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Effect of rat hypophysectomy and growth hormone treatment on cardiac polysomes and ribonucleic acid
.\I<(‘HIVER
OF
Effect
BIOCHEMISTRY
AND
115, 44s3-449 (1966)
BIOPHYSICS
of Rat Hypophysectomy on Cardiac
and
Polysomes
and
D. C. IV. EARL’ Department
of Biochemistry,
AND
University
Received
Growth
Ribonucleic
Treatment
Acid
A. KORKER
of Cambridge,
February
Hormone
Cambridge
England
9, 19GG
Hypophysectomy of the rat lowered the ribosome cont,ent of rat, heart but not the soluble RNA content. Treatment of rats with growth hormone stimulated the syn-
thesis of ribosomal structural RNA and, to a much lesser extent, of sRNA. Hypophysectomy lowered, and growth hormone treatment of rats restored, the incorporation of perfused amino acids into cardiac protein. Hypophysectomy did not affect the specific activity of the amino acid pool. Incorporation of amino acids into protein in vitro was depressed by hypophysectomy, yet the proportion of polysomes remained unaltered. Ribosomes were less able, after hypophysectomy, to react with poly U and stimulate phenylalanine incorporation.
There is substantial evidence in favor of the view that growth hormone can act to increase the rate of protein biosynthesis (1). Treatment of rats with growth hormone results in a marked stimulation of RKB synthesis of liver (2, 3), increases the number of ribosomes in the liver cells, stimulates their ability t,o incorporate amino acids into protein in vitro, and, simultaneously, increases the proportion of polysomes in the total ribonucleoprotein part’icle population (2). The preparation of active ribosomes and polysomes from rat heart (4) has opened t,he way to a study of the effects of growth hormone on the protein synthetic ability of this tissue. It is known (5) that the heart loses proportionally more weight than the body as a whole after hypophysectomy and that admir&ration of growth hormone to the hypophysectomized rat, increases the weight of the heart and its prot,ein content (6, 7, S). MATERIBLS
AND
METHODS
MuteriaZs. Human growth hormone, prepared by the method of Raben (9), was used. It was dissolved in 0.001 N NaOII, adjusted to neutrality with dilut,e HCl, and administered by subcutane1 Present address: Vanderbilt University,
Department Nashville,
of Physiology; Tennessee. 445
ous injection. The source of other materials has been described (10). ~1nin~aZ.s. Female albino rats weighing 150-200 gm were used. They were bred in the laboratory or were supplied by Tucks Ltd., Rayleigh, Essex. IIypophysectomy was carried out, under ether anesthesia, by the parapharyugeal method (11). The successful removal of t,he hypophysis was checked by manifestations such as loss of body weight., soft fur, and atrophied adrenals and ovaries. Jlethods. All the methods used have been described (10, 4). RESULTS
Table I shows the effect of hypophysectomy of the rat and growth hormone administration to the hypophysectomized animal on t#he incorporation of injected 3H-orotic acid and of perfused 14C-amino acids into RSA and protein, respectively. It is typical of rnariy similar experiments. It is notable that hypophysectomy results in a fall of incorporation into both RNA and protein, while the administration of growth hormone can reverse this inhibition. Ko effect of hypophysectomy was observed on the specific activity of the amino acid pool. It is worth mentioning that, with the same met,hod, a decrease in the specific
446
EARL
AND
activity of t,he pool was noted aft,er actinomycin treatment of rats (10). Labelling of RNA of caxk’ac ribosome preparations after hypoph,ysectouq and growth hormone. One hour after administration of 321,to rats by injection, most of the labellcd RKA exkacted f:rom cardiac ribosomes sediment’s at 28 S on sucrose-densit’y gradient centrifugation (10). Two hours after the pulse is given, radioactive RXA is present in all t,he t’ypes of RKA on the gradient: sRNA, 19 S and 28 S structural RSA of ribosomes, and R?CA sedimenting between 4 and 19 S. The specific activit’y of these fractions was measured after extraction and centrifugation analysis of RNA from cardiac ribosomes of normal, hypophysectomized and growth-hormone-treated rats. Typical results are shown in Table II. It is clear that hypophysect’omy has resulted in a lower incorporation of precursor into all types of RXA while a single dose of growth hormone stimulated incorporation into all t,ypes of RNA. Less stimulation of labelling of sRSA by growth hormone compared with t,hat of ribosomal RNA was noted. It was also noted t’hat the yield of ribosomal RNA from the heart muscle was consistently lower from hearts of hypophyseci,omized animals than from normal animals. For example, the yield from normal hearts was 23-28 mg ribosomal RNA/100 gm wet weight of heart muscle, while from hypophysectomized animals yields of 15-18 mg mere obtained. The yields of soluble RNA from the cell sap were similar from hearts of both normal and hypophysectomized nnimals.
