Changes In Rate Of Synthesis Research Articles

Measurement of DNA integrity and structure in Xenopus embryos in the presence of hydroxyurea, actinomycin-D, and triethylenemelamine using the fluorescent probe Hoechst 33258.

Cell health assay of water quality (CHAWQ) is an assay using intracellular biomarkers measured by optical techniques. CHAWQ uses embryos of the South African clawed frog, Xenopus laevis, and optical transducers of intracellular biomarkers to obtain rapid assessment of toxicity to frog embryos. Since the biomarkers are common to all cells, CHAWQ can indicate toxicity of different classes of chemicals. Among the biomarkers used are 1) the change in synthesis rate, 2) the structure, or 3) the environment of DNA. Measurement of DNA to detect genotoxicants has previously used extracted DNA or flow cytometry to detect alterations in DNA content or configuration. We report the use of viable frog embryos and the fluorescent probe Hoechst 33258 to detect the effect of three DNA-active chemicals--actinomycin-D, hydroxyurea, and triethylenemelamine (TEM)--on DNA in intact embryos. We found that we can detect changes in the DNA in the presence of toxicants at concentrations comparable to longer-term assays but following a much shorter time of drug exposure. Actinomycin-D caused a fluorescence decrease, TEM caused a fluorescence increase, whereas hydroxyurea gave a biphasic response. Hydroxyurea caused a decrease at low concentrations and an increase at higher concentrations. Concentration-response data for TEM, hydroxyurea, and actinomycin-D generated EC50 values of 0.1 mg/ml, 1.4 mg/ml, and 6.34 micrograms/ml, respectively.

Variations in hepatic carbon flux during unrestricted feeding.

Previous findings have established a pivotal role for hepatic pyruvate dehydrogenase complex (PDH) in regulating hepatic carbon flux during the starved-to-fed and fed-to-starved nutritional transitions [Holness, McLennan, Palmer & Sugden (1988) Biochem. J. 252, 325-330; Holness & Sugden (1990) Biochem. J. 268, 77-81]. We have therefore examined liver PDH activities during the light and dark phases of the feeding cycle in the adult rat in relation to hepatic glycogenesis, fatty acid synthesis and cholesterogenesis. There was significant synchronous suppression of lipogenesis and glycogenesis during the light phase; rates were restored asynchronously during the dark (feeding) phase. Glycogen concentrations declined during the light phase and increased during the dark phase. Despite quite dramatic changes in rates of glycogen and lipid synthesis and hepatic glycogen concentrations during the light and dark phases, hepatic PDHa (active form) activity remained relatively unchanged. Qualitative and quantitative differences in the pattern of change in rates of synthesis of fatty acid and cholesterol suggested regulation at pathway-specific sites distal to PDH.

On the molecular mechanism of the circadian clock: The 64000-Mr protein of Chlamydomonas reinhardtii might be related to the biological clock

Labelling of Chlamydomonas reinhardtii cells with [(35)S] methionine led to the detection of a 64-kDa polypeptide which is synthesized according to a circadian rhythm. The change in synthesis rate could be demonstrated to exist under constant dim-light conditions as well as in darkness. Maximum synthesis of the 64-kDa polypeptide occurred at about 10 h after onset of constant conditions, and the period length of its oscillation was about 29 h. The 64-kDa polypeptide was synthesized on 80S ribosomes as shown by experiments in which cycloheximide and chloramphenicol were supplied to the cultures. Peptide-microsequence analysis yielded an N-terminal sequence of 14 amino acids. No significant homology to any other known polypeptide could be demonstrated in searches of current databases. The possible role of the 64-kDa polypeptide and its relationship to the biological clock is discussed.

Effect of interferon-gamma on complement gene expression in different cell types.

