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While there are indications that some other non-J3 PLC enzymes interact with G proteins, their identities need to be determined. The observations that [37subunits can regulate some forms of PLC raise questions about the specificity of PLC stimulation but also offer a putative pathway for PTX-sensitive stimulation of this signalling pathway. Finally, the emergence of the Gq proteins as regulators of Ptdlns-PLC invite the speculation that they may participate in hormonal regulation of other lipid-derived second messengers. The tools are at hand to dramatically improve our understanding of this intricate network for regulation of these important second messengers. References 1 Berridge, M. J. (1987) Annu. Rev. Biochem. 56, 159-193 2 Boyer, J. L., Hepler, J. R. and Harden, T. K, (1989) Trends Pharmacol. Sci. 10, 360-364 3 Ross, E. M. (1989) Neuron 3, 141-152 4 Sternweis, P. C. (1990) Trends Neurosci. 13, 122-126 5 Simon, M. I,, Strathmann, M. P. and Gautam, N. (1991) Science 252,802-808 6 Kazir, *{. et al. (1991) Annu. Rev. Biochem. 60,

DESCRIPTIONS OF SEVERALregulatory phenomena that are responsible for short-term adaptations by microbes have hitherto been based on physiological observations; however, increased knowledge of the underlying biochemical and genetic mechanisms now means that the terms used can be defined quite rigorously to remove the confusion that exists surrounding nomenclature. Ambiguities in the nomenclature of regulatory mechanisms stem partly from applying some expressions, already used for physiological changes, to molecular events. For example, either one or both of the two different physiological controls, 'induction' and 'derepression', may regulate the synthesis of enzymes involved in utilizing a given substrate. However, in molecular terms, induction inactivates a repressor, so induction and derepression may seem to be the same phenomenon. The extensive studies that have been published K-D1 Entian is at the Institut for M~krobiologie der Johann Wolfgang Goethe-Universit&t Frankfurt, Theodor-Stern-Kai 7, Haus 75A, D-6000 Frankfurt/M, Germany. J. A. Barnett is at the School of Biological Sciences, University of East Angiia, Norwich, UK NR4 7TJ.

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349-400 7 Brown, A. M, (1990) Annu. Rev. Biochem. 52, 197-213 8 Linder, M. E. and Gilman, A. G. (1992) Sci. Am. 267, 56-65 9 Pang, I-H. and Sternweis, P. C. (1990) J. Biol. Chem. 265, 18707-18712 10 Strathmann, M. and Simon, M. (1990) Prec. Natl Acad. Sci. USA 87, 9113-9117 11 Smrcka, A, V., Hepler, J. R., Brown, K. O. and Sternweis, P. C. (1991) Science 251,804-807 12 Taylor, S. J., Smith, J. A. and Exton, J. H. (1991) J. Biol. Chem. 265, 17150-17156 13 Taylor, S. J., Chae, H. Z., Rhee, S. G. and Exton, J. H. (1991) Nature 350, 516-518 14 Waldo, G. L, Boyer, J. L, Morris, A. J. and Harden, T. K. (1991) J. Biol. Chem. 266, 14217-14225 15 Berstein, G. et al. (1992) J. Biol. Chem. 267, 8081-8088 16 Gutowski, S. et al. (1991) J. Biol. Chem. 266, 20519-20524 17 Shenker, A., Goldsmith, P,, Unson, C. G. and Spiegel, A. M. (1991) J, Biol. Chem. 266, 9309-9313 18 Wange, R. L,, Smrcka, A. V., Sternweis, P. C. and Exton, J. H. (1991) J, Biol. Chem. 266, 11409-11412 19 Wu, D., Lee, C. H., Rhee, S. G. and Simon, M. I. (1992) J. Biol, Chem. 267, 1811-1817 20 Lee, C. H. et al. (1992) J. Biol. Chem. 267, 16044-~16047 21 Rhee, S. G., Suh, P. G., Ryu, S. H. and Lee, S. Y. (1989) Science 244, 546-550 22 Crooke, S. T. and Bennett, C. F. (1989) Cell Cal.

