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Adamson, S. D., Herbert, E. & Godschaux, W. (1968) Arch. Biochem. Biophys. 125,671-683
Adamson, S. D., Howard, G. A. & Herbert, E. (1969) Cold Spring Harbor Symp. Quant. Biof.
34,547-554
Burgess, A. B. & Mach, B. (1971) Nature (London) New Biol. 233, 209-210
Darnbrough, C., Hunt, T. & Jackson, R. J. (1972) Biochem. Biophys. Res. Commun. 48,
1556-1564
Ehrenfeld, E. & Hunt, T. (1971) Proc. Nut. Acad. Sci. US.68, 1075-1078
Honjo, T., Nishizuka, Y . , Hayaishi, 0. & Kato, I. (1968) J. Biol. Chem. 243, 3553-3555
Housman, D., Jacobs-Lorena, M., RajBhandary, U. L. & Lodish, H. F. (1970) Nature (London)
227,913-918
Howard, G. A,, Adamson, S. D. & Herbert, E. (1970) J . Biol. Chem. 245, 6237-6239
Hunt, T., Vanderhoff, G . A. & London, I. M. (1972) J. Mol. Biol. 66, 471-481
Hunter, A. R. &Jackson, R. J. (1971) Eur. J. Biochern. 19, 316-322
Hunter, A. R., Hunt, T., Jackson, R. J. & Robertson, H. D. (1972) in Synthesis, Structure and
Funcriorz of Hemoglobin (Martin, H. & Nowicki, L., eds.), pp. 133-145, J. F. Lehmanns
Verlag, Munich
Jackson, R. J. & Hunter, A. R. (1970) Nature (London) 227, 672-676
Jacobs-Lorena, M. & Baglioni, C. (1972) Proc. Nut. Acad. Sci. US.69, 1425-1428
Kaempfer, R. (1969) Nature (London) 222, 950-953
Kosower, N. S., Vanderhoff, G . A., Benerofe, B. & Hunt, T. (1971) Biocheni. Biophys. Res.
Commun. 45,816-821
Kosower, N. S., Vanderhoff, G . A. & Kosower, E. M. (1972) Biochim. Biophys. Acta 272,
623-637
Legon, S., Jackson, R. J. &Hunt, T. (1973) Nature (London) New Biol. 241,150-152
Lengyel, P. (1969) Cold Spring Harbor Symp. Quant. Biof. 34, 828-841
Lucas-Lenard, J. & Lipmann, F. (1971) Annu. Rev. Biochem. 4 0 , 4 0 9 4 8
Mathews, M. B., Osborn, M. & Lingrel, J. B. (1971) Nature (London) New Biol. 233, 206-209
Maxwell, C. R. & Rabinovitz, M. (1969) Biochem. Biophys. Res. Commun. 35, 79-85
Olsen, G . D., Gaskill, P. & Kabat, D. (1972) Biochim. Biophys. Acta 272, 297-304
Schreier, M. & Staehelin, T. (1973~)J . Mol. Biof.73,329-349
Schreier, M. & Staehelin, T. (19736) Nature (London) in the press
Smith, A. E. & Marcker, K. A. (1970) Nature (London) 226, 607-610
Terada, M., Metafora, S., Banks, J., Dow, L. W., Bank, A. & Marks, P. A. (1972) Biochem.
Biophys. Res. Commun. 47, 766-774
Weeks, D. P., Verma, D. P. S., Seal, S. N. & Marcus, A. (1972) Nature (London)236, 167-168
Translational C o n t d of ImmunogPobuPin Synthesis
RONALD H. STEVENS
National Institute for Medical Research, 7 h e Ridgewuy, Mill Hill, London N W7 1A A ,
U.K.
Mouse plasmacytoma cells (myeloma 5563) when grown in tissue culture synthesize a
7 S immunoglobulin composed of two heavy chains (H-chains) and two light chains
(L-chains). The H-chains and L-chains are synthesized on separate polyribosomes
(Williamson & Askonas, 1967), assembled and transported into an intracellular pool
of myeloma protein (H2L2) before secretion from the cell. The intracellular pool of
H2L2 is large, representing over 2h of synthesis of H-chains and L-chains. A large
intracellular pool of free L-chains also exists, which on a molar basis is over four times
the size of the H2L2 pool. There is, however, only a small pool of free H-chains
(Williamson & Askonas, 1968).