KOILNEI:
Figure 1 show-s sucrose density gradient analyses of ribosome preparations from normal and hypophysectomized rat,s. Lit,t.le difference in t,he sedimentation patt,ern can be seen, either immediately aft,er hypophysectomy or S weeks after the operation. Table III shows t,he calculated data on the proportion of polysomes in preparations from normal and hypophysectomized rat)s and after growth hormone treatment; no significant difference was detectjed. This suggests that, the mRXA content, of such preparations is not, changed with respect to the amount’ of ribosomes. Similar observations were made in hearts from alloxan-diabetic rats, alloxan-diabetic treated with insulin, and normal insulinized rats. In these conditions also, while the rate of proteinsynthesis TABLE
HYPOPHYSECTOX~
Normal Hypophysectomiaed Hypophysectomized treated with growth hormoneh
sRNA Ribosomal
RNA 2% 19s 10s
800 172 180 800
550 436 GGO
320 244 325
II
AND GROWTH HORMONE PHOSPHATE INTO HEART
Normal rats
Incorporation into Protein (CPdW total protein)
a3H-Orot,ic acid (1 mC) was injected 1 hour before killing. b Growth hormone (500 rg/day) was given for 4 consecutive days.
TREATMEN.T ON THE MUSCLE RNAa Incorporation
RNA Species
Incorporation into RNA (crm/w total RNA)
Condition of animal
TABLE
EFFECTS OF
I
EFFECT OF HYPOPHYSECTOMY AND GROWTH HORMONE TREATMENT ON THE INCORPORATION OF 3H-OROTIC ACID INTO IZNA IX VIVO AND OF I$-LABELED AMINO ACIDSINTO PROTEIN DURING PERFUSIONS
Hypwfrh~to-
484 92 118 470
INCORPORATION
(cpm/mg RNA)
OF 32P-
Hypophys. as percentage of normal
Hypophysectomized + growth hormone”
.__ Percentage stimulation by growth hormone
GO 54 64 59
1050 410 620 1550
220 450 540 390
0 2 mC a-p-phosphate was given by intravenous injection 2 hours before killing. 6 500 pg growth hormone was administered 12 hours beforehand.
I:AT
E260mp O-5
A.
447
HYPOPHYSIZTORI~-
SOS
E260mp
A
B.
E260mp
80s
805
C.
A
0
IO
20
30
SAMPLE
0
NUMBER
IO FROM
BOTTOM
FIG. 1. Sucrose density-gradient centrifugation (A) normal rats; (B) 3 days after hypophysect,omy; tion is from right to left,. TABLE
III
PROPORTION OF POLYSOMES IN MIBOSOME PREPARATIONS FROM HEaRTS OF NORMAL HYPOPHYSECTOMIZED ANIMALS AND HYPOPHYSECTOMIZED I~ATS TREATED WITH GROWTH HORMONE' Animal Normal Hypophysectomized (12 weeks) Hypophysectomized (12 weeks) + growth hormone Normal + growth hormone Hypophysectomized (2 days) Hypophysectomized (2 weeks) Sham operated
GO-G9 65-70
GO-G2 WG5 FR G2 64
a 1 mg of growth hormone was given subcutaneously 12 hours before killing. The proportion of polysomes is expressed as a percentage of tot,al extinction of plots of extinction at 200 mfi after sucrose density gradient analysis of ribosome preparat,ions.