We have studied the expression of the complement components C2, C3, factor B, C1 inhibitor (C1-inh), C4-binding protein (C4-bp) and factor H in human peripheral blood monocytes, skin fibroblasts, umbilical vein endothelial cells (HUVEC) and the human hepatoma cell line G2 (Hep G2) in the absence and the presence of interferon-gamma (IFN-gamma). E.l.i.s.a. performed on culture fluids, run-on transcription assays, Northern blot and double-dilution dot-blot techniques confirmed that monocytes expressed all six components, whereas fibroblasts, HUVEC and HepG2 each expressed five of the six components. Fibroblasts and HUVEC did not synthesize C4-bp, and Hep G2 did not produce factor H. In addition to these differences, the synthesis rates of C3, C1-inh and factor H were not the same in all cell types. However, the synthesis rates of C2 and factor B were similar in all four cell types. The half-lives of the mRNAs were shorter in monocytes than in other cell types. Monocyte factor H mRNA had a half-life of 12 min in monocytes, compared with over 3 h in fibroblasts and HUVEC. The instability of factor H mRNA in monocytes may contribute to their low factor H secretion rate. IFN-gamma produced dose-dependent stimulation of C2, factor B, C1-inh, C4-bp and factor H synthesis by all cell types expressing these proteins, but decreased C3 synthesis in all four cell types. Cell-specific differences in the response to IFN-gamma were observed. The increased rates of transcription of the C1-inh and factor H genes in HUVEC were greater than in other cell types, while the increased rate of transcription of the C2, factor B and C1-inh genes in Hep G2 cells was less than in other cell types. IFN-gamma did not affect the stability of C3, factor H or C4 bp mRNAs, but increased the stability of factor B and C1-inh mRNAs and decreased the stability of C2 mRNA. Although these changes occurred in all four cell types studied, the half-life of C1-inh mRNA in monocytes was increased almost 4-fold, whereas the increases in the other cell types were less than 30%. These data show that the constitutive synthesis rates of complement components may vary in the different cell types. They also show that the degree of change in synthesis rates in response to IFN-gamma in each of the cell types often varies due to differences in transcriptional response, sometimes in association with changes in mRNA stability.

Ferritin gene expression in health and malignancy.

Intracellular iron can be stored in the protein shell of ferritin to protect the cell against the toxic action of the iron. In response to increased cell iron, more ferritin subunits are synthesized using translational and transcriptional mechanisms. Translational control involves a unique stem-loop structure in the 5' untranslated region of the subunit messengers. When iron level is low, a protein binds to this stem-loop structure and prevents translation. When intracellular iron level rises, the repressor protein is discharged and the large population of messengers begins to translate subunits. Similar stem-loop motifs occur in the 3' untranslated region of the transferrin receptor messenger where they regulate breakdown of the receptor mRNA. Finally, the presence of excess iron preferentially stimulates transcription of more ferritin message of one type (L-mRNA) which produces ferritin shells favoring iron storage. In this way, protection of the cell against iron excess is enhanced by coordinate changes in rate of synthesis of ferritin mRNA of the L-type, by release of ferritin mRNA stored in the cytoplasm, and by a reduction in the number of receptors for accepting iron into the cell. The application of these principles with reference to malignant cells is discussed.

Changes in gene expression patterns during the sexual life cycle of Chlamydomonas reinhardtii

In the sexual life cycle of Chlamydomonas reinhardtii, three types of cells can be distinguished: vegetative cells, gametes, and zygotes. Using is vivo pulse labelling of proteins with 35S-sulfate, followed by two-dimensional separation of the polypeptides on polyacrylamide gels, we analyzed the patterns of protein synthesis typical for each cell type. Approximately 20% of the proteins detected were synthesized at very different rates when the pattern from vegetative cells was compared to that of gametes or zygotes. Gamete formation from vegetative cells is a two-step process controlled by two extrinsic signals: first, nitrogen-starvation induces the differentiation of vegetative cells to pregaroetes, which are then competent for a light–induced differentiation to mature gametes. The majority of the changes in rate of synthesis of different proteins during the switch from vegetative cells to gametes was observed already in pregametes. Between the stages of pregametes and gametes, changes in the rates of synthesis of four proteins were detected. Induction of zygote germination resulted in complex changes in the patterns of gene expression. Evidence is presented for three groups of proteins whose synthesis is turned on or turned off during zygote germination.