The regulatory mechanisms controlling the intracellular concentrations of H-chains,
L-chains and H2L2 protein are poorly understood. Experiments have therefore been
conducted (in collaboration with Dr. A. R. Williamson) to investigate the regulation
of the intracellular concentration of H2L2.
When myeloma cells are incubated a t 25"C, secretion of H2L2from the cell ceases
within 20min. The synthesis of H-chains and L-chains and the subsequent assembly
into H2Lr continues, however, a t nearly 40% of the 37°C synthetic rate, resulting in a n
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BIOCHEMICAL SOCIETY TRANSACTIONS
increased intracellular concentration of HzL2. Myeloma cells that have been incubated
at 25°C for 90min contain a 20-30x-increased intracellular H2Lz pool. When such
cells are reincubated at 37°C there is an apparent decrease in H-chain synthesis relative
to L-chain or total protein synthesis. Whereas L-chain synthesis returns to the pre25°C synthetic rate within 15min, the synthesis of H-chain requires over 60min to return
to the pre-25°C rate.
A similar decrease in H-chain synthesis occurs during prolonged incubation at
25°C. Myeloma cells maintained in exponential growth contain a larger intracellular
pool of HzLz than do cells in late stationary phase. When both populations of cells
were incubated at 25°C and the synthesis of H-chains and L-chains was measured at
intervals after the lowering of the temperature, repression of H-chain synthesis was
observed. Exponentially growing cells showed an 80 % decrease in H-chain synthesis
after lOOmin at 25°C. Stationary cells, with the diminished intracellular concentration
of H2L2,required 2lOmin at 25°C to effect an equivalent decrease in H-chain synthesis,
the additional time being sufficient to increase the intracellular concentration of H2Lz
to that of exponentially growing cells.
From these experiments it appeared that an increased intracellular concentration of
HzL2 caused a decrease in H-chain synthesis. The effect of a diminished intracellular
H2Lz pool on subsequent synthesis of myeloma protein was also examined. Total
cellular protein synthesis was decreased by 98 % with cycloheximide. Synthesis of the
H-chains and L-chains ceases although secretion proceeds normally (Scharff et al.,
1967). After a 40% depletion of the H2L2pool, the cycloheximide was removed, the
cells were radioactively labelled at lOmin intervals and the synthesis of total protein
and HzLzwas measured. Between 20 and 40min after the release of the cells from protein
inhibition the ratio of H2L2to total protein synthesized increases from 6 to 13%. After
an additional 20min the ratio returns to 6%, the increased synthesis having been
sufficient to re-form the intracellular H2L2pool.
The effect of H-chain repression by HzLz can also be demonstrated in vitro. When
polyribosomes prepared from myeloma cells were injected with various concentrations
of HzLz into oocytes from Xenopus laevis there was a decrease in the-amount of
myeloma protein synthesized. This decrease was mainly due to an 80% decrease in
H-chain synthesis.
From the above experiments it is apparent that in uioo the synthesis of H-chains is
affected by the intracellular concentration of H2L2.The rapidity with which H-chain
synthesis is repressed and released from repression suggests that regulation is occurring
at the level of mRNA translation. It therefore appears likely that one mechanism
regulating the intracellular concentration of myeloma protein involves the feedback
repression of H2Lzon H-chain mRNA. The exact nature of the repression and whether
or not it is the sole means of regulating the intracellular HzLzpool have not yet been
elucidated.
Scharff, M., Shapiro, A. & Ginsberg, B. (1967) Cold Spring Harbor Synzp. Quaiit. Biol. 32,
237-241
Williamson, A. R. & Askonas, B. A. (1967) J. Mol. Biol. 23, 201-216
Williamson, A. R. & Askonas, B. A. (1968) Arch. Biochem. Biophys. 125, 401-409
Translation of Lens Messenger Ribonucleic Acid in Heterologous Systems
HANS BLOEMENDAL, ANTON J. M. BERNS and GER J. A. M. STROUS
Department of Biochemistry, University of Nijmegen, Geert Grooteplein N 21,
Nijinegen, The Netherlands
The mammalian eye lens represents a very convenient system for the study of a number
of fundamental biological processes, e.g. protein biosynthesis, aging and differentiation.
One of the major constituents of lens tissue is a-crystallin, a protein that is composed of
1973