in the whole tissue has changed, the mRX\‘,Lz ribosome ratio has remained constant. Studies m the incorporation
20
of amino acids
info protein in vitro. In all experiments reported in this section, optimal conditions for the incorporation of amino acids in vitro were assumed t,o be similar to those elucidat,ed for czardiac ribosome preparation from
30 OF
0
IO
20
30
GRADIENT
analysis of ribosome and (C) 8 weeks after
preparations from hearts of hypophysectomy. Sedimenta-
hearts of normal rats (4). Results are expressed as cpm/mg of ribosomal RNA incubat,ed to relate the incorporation by different particle preparations. Table IV shows the incorporation in vitro, by cardiac ribosome preparations from heart’s of normal, hypophysectomized, and hypophysectomized animals treated with growth hormone. The growth hormone treatment used resulted in an increased body weight. Hypophysectomy reduced the amino acid incorporating activity Gz vitro. Treatment of t.hc hypophysectomized rats with 500 pg per day of growth hormone for 4 consecutive days failed to improve the incorporation in v&o, but treatment with growth hormone for 10 days effected it11 improvement,. Thus ribosome preparations from the hearts of hypophysectomized animals have been shown t,o contain a normal proportion of polysomes, yet their ability to incorporate amino acids into protein in vitro is markedly reduced. A similar disnbilit’y was noted after act,inomycin t,reatment of rats (10). The results in Table IV also show the effect on phcnylalanine incorporation of the additions of Poly U to in vitro systems containing cardiac ribosomes from normal, hypophysectomized, and growth-hormone
448
EARL AND KORNER TABLE IV
IN VITRO AMINO
ACID INCORPORATING ACTIVITY HYPOPHYSECTOMIZED, AND
OF RIBOSOME PREPARATIONS GROWTH-HORMONE-TREATED
FROM HEARTS RATS”
Hypophysectomized WYPOX.)
IIypox + G.H. 0.5 muday for 4 days
-.
Leu Phe Phe + Poly U
8120 5935 9380 (160)
4850 3075 4885 (150)
OF NORMAL,
--~___
52 52 52
4495 3155 4435 (140)
a L-(G-l%) Leucine (222mC/mmole) was used. Incubation, I hour at 37”. Incorporation expressedas cpm/mg ribosomal RNA incubated. Figures in parent’heses are percentage stimulation by poly IT. treated hypophysectomized rats. Although Poly U stimulates the incorporation of phenylalanine by ribosomes from hypophysectomized rats to an extent similar to that seen with normal ribosomes, the overall incorporation is still inhibited and to the extent observed in the absence of polyuridylic acid. This implies that after hypophysectomy, the ribosomes themselves are impaired in their ability to incorporate amino acids in vitro, and supports the contention that this inactivity is not solely due to a lack of mRNA. A similar finding was made for liver ribosomes (12). It is worth noting that cell sap prepared from hearts of hypophysectomized rats showed the same activity as normal sap when assayed in vitro with normal ribosomes. Both saps caused optimal incorporation at the same proportions of sap protein to ribosomal protein (see Ref. 4). DISCUSSION Growth hormone treatment of rats stimulates the labelling in the heart of both sizes of st,ructural RNA of ribosomes, and, to a lesser extent, of sRNA and also of material sedimenting between 4 and 19 S. These results are similar to those obtained with rat liver (2). In heart muscle (as in liver) the ribosome content falls after hypophysectomy, but the sRNA content probably remains almost unchanged. Despite the fall in ribosome content after hypophysectomy, the proportion of polysomes remains virtually unaltered. This is contrary to the finding with liver (a), but the analyses of liver were carried out on the 10,OOOgsupernatant material without prior
sedimentation of the ribosome fraction. Despite the presence of similar proportions of polysomes in preparations from normal and hypophysectomized rats, the ability of the particles to incorporate amino acids into protein in vitro and during perfusion was markedly altered. Clearly no simple correlation between polysome content and incorporating ability can be drawn. Furthermore, although the percentage stimulation of phenylalanine incorporation on addition to poly U was about the same with cardiac ribosomes from normal and hypophysectomized rats, the absolut’e values of incorporation mere lower wit,h ribosomes from rats deprived of growth hormone. Clearly the disability of these ribosomcs is not confined t’o lack of mRSA but must lie in some change in the ribosomcs thcmselvcs. Similar conclusions were reached wit’h regard to the effect of hypophysect’omy on liver ribosomes (1, 12), the effect of actinomycin treatment on cardiac ribosomes (lo), and the effect of alloxan diabetes on muscle ribosomes (13). ACKNOWLEDGMENTS The authors are grateful to the Medical Research Council, The Royal Society, and the British Empire Cancer Campaign for grants toward the cost of this work, and one of (D.C.N.E.) to the D.S.I.R. for a research studentship. We thank Professor F. G. Young for his encouragement. REFERENCES 1. KORNER, A., Recent Progr. Hormone Res. 21, 205 (1951). 2. KORXER, A., Hiochem. J. 92, 449 (1964). 3. TALWAR, G. P., PANDA, N. C., SARIN, G. S., AND TOLANI, 9. J., Biochenr. J. 82, Ii3 (1962).
