereas in mechanism 2 the affinity of Na exceeds that of K by a factor of about20.

Absorption of K-Rb by the high affinity mechanism 1 is autonomous cationtransport independent of the concentration of the anion and the rate of absorptionof the anion. In the Cl experiments, CaCl2 was present throughout at 0.5 mM.The concentration of Cl therefore varied between the narrow limits of 1.0 and 1.2mM, over the 0.002 to 0.20 mM range of K or Rb concentrations. At 0.20 mM Kor Rb, this mechanism operates virtually at saturation level, while yet the Cl con-centration is 6 times that of the cation. The rate of cation transport is therefore afunction of the degree of saturation of entities binding the cations in the process oftransport, irrespective of the extent of concomitant anion transport. The rate ofK-Rb transport in the presence of Cl is identical over this range with that observedwith S04 as the counterion-an ion whose rate of absorption is extremely low com-pared with that of K-Rb and of Cl. This is further evidence for the autonomousnature of K-Rb transport, in keeping with earlier conclusions.1 4. 6 9 The mecha-nism is by exchange of the cations absorbed with H ions which are released.6 9 10Over the high concentration range, Rb absorption from S04 solutions is greatly

depressed, compared with absorption from Cl solutions. This influence of theanion on the rate of absorption of the cation has been the general experience in thepast (except 9). For example, Hoagland"1 reported that "accumulation of potas-sium from the sulphate was much smaller than from the chloride, bromide, or nitrate,as though the rate of accumulation of the cation, or the equilibrium value attained,was markedly influenced by the mobility of the anion." The experiments leadingto those conclusions were done at concentrations of salt falling within the highrange where mechanism 2 (the low affinity mechanism) participates in absorption.The present experiments show that the anion has no such influence on K-Rb ab-sorption mediated by the high affinity mechanism 1 operating at low external con-centrations. The differential response of K-Rb transport to the identity of theanion at low and high cation concentrations constitutes further evidence for opera-tion of two discrete K-Rb absorption mechanisms.

Experiments on mechanisms of ion transport are conventionally done at con-centrations which are high in terms of the K and Rb Km, and KMI values reported

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here- so high that both mechanisms can be expected to make contributions to thetotal observed rate of absorption. Quantitative relationships are often sought be-tween the velocities observed and parameters such as respiration rates'2 or mem-brane potentials.'3 It will be necessary, in attempting such correlations, to takecognizance of the evidence that rates of absorption observed at these concentrationsmay represent summations of terms contributed by different transport mechanisms.The duality of K-Rb transport does not imply that Michaelis-Menten kinetics

are not followed, as has been claimed.'4 Such duality was first inferred from ex-periments showing conformity with double Michaelis-Menten kinetics,2 and sub-sequent confirmation of this concept has similarly been based mainly,8' 9, 15, 16though not exclusively,4' 1' on analysis in terms of Michaelis-Menten kinetics. Thepresent experiments, done unlike the others in the presence of Ca, sharply differen-tiate between two K-Rb transport mechanisms, and demonstrate the applicabilityof Michaelis-Menten kinetics to both.An alternative interpretation: Dual Michaelis-Menten terms in ion transport

have heretofore been interpreted on the basis of two kinds of binding sites withdifferent affinities for one and the same ionic species. Another interpretation ofsuch kinetics is possible. If, at low concentrations, one substrate ion is bound percarrier site, but at higher concentrations two ions of the substrate element are boundper active carrier site, and are bound more loosely than is the single ion, thenkinetics like those observed would be expected (cf. ref. 18). If we are dealing withdiscrete sites, the possibility must be borne in mind that they may deliver ions intodifferent compartments or "inner spaces."

Physiological significance: The high affinity mechanism 1 of K-Rb absorptionoperates at half-maximal velocity at an external concentration of about 0.018 mM.This corresponds to 0.7 ppm K-a concentration of the order expected in soil solu-tions. Much lower concentrations yet may prevail in the immediate microenviron-ment of roots in soil. It is therefore very significant that of two demonstrable Kabsorption mechanisms in barley roots, one has a very high affinity for K ions.Recent experiments'9 have demonstrated that barley plants can grow well in solu-tions maintained at the very low level of 0.02 to 0.03 ppm K, or 0.0005 to 0.00075mM. At these concentrations, the high affinity mechanism 1 would operate, underour experimental conditions, at several per cent of its maximal rate, while the con-tribution from the low affinity mechanism 2 would be infinitesimal.