Changes in rates of synthesis of individual proteins in a psychrophilic bacterium after a shift in temperature

In the psychrophilic bacterium Vibrio sp. strain ANT-300, which has the ability to grow efficiently between 13 and -2 degrees C, with an optimum at 7 degrees C, cells in steady-state growth at 0 and 13 degrees C appeared to exhibit different patterns in the levels of certain individual proteins. With a shift in temperature, the steady-state level of individual proteins was achieved only after dramatic transient changes in the rates of synthesis of a small number of those proteins whose levels would be adjusted. Upon a shift up from 0 to 13 degrees C, the rates of synthesis of at least 25 proteins increased transiently, while increased rates of synthesis of 39 proteins were induced immediately upon a shift down from 13 to 0 degrees C. The proteins of which the levels would be adjusted were synthesized at differential rates, which varied conspicuously with respect to timing after the shifts in temperature. Such changes appear to be active regulatory responses to changes in temperature.

Differences between the regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase and low density lipoprotein receptor in human hepatoma cells and fibroblasts reside primarily at the translational and post-translational levels.

We have carried out parallel analyses of the regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGr) and low density lipoprotein receptor (LDLr) in two highly differentiated human hepatoma cell lines, HepG2 and Hep3B, and primary cultures of human fibroblasts. Analyses of the levels of HMGr and LDLr mRNAs under a variety of culture conditions that perturb intracellular sterol metabolism, or which differ in the levels of extracellular sterols, indicated that the hepatoma cells and fibroblasts responded similarly in terms of the repression or induction ratios of both mRNAs. However, the absolute levels of the mRNAs were severalfold higher in the hepatoma cells. The major difference between the responses of the hepatoma cells and fibroblasts involved the increase in expression of LDLr which occurred upon shifting the cells from complete to lipoprotein-depleted serum. Under these conditions, the 3-fold increase in rate of synthesis of LDLr in the hepatomas was closely matched by increases in the level of its mRNA. In the case of fibroblasts, a 10-fold increase in translational efficiency was required to explain the 30-fold change in rate of synthesis of LDLr. Polysome profiles from both hepatoma cells and fibroblasts suggest that the rate of elongation or termination on LDLr mRNA is relatively low in the presence of reconstituted complete serum, and that it increases in fibroblasts upon lipoprotein depletion, but not in the hepatoma cells. These data indicate that hepatic expression of LDLr may be relatively refractory to induction by decreased circulating levels of lipoprotein when compared with peripheral tissues.

Quantitative analysis of protein synthesis in mouse embryos. I. Extensive reprogramming at the one- and two-cell stages

A quantitative, high-resolution, two-dimensional gel protein database has been constructed for the mouse embryo. This database has been used to obtain a detailed accounting of the amount and relative timing of changes in protein synthesis that occur during the 1-, 2-, and 4-cell stages along with a description of the most prevalent developmentally regulated patterns of synthesis. We find that during these early stages of development the pattern of proteins synthesized changes to a much greater extent than previously appreciated. During the 1- and 2-cell stages, the majority (60% and 85%, respectively) of the analyzed proteins exhibit twofold or greater changes in their rates of synthesis. The periods of greatest change are the late 1-cell and mid 2-cell stages, during which an extensive remodelling of protein synthetic pattern occurs that is largely complete by 15 h following the first cleavage. Once this reprogramming is complete, very little change is observed during the late 2-cell and 4-cell stages. Cluster analysis of individual protein synthesis patterns reveals a limited number of coordinately regulated protein sets that are responsible for most of the changes observed during the 1- and 2-cell stages. During the 2-cell stage, one third of the proteins increase by an average of fivefold, another third decrease by an average of sevenfold, and 10% undergo transient changes in rates of synthesis. These patterns reflect the switch from zygotic to maternal mRNA utilization following transcriptional activation.(ABSTRACT TRUNCATED AT 250 WORDS)