K4T
HYPOPHYSECTOMY
4. EARL,D.C.N.,ANDKORNER, A., Kiochem.J. 94, 721 (1965). 5. BEZNAK, M., J. Physiol. 116, 74 (1952). 6. BEZNAK, M., J. Physiol. 120, 23P (1953). 7. BEZNAK, M., J. Physiol. 124, 64 (1954). 8. SIMPSON, M.E., EVANS, H.M., AND L1,C.N. Growth 13, 151 (1949). 9. RABEN, hl. S., Recent Progr. Hormone Res., 15,71 (1959).
449
10. EARL, D.C. N., AND KORIXER, A., drch. Biothem. Riophys. 115, 437 (1966). 11. ENGLE, D. J., AXD GRIFFITH, J. Q., in “The Rat in Laboratory Investigation” (J. Q. Griffith and E. J. Farris, eds.), p. 380. Lippincott, Philadelphia (1942). 12. KORNER, A., J. Cellular Camp. Physiol. 66, Suppl. 1, 153 (1965). 13. B.4MERSAD, O., AND WOOL, I. G., Federation Proc. 24, 511 (1965).
OF
Effect
BIOCHEMISTRY
AND
115, 44s3-449 (1966)
BIOPHYSICS
of Rat Hypophysectomy on Cardiac
and
Polysomes
and
D. C. IV. EARL’ Department
of Biochemistry,
AND
University
Received
Growth
Ribonucleic
Treatment
Acid
A. KORKER
of Cambridge,
February
Hormone
Cambridge
England
9, 19GG
Hypophysectomy of the rat lowered the ribosome cont,ent of rat, heart but not the soluble RNA content. Treatment of rats with growth hormone stimulated the syn-
thesis of ribosomal structural RNA and, to a much lesser extent, of sRNA. Hypophysectomy lowered, and growth hormone treatment of rats restored, the incorporation of perfused amino acids into cardiac protein. Hypophysectomy did not affect the specific activity of the amino acid pool. Incorporation of amino acids into protein in vitro was depressed by hypophysectomy, yet the proportion of polysomes remained unaltered. Ribosomes were less able, after hypophysectomy, to react with poly U and stimulate phenylalanine incorporation.