Selectivity in the absorption of K and Na by growing plants is likely to be anexpression of the operation of specific transport mechanisms like those describedhere. Plants growing in many saline soils absorb K from solutions with very wideNa/K ratios. This suggests existence of absorption mechanisms with much higheraffinity for K than for Na, like the high affinity mechanism 1 described here. Bothmutual interference between these elements and failure to interfere have been en-countered in conventional, long-term experiments. Interpretation of thesephenomena in terms of dual mechanisms with different affinities for K and Na willprove rewarding.Summary.-The relationship between the rates of absorption of K and Rb by

barley roots and the concentration of these ions in the external solution, over therange 0.002 to 50 mM, is predictable on the assumption that two carrier sites bindand transport the ions. One of these operates at half-maximal velocity at a con-

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692 BIOCHEMISTRY: CHIPCHASE AND BIRNSTIEL PROC. N. A. S.

centration of about 0.018 mM, with very low affinity for Na, the second at about16 mM, with severe competition by Na. The latter but not the former mechanismis inhibited when SO4 is the anion instead of Cl.

* This research was supported by grants from the National Science Foundation and the Office ofSaline Water, U.S. Department of the Interior. Rb86 was furnished by Oak Ridge NationalLaboratory, Oak Ridge, Tennessee. K4' was obtained by neutron bombardment of K2CO3 in theLPTR facility of the University of California Lawrence Radiation Laboratory at Livermore,through the courtesy of the Director, Dr. John S. Foster, Jr., and Dr. Edward Teller. We appre-ciate the interest and cooperation of Mr. Ernest E. Hill, Reactor Supervisor.

I Epstein, E., Ann. Rev. Plant Physiol., 7, 1 (1956).2 Epstein, E., and C. E. Hagen, Plant Physiol., 27, 457 (1952).3Epstein, E., Agrochim., 6, 293 (1962).4Epstein, E., Plant Physiol., 36, 437 (1961).5 Epstein, E., W. E. Schmid, and D. W. Rains, Plant and Cell Physiology, in press.6 Laties, G. G., Ann. Rev. Plant Physiol., 10, 87 (1959).7Epstein, E., D. W. Rains, and W. E. Schmid, Science, 136, 1051 (1962).8 Leggett, J. E., and E. Epstein, Plant Physiol., 31, 222 (1956).9 Jackson, P. C., and H. R. Adams, J. Gen. Physiol., 46, 369 (1963).

10 Jacobson, L., and L. Ordin, Plant Physiol., 29, 70 (1954).11 Hoagland, D. R., in Permeability and the Nature of Cell Membranes, Cold Spring Harbor Sym-

posia on Quantitative Biology, vol. 8 (1940), p. 181.12 Robertson, R. N., Biol. Rev., 35, 231 (1960).13 Dainty, J., Ann. Rev. Plant Physiol., 13, 379 (1962).14 Briggs, G. E., A. B. Hope, and R. N. Robertson, Electrolytes and Plant Cells (Oxford: Black-

well Scientific Publications, 1961), p. 146.15 Fried, M., and J. C. Noggle, Plant Physiol., 33, 139 (1958).16 Bange, G. G. J., Plant and Soil, 11, 17 (1959).17 Bange, G. G. J., and R. Overstreet, Plant Physiol., 35, 605 (1960).18 Stein, W. D., Biochim. Biophys. Acta, 59, 66 (1962).19 Williams, D. E., Plant and Soil, 15, 387 (1961).

SYNTHESIS OF TRANSFER RNA BY ISOLATED NUCLEI*

BY MARGARET I. H. CHIPCHASE AND MAX L. BIRNSTIEL

DIVISION OF BIOLOGY, CALIFORNIA INSTITUTE OF TECHNOLOGY

Communicated by James Bonner, March 15, 1963

In the course of earlier experiments on the incorporation of labeled nucleosidesinto RNA by isolated nuclei1 we observed that much of the newly synthesized RNAis soluble in 1 M NaCl, as is transfer RNA.'-4 Sirlin has reported the incorporationof pseudo-uridine into nuclei, allegedly into transfer RNA,8 6 and the presenceof amino-acyl RNA in thymus nuclei has been shown by Hopkins.7 Since the peanuclei with which we work are capable of protein synthesis,8 they might therefore besuspected of containing transfer RNA. It will be shown below that isolated peanuclei not only contain, but possess the ability to synthesize, transfer RNA.

Materials and Methods.-Analytical reagent grade chemicals were used throughout. ATP,CTP, GTP, UTP, UMP, uridine, phosphocreatine, and crystalline DNAse were obtained fromSigma. Creatine phosphokinase was obtained from the California Corp. for Biochemical Re-search. Sodium penicillin-G was a gift of Chas. Pfizer and Co., New York. 2-hydroxy-3-naphthoic

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