Starvation Effects on Protein Synthesis in Red and White Muscle of the Barred Sand Bass,Paralabrax nebulifer

Starvation of the fish Paralabrax nebulifer was accompanied by complex changes in the synthesis rates of different classes of proteins within the white muscle. Synthesis of white-muscle myofibrillar proteins, as measured by incorporation of 14C-leucine, decreased rapidly during starvation, whereas synthesis of total sarcoplasmic proteins was unchanged. Although actin and aldolase label incorporation in white muscle underwent an immediate decrease to approximately 10% of control, lactate dehydrogenase and creatine phosphokinase showed a smaller initial decrease. Relative activities of white-muscle glycolytic enzymes were conserved despite a significant loss in overall glycolytic capacity. Control of specific synthesis rates may be important in maintaining an overall balance of proteins with different half-lives. In red muscle, starvation led to no reductions in the synthesis of any protein examined. Changes in muscle protein synthesis rates were reflected in a larger decrease in RNA/DNA ratio in white muscle than in red muscle during starvation. These results suggest that balanced metabolic function is retained during starvation in fishes, and that starvation may have differential effects on high- and low-speed swimming abilities.

Selective activation of the N-glycosylation apparatus in uteri by estrogen.

Estrogen rapidly, preferentially and markedly enhances the rate of N-linked glycoprotein synthesis in mouse uteri. In contrast, the rate of glycoprotein turnover is unaffected by the hormone. Estrogen's effect on the expression of mRNA coding for glycoproteins was studied using an in vitro translation-glycosylation system as well as by Northern/slot blot analyses. Both approaches indicated that estrogen did not have a preferential stimulatory effect on the general expression of glycoprotein mRNA. Neither was there a significant change in the relative levels of specific mRNA coding for several N-linked glycoproteins, i.e. laminin B1 and B2, fibronectin, and uvomorulin, as a function of estrogen treatment. Immunoprecipitation studies also demonstrated no change in the relative rates of synthesis of the corresponding core proteins for laminin or fibronectin. Taken together, these results suggested that estrogen primarily stimulated glycoprotein synthesis by stimulating the glycosylation apparatus, and not by increasing synthesis of protein acceptors. Previous studies have indicated that of a variety of potential regulatory points in the pathway of N-linked glycoprotein assembly, only expression of mannosylphosphoryldolichol synthase (MPDS) increases sufficiently to account for the increase in glycoprotein expression observed in response to estrogen. Consistent with these observations, it was found that injection of uterine poly(A+) RNA from estrogen-treated uteri into Xenopus oocytes markedly stimulated MPDS activity in the oocytes. In contrast, injection of RNA from non-estrogen-treated uteri did not stimulate MPDS activity in oocytes. Collectively, these results indicate that steroid hormones can modulate glycoprotein expression by preferentially stimulating the glycosylation apparatus. Nonetheless, one of estrogen's effects on the glycosylation apparatus, induction of MPDS activity, appears to occur at a transcriptional level.