There is substantial evidence in favor of the view that growth hormone can act to increase the rate of protein biosynthesis (1). Treatment of rats with growth hormone results in a marked stimulation of RKB synthesis of liver (2, 3), increases the number of ribosomes in the liver cells, stimulates their ability t,o incorporate amino acids into protein in vitro, and, simultaneously, increases the proportion of polysomes in the total ribonucleoprotein part’icle population (2). The preparation of active ribosomes and polysomes from rat heart (4) has opened t,he way to a study of the effects of growth hormone on the protein synthetic ability of this tissue. It is known (5) that the heart loses proportionally more weight than the body as a whole after hypophysectomy and that admir&ration of growth hormone to the hypophysectomized rat, increases the weight of the heart and its prot,ein content (6, 7, S). MATERIBLS
AND
METHODS
MuteriaZs. Human growth hormone, prepared by the method of Raben (9), was used. It was dissolved in 0.001 N NaOII, adjusted to neutrality with dilut,e HCl, and administered by subcutane1 Present address: Vanderbilt University,
Department Nashville,
of Physiology; Tennessee. 445
ous injection. The source of other materials has been described (10). ~1nin~aZ.s. Female albino rats weighing 150-200 gm were used. They were bred in the laboratory or were supplied by Tucks Ltd., Rayleigh, Essex. IIypophysectomy was carried out, under ether anesthesia, by the parapharyugeal method (11). The successful removal of t,he hypophysis was checked by manifestations such as loss of body weight., soft fur, and atrophied adrenals and ovaries. Jlethods. All the methods used have been described (10, 4). RESULTS
Table I shows the effect of hypophysectomy of the rat and growth hormone administration to the hypophysectomized animal on t#he incorporation of injected 3H-orotic acid and of perfused 14C-amino acids into RSA and protein, respectively. It is typical of rnariy similar experiments. It is notable that hypophysectomy results in a fall of incorporation into both RNA and protein, while the administration of growth hormone can reverse this inhibition. Ko effect of hypophysectomy was observed on the specific activity of the amino acid pool. It is worth mentioning that, with the same met,hod, a decrease in the specific
446
EARL
AND
activity of t,he pool was noted aft,er actinomycin treatment of rats (10). Labelling of RNA of caxk’ac ribosome preparations after hypoph,ysectouq and growth hormone. One hour after administration of 321,to rats by injection, most of the labellcd RKA exkacted f:rom cardiac ribosomes sediment’s at 28 S on sucrose-densit’y gradient centrifugation (10). Two hours after the pulse is given, radioactive RXA is present in all t,he t’ypes of RKA on the gradient: sRNA, 19 S and 28 S structural RSA of ribosomes, and R?CA sedimenting between 4 and 19 S. The specific activit’y of these fractions was measured after extraction and centrifugation analysis of RNA from cardiac ribosomes of normal, hypophysectomized and growth-hormone-treated rats. Typical results are shown in Table II. It is clear that hypophysect’omy has resulted in a lower incorporation of precursor into all types of RXA while a single dose of growth hormone stimulated incorporation into all t,ypes of RNA. Less stimulation of labelling of sRSA by growth hormone compared with t,hat of ribosomal RNA was noted. It was also noted t’hat the yield of ribosomal RNA from the heart muscle was consistently lower from hearts of hypophyseci,omized animals than from normal animals. For example, the yield from normal hearts was 23-28 mg ribosomal RNA/100 gm wet weight of heart muscle, while from hypophysectomized animals yields of 15-18 mg mere obtained. The yields of soluble RNA from the cell sap were similar from hearts of both normal and hypophysectomized nnimals.
KOILNEI:
Figure 1 show-s sucrose density gradient analyses of ribosome preparations from normal and hypophysectomized rat,s. Lit,t.le difference in t,he sedimentation patt,ern can be seen, either immediately aft,er hypophysectomy or S weeks after the operation. Table III shows t,he calculated data on the proportion of polysomes in preparations from normal and hypophysectomized rat)s and after growth hormone treatment; no significant difference was detectjed. This suggests that, the mRXA content, of such preparations is not, changed with respect to the amount’ of ribosomes. Similar observations were made in hearts from alloxan-diabetic rats, alloxan-diabetic treated with insulin, and normal insulinized rats. In these conditions also, while the rate of proteinsynthesis TABLE
HYPOPHYSECTOX~
Normal Hypophysectomiaed Hypophysectomized treated with growth hormoneh
sRNA Ribosomal
RNA 2% 19s 10s
800 172 180 800
550 436 GGO
320 244 325
II
AND GROWTH HORMONE PHOSPHATE INTO HEART
Normal rats
Incorporation into Protein (CPdW total protein)
a3H-Orot,ic acid (1 mC) was injected 1 hour before killing. b Growth hormone (500 rg/day) was given for 4 consecutive days.
TREATMEN.T ON THE MUSCLE RNAa Incorporation
RNA Species
Incorporation into RNA (crm/w total RNA)
Condition of animal
TABLE
EFFECTS OF
I
EFFECT OF HYPOPHYSECTOMY AND GROWTH HORMONE TREATMENT ON THE INCORPORATION OF 3H-OROTIC ACID INTO IZNA IX VIVO AND OF I$-LABELED AMINO ACIDSINTO PROTEIN DURING PERFUSIONS
Hypwfrh~to-
484 92 118 470
INCORPORATION
(cpm/mg RNA)
OF 32P-
Hypophys. as percentage of normal
Hypophysectomized + growth hormone”
.__ Percentage stimulation by growth hormone
GO 54 64 59
1050 410 620 1550
220 450 540 390
0 2 mC a-p-phosphate was given by intravenous injection 2 hours before killing. 6 500 pg growth hormone was administered 12 hours beforehand.
I:AT
E260mp O-5
A.
447
HYPOPHYSIZTORI~-
SOS
E260mp
A
B.
E260mp
80s
805
C.
A
0
IO
20
30
SAMPLE
0
NUMBER
IO FROM
BOTTOM
FIG. 1. Sucrose density-gradient centrifugation (A) normal rats; (B) 3 days after hypophysect,omy; tion is from right to left,. TABLE
III
PROPORTION OF POLYSOMES IN MIBOSOME PREPARATIONS FROM HEaRTS OF NORMAL HYPOPHYSECTOMIZED ANIMALS AND HYPOPHYSECTOMIZED I~ATS TREATED WITH GROWTH HORMONE' Animal Normal Hypophysectomized (12 weeks) Hypophysectomized (12 weeks) + growth hormone Normal + growth hormone Hypophysectomized (2 days) Hypophysectomized (2 weeks) Sham operated
GO-G9 65-70
GO-G2 WG5 FR G2 64
a 1 mg of growth hormone was given subcutaneously 12 hours before killing. The proportion of polysomes is expressed as a percentage of tot,al extinction of plots of extinction at 200 mfi after sucrose density gradient analysis of ribosome preparat,ions.
in the whole tissue has changed, the mRX\‘,Lz ribosome ratio has remained constant. Studies m the incorporation
20
of amino acids
info protein in vitro. In all experiments reported in this section, optimal conditions for the incorporation of amino acids in vitro were assumed t,o be similar to those elucidat,ed for czardiac ribosome preparation from
30 OF
0
IO
20
30
GRADIENT
analysis of ribosome and (C) 8 weeks after
preparations from hearts of hypophysectomy. Sedimenta-
hearts of normal rats (4). Results are expressed as cpm/mg of ribosomal RNA incubat,ed to relate the incorporation by different particle preparations. Table IV shows the incorporation in vitro, by cardiac ribosome preparations from heart’s of normal, hypophysectomized, and hypophysectomized animals treated with growth hormone. The growth hormone treatment used resulted in an increased body weight. Hypophysectomy reduced the amino acid incorporating activity Gz vitro. Treatment of t.hc hypophysectomized rats with 500 pg per day of growth hormone for 4 consecutive days failed to improve the incorporation in v&o, but treatment with growth hormone for 10 days effected it11 improvement,. Thus ribosome preparations from the hearts of hypophysectomized animals have been shown t,o contain a normal proportion of polysomes, yet their ability to incorporate amino acids into protein in vitro is markedly reduced. A similar disnbilit’y was noted after act,inomycin t,reatment of rats (10). The results in Table IV also show the effect on phcnylalanine incorporation of the additions of Poly U to in vitro systems containing cardiac ribosomes from normal, hypophysectomized, and growth-hormone
448
EARL AND KORNER TABLE IV
IN VITRO AMINO
ACID INCORPORATING ACTIVITY HYPOPHYSECTOMIZED, AND
OF RIBOSOME PREPARATIONS GROWTH-HORMONE-TREATED
FROM HEARTS RATS”
Hypophysectomized WYPOX.)
IIypox + G.H. 0.5 muday for 4 days
-.