Developmental and diurnal changes in ecdysteroid biosynthesis by prothoracic glands of Rhodnius prolixus (Hemiptera) in vitro during the last larval instar

The synthesis of ecdysteroids by prothoracic glands (PGs) of male last instar larvae of Rhodnius prolixus was measured in vitro by radioimmunoassay throughout the course of larval-adult development. Large and systematic changes in relative rates of synthesis occur during development. Two bursts of elevated synthetic activity were found. The first commences as soon as development is initiated by a blood meal and lasts approximately 1 day. The second commences 4 days later and increases progressively to a peak at Days 11-13 after feeding (up to 25 ng of 20-hydroxyecdysone eq. gland-1/4 hr-1). The onset of each of these bursts of activity coincides with apparent times of PG stimulation in vivo by release of the prothoracicotropic hormone from the brain. Both bursts result in increases in hemolymph ecdysteroid titer measured in the donor animals. PGs exhibit an abrupt attenuation of synthesis on Day 14, which is followed by a rapid decline in the hemolymph ecdysteroid titer. Clearly, ecdysteroid synthesis by PGs is a major factor regulating the hemolymph titer. Ecdysteroid synthesis by PGs exhibits diurnal changes in vitro. The amount of ecdysteroid synthesized by PGs from animals during the scotophase is two to five times higher than that from animals during the photophase. A corresponding rhythm is seen in the hemolymph ecdysteroid titer. The rhythm in the titer is known to be under circadian control. It is therefore suggested that ecdysteroid synthesis in PGs of Rhodnius is regulated by a circadian system, possibly located in the PGs themselves.

Interferon β2/B‐Cell Stimulating Factor 2/Interleukin 6 Affects Glycosylation of Acute Phase Proteins in Human Hepatoma Cell Lines

Undefined monocyte-derived cytokines have previously been shown to affect glycan processing in glycoproteins secreted by human hepatoma cell lines. Hep 3B cells, when incubated with the cytokine interferon beta 2/B-cell stimulating factor 2/interleukin 6, secreted forms of alpha 1-protease inhibitor, ceruloplasmin, and alpha-fetoprotein with increased reactivity with concanavalin A (Con A) while incubation of Hep G2 cells with this cytokine led to secretion of forms of these proteins with decreased reactivity with Con A, reflecting changes in their oligosaccharide chains. The difference in response of these two transformed cell lines to this cytokine undoubtedly reflects differences in their intracellular glycan processing mechanisms. Changes in glycosylation patterns were dissociated from changes in rate of synthesis: this cytokine caused increased synthesis of alpha 1-protease inhibitor and ceruloplasmin, and decreased synthesis of alpha-fetoprotein in both cell lines.

Stimulus-evoked changes in RNA translation in cholinergic neurons

The electromotor system of Torpedo marmorata was used to investigate effects of evoked neuronal activity on protein synthesis in neuronal perikarya. At different times after electrical or mechanical stimulation, fish were killed and the RNA of the electric lobes isolated and used to prime a cell-free translation system. The radioactively labeled synthesis products were separated by 2-D electrophoresis and the patterns obtained from different specific RNAs were compared. Several proteins exhibited time-dependent changes in rate of synthesis as indicated by the relative intensity of their autoradiographic spots. These proteins appear/disappear in a specific way, indicating a modulated translational response to neuronal activity.

Cloning of rat brain succinyl-CoA:3-oxoacid CoA-transferase cDNA. Regulation of the mRNA in different rat tissues and during brain development.

3-Oxoacid CoA-transferase, which catalyses the first committed step in the oxidation of ketone bodies, is uniquely regulated in developing rat brain. Changes in 3-oxoacid CoA-transferase activity in rat brain during the postnatal period are due to changes in the relative rate of synthesis of the enzyme. To study the regulation of this enzyme, we identified, with a specific polyclonal rabbit anti-(rat 3-oxoacid CoA-transferase), two positive cDNA clones (approx. 800 bp) in a lambda gtll expression library, constructed from poly(A)+ RNA from brains of 12-day-old rats. One of these clones (lambda CoA3) was subcloned into M13mp18 and subjected to further characterization. Labelled single-stranded probes prepared by primer extension of the M13mp18 recombinant hybridized to a 3.6 kb mRNA. Rat brain mRNA enriched by polysome immunoadsorption for a single protein of size 60 kDa which corresponds to the precursor form of 3-oxoacid CoA-transferase was also found to be similarly enriched for the hybridizable 3.6 kb mRNA complementary to lambda CoA3. Affinity-selected antibody to the lambda CoA3 fusion protein inhibited 3-oxoacid CoA-transferase activity present in rat brain mitochondrial extracts. The 3.6 kb mRNA for 3-oxoacid CoA-transferase was present in relative abundance in rat kidney and heart, to a lesser extent in suckling brain and mammary gland and negligible in the liver. The specific mRNA was also found to be 3-fold more abundant in the brain from 12-day-old rats as compared with 18-day-old foetuses and adult rats, corresponding to the enzyme activity and relative rate of synthesis profile during development. These data suggest that 3-oxoacid CoA-transferase enzyme activity is regulated at a pretranslational level.

Stimulation of pancreatic secretory process in the rat by low-molecular weight proteinase inhibitor

Oral application of a single dose of a new synthetic proteinase inhibitor Camostate (Foy-305) in male Wistar rats was carried out together with studies of in vitro amino acid incorporation followed by separation of proteins by two-dimensional gel electrophoresis. The aim of this experiment was to analyze changes produced by the inhibitor in total protein and individual enzyme biosynthesis. Administration of 100 mg/kg Foy-305 resulted in significant inhibition of total pancreatic protein synthesis, without changes in fractional rates for individual enzymes. 50 mg/kg Foy-305 induced a 10-fold elevation of cholecystokinin (CCK) levels in serum; this persisted for 3 h and led to a significant increase in the total rate of protein synthesis with peak values at 6 and 9 h (78% and 84% above control levels, respectively), returning to control by 15 h. Changes in fractional rates of synthesis occurred with a latency of 6 h and were restricted to amylase and the anionic form of trypsinogen and chymotrypsinogen. Amylase biosynthesis decreased by about 40% from control levels at 9 h to return to control levels by 15 h. Increased synthesis of trypsinogen and chymotrypsinogen was observed; this was also phasic. The results show similar enzyme-specific regulation as previously described for exogenous CCK stimulation and for the adaptation of the pancreas to diets enriched in protein. They demonstrate the effectiveness of pulsatory endogenous hormone release in the regulation of protein synthesis.

Minor Burns Lead to Major Changes in Synthesis Rates of Plasma Proteins in the Liver

The effect of minor burns on the rates of synthesis of plasma proteins in the liver was studied in white Buffalo rats. Burns of second to third degree, covering 0.8% of total body surface, were produced by short application of a hot piece of metal to the skin under ether anesthesia. Levels of mRNA in extracts from liver removed after 24 hr were measured by hybridization to radioactively labeled specific cDNA probes. The level of mRNA for major acute phase alpha 1-protein (also called cysteine proteinase inhibitor or T1-kininogen) increased 20-fold, that of fibrinogen mRNA 8-fold, and that of alpha 1-acid glycoprotein mRNA about 9-fold. The levels of albumin mRNA and transthyretin mRNA (also called prealbumin) decreased to about 80% of normal and the level of transferrin mRNA did not change significantly. Thus, although the percentage of burnt skin was only very small, a typical acute phase response of plasma protein synthesis in liver was observed.

Early changes in the synthesis of nuclear and cytoplasmic proteins are induced by nerve growth factor in differentiating rat PC12 cells.