Leu Phe Phe + Poly U
8120 5935 9380 (160)
4850 3075 4885 (150)
OF NORMAL,
--~___
52 52 52
4495 3155 4435 (140)
a L-(G-l%) Leucine (222mC/mmole) was used. Incubation, I hour at 37”. Incorporation expressedas cpm/mg ribosomal RNA incubated. Figures in parent’heses are percentage stimulation by poly IT. treated hypophysectomized rats. Although Poly U stimulates the incorporation of phenylalanine by ribosomes from hypophysectomized rats to an extent similar to that seen with normal ribosomes, the overall incorporation is still inhibited and to the extent observed in the absence of polyuridylic acid. This implies that after hypophysectomy, the ribosomes themselves are impaired in their ability to incorporate amino acids in vitro, and supports the contention that this inactivity is not solely due to a lack of mRNA. A similar finding was made for liver ribosomes (12). It is worth noting that cell sap prepared from hearts of hypophysectomized rats showed the same activity as normal sap when assayed in vitro with normal ribosomes. Both saps caused optimal incorporation at the same proportions of sap protein to ribosomal protein (see Ref. 4). DISCUSSION Growth hormone treatment of rats stimulates the labelling in the heart of both sizes of st,ructural RNA of ribosomes, and, to a lesser extent, of sRNA and also of material sedimenting between 4 and 19 S. These results are similar to those obtained with rat liver (2). In heart muscle (as in liver) the ribosome content falls after hypophysectomy, but the sRNA content probably remains almost unchanged. Despite the fall in ribosome content after hypophysectomy, the proportion of polysomes remains virtually unaltered. This is contrary to the finding with liver (a), but the analyses of liver were carried out on the 10,OOOgsupernatant material without prior
sedimentation of the ribosome fraction. Despite the presence of similar proportions of polysomes in preparations from normal and hypophysectomized rats, the ability of the particles to incorporate amino acids into protein in vitro and during perfusion was markedly altered. Clearly no simple correlation between polysome content and incorporating ability can be drawn. Furthermore, although the percentage stimulation of phenylalanine incorporation on addition to poly U was about the same with cardiac ribosomes from normal and hypophysectomized rats, the absolut’e values of incorporation mere lower wit,h ribosomes from rats deprived of growth hormone. Clearly the disability of these ribosomcs is not confined t’o lack of mRSA but must lie in some change in the ribosomcs thcmselvcs. Similar conclusions were reached wit’h regard to the effect of hypophysect’omy on liver ribosomes (1, 12), the effect of actinomycin treatment on cardiac ribosomes (lo), and the effect of alloxan diabetes on muscle ribosomes (13). ACKNOWLEDGMENTS The authors are grateful to the Medical Research Council, The Royal Society, and the British Empire Cancer Campaign for grants toward the cost of this work, and one of (D.C.N.E.) to the D.S.I.R. for a research studentship. We thank Professor F. G. Young for his encouragement. REFERENCES 1. KORNER, A., Recent Progr. Hormone Res. 21, 205 (1951). 2. KORXER, A., Hiochem. J. 92, 449 (1964). 3. TALWAR, G. P., PANDA, N. C., SARIN, G. S., AND TOLANI, 9. J., Biochenr. J. 82, Ii3 (1962).
K4T
HYPOPHYSECTOMY
4. EARL,D.C.N.,ANDKORNER, A., Kiochem.J. 94, 721 (1965). 5. BEZNAK, M., J. Physiol. 116, 74 (1952). 6. BEZNAK, M., J. Physiol. 120, 23P (1953). 7. BEZNAK, M., J. Physiol. 124, 64 (1954). 8. SIMPSON, M.E., EVANS, H.M., AND L1,C.N. Growth 13, 151 (1949). 9. RABEN, hl. S., Recent Progr. Hormone Res., 15,71 (1959).
449
10. EARL, D.C. N., AND KORIXER, A., drch. Biothem. Riophys. 115, 437 (1966). 11. ENGLE, D. J., AXD GRIFFITH, J. Q., in “The Rat in Laboratory Investigation” (J. Q. Griffith and E. J. Farris, eds.), p. 380. Lippincott, Philadelphia (1942). 12. KORNER, A., J. Cellular Camp. Physiol. 66, Suppl. 1, 153 (1965). 13. B.4MERSAD, O., AND WOOL, I. G., Federation Proc. 24, 511 (1965).