Differentiation of rat pheochromocytoma PC12 cells into neuron-like cells was induced by nerve growth factor (NGF) and changes in the apparent rate of synthesis of cellular proteins were analyzed. Attention was particularly focused on the first few hours of exposure to NGF before significant neurite outgrowth was detectable. Cultures were pulse-labeled for 1-h periods with [35S]methionine and proteins were extracted from various subcellular fractions and analyzed by one-dimensional gradient and two-dimensional equilibrium and nonequilibrium gel electrophoresis. The results showed that although the general level of protein synthesis remained constant, by 8 h NGF increased the apparent rate of synthesis of approximately 11 cytoplasmic and 5 nuclear proteins. For several of these proteins, the effect was apparently NGF-specific, since no induction was observed in dibutyryl cAMP-treated cells. Of interest was the following observation: of the five nuclear proteins, NGF increased the synthesis of two proteins with MrS of 56,000 [doublet] and 50,000 D that were associated with a biochemically and morphologically defined nuclear matrix fraction. A cytoplasmic protein, with an Mr of 92,000 D (pI 4.8) appeared to be induced de novo by NGF. NGF also decreased the rate of synthesis of several cytoplasmic and nuclear proteins of low molecular mass (less than 40,000 D). Since only 1-h pulses of [35S]methionine were used, and since experiments with actinomycin D showed that most of these NGF-induced early changes in rates of synthesis included a transcription-dependent step, it seems likely that early effects of NGF include activation of specific genes.

Transcriptional regulation of ribosomal proteins during a nutritional upshift in Saccharomyces cerevisiae.

The relative rates of synthesis of Saccharomyces cerevisiae ribosomal proteins increase coordinately during a nutritional upshift. We constructed a gene fusion which contained 528 base pairs of sequence upstream from and including the TATA box of ribosomal protein gene rp55-1 (S16A-1) fused to a CYC1-lacZ fusion. This fusion was integrated in single copy at the rp55-1 locus in the yeast genome. During a nutritional upshift, in which glucose was added to cells growing in an ethanol-based medium, we found that the increase in the relative rate of synthesis of the beta-galactosidase protein product followed the same kinetics as the change in relative rates of synthesis of several ribosomal proteins measured in the same experiment. This demonstrates that the nontranscribed sequences upstream from the rp55-1 gene, which are present in the fusion, are sufficient to mediate the change in rates of synthesis characteristic of ribosomal proteins under these conditions. The results also suggest that a change in transcription rates is mainly responsible for the increase in relative rates of synthesis of ribosomal proteins during a nutritional upshift in S. cerevisiae.

The physiological bases of nutrient responses during lactation.

The lactating dairy cow responds to dietary changes according to the nutrients provided and her propensity to produce milk. The allocation of nutrient use to milk production or body gain is generally termed ‘partition’ and has been shown to be a function of both the animal and its diet. As feed intake is increased, the lactating cow responds with progressively smaller increments of increased milk yield and progressively larger increments of body tissue. This ‘diminishing return’ is not associated with a change in efficiency of any physiological process. It is the result of changes in the rates of synthesis of milk components and of body tissue. The individual rates of milk synthesis and body tissue synthesis may be influenced independently. It is therefore somewhat awkward to speak of control of partition as though it were a single physiological process. We are concerned with the synthesis of specific milk constituents and the role of specific metabolites in influencing the rates of synthesis and, thereby, milk yield and composition. It is unfortunately very difficult to tell if milk production is limited by nutritional state, i.e. nutrient supply, or by the hormone balance or other physiological factors involved in milk synthesis. There is very clearly a close interaction between nutrient supply and physiological control of milk synthesis. Bauman & Currie (1980) used the term ‘homeorhesis’ which they credit to Kennedy (1967) to refer to the coordinated changes in metabolism of body tissues necessary to support a physiological state such as lactation. It is clear that nutrient response is altered by physiological state and that for a given physiological state, milk production will be influenced by nutrient supply. Recent reviews have extensively discussed the endocrine control of partition and milk biosynthesis (Hart, 1983; Akers, 1985)’ mammary-gland differentiation (Houdebine, 1985), uptake and utilization of amino acids by the mammary gland (Clark, 1975; Mepham, 1982), and glucose homeostasis (McDowell, 1983). It is apparent that the control of milk synthesis or partition involves both substrate availability and endocrine control. The effects of substrate availability may be presumed from experiments in which dietary manipulation has caused a change in rate of milk synthesis and a corresponding change in body-tissue energy balance.