10, 309-323 23 Rhee, S. G. (1991) Trends Biochem. Sci. 16,

297-301 24 Wahl, M. I., Daniel, T. O. and Carpenter, G. (1988) Science 241, 968-970 25 Meisenhelder, J., Suh, P. G., Rhee, S. G. and Hunter, T. (1989) Cell 57, 1109-1122 26 Wahl, M. I. et al. (1989) Prec. Natl Acad. Sci. USA 86, 1568-1572 27 Kim, U. H., Kim, H. S. and Rhee, S. G, (1990) FEBS Lett. 270, 33-36 28 Kriz, R. eta/. (1990) Ciba Found. Syrup. 150,

112-127 29 Park, D. eta/. (1992) J. Biol. Chem, 267,

16048-16055 30 Amatruda Ill, T. T., Steele, D. A., Slepak, V. Z. and Simon, M. I. (1991) Prec. Natl Acad. Sci. USA 88, 5587-5591 31 Thomas, M. H.,Geraint, Geny, B. and Cockcroft, S. (1991) EMBOJ. 10, 2507-2512 32 Sternweis, P. C., Smrcka, A. V. and Gutowski, S. (1992) Phil. Trans. R. Sec. Lend. Ser. B 336,

35-42 33 Camps, M. et al. (1992) Eur. J. Biochem. 206,

821-831 .34 Tang, W. J. and Gilman, A. G. (1992) Science

254, 1500-1503 35 Ryu, S. H. eta/. (1990) J. Biol. Chem. 265,

17941-17945 36 Geet, C. V. et al. (1990) J. Biol. Chem. 265,

7920-7926 37 Bizzarri, C., Girolamo, M. D., D'Orazio, M. C. and Corda, D. (1990) Prec. Natl Acad. Sci. USA 87,

4889-4893 38 Litosch, I. (1989) Biochem. J. 261,245-251

There are several kinds of regulation that enable microbes to cope with rapidly changing supplies of nutrients. This is exemplified by sugar metabolism in Saccharomyces cerevisiae. Some readily reversible controls affect the activity of enzymes, either by allosteric activation and deactivation, which often occur within seconds, or by covalent modification, within minutes. Other controls regulate the amount of enzyme present in the cells, either by irreversible proteolytic inactivation of the enzyme, or by influencing enzymic synthesis. The nomenclature of these processes is often confused. on sugar metabolism by the yeast, Saccharomyces cerevisiae, now make it possible to integrate many physiological and genetic findings more precisely than before, allowing the clarification of the nomenclature and the regulatory processes themselves. The utilization of sugars by yeasts is regulated by several kinds of enzymic control, described in physiological terms as allosteric activation and dea~:ti-

vation, interconversion, specific proteolysis (inactivation) and induction, repression and derepression. Allosteric activation, deactivation and interconversion refer to the regulation of the activities of enzymes; whereas inactivation, induction, repression and derepression are mechanisms by which the amount of enzyme is regulated. Like other yeasts, S. cerevisiae grows on hexose sugars, such as glucose, © 1992,ElsevierSciencePublishers,(UK)

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GLUCOSE

GLYCOLYSIS f DEACTIVATOR

L

ATP

)\

I FRUCTOSE 6-PHOSPHATE

i

phosphofructokinase

/

AMp

/

L

fruct°se2'6-blsphosphate

)

DEACTIVATORS 1 AMP fructose 2,6bisphosphate

/

fructose 1,6-bisphosphatase

FRUCTOSE I,i-B|SPHOSPIIATE

I

}

PHOSPHOENOLPYRUVATE ~

I ( xcIVTO' 1 citrate

/

pyruvate kinaseI

O/ALOACETATE

/

PYRUVATE

I I

fructose 1,6-bisphosphate

TCA

GLYOXYLATE CYCLE

//'

T

CYCLE

i

GLUCONEOGENESIS

Figure 1 The regulation of glycolysis and gluconeogenesis is by activators and deactivators.

when they are available1. However. unlike most other yeasts, S. cerevisiae catabolizes hexoses mainly to ethanol (fermentation), even under aerobic conditions2; catabolism is therefore almost solely anaerobic, by glycolysis, and does not involve the tricarboxylic acid cycle. After exhausting the exogenous sugar, this yeast can then catabolise ethanol aerobically, by the tricarboxylic acid cycle. When a yeast is utilizing ethanol (or other Cz or C3 carbon compounds) for growth, the hexoses that are necessary for cell wall biosynthesis are provided by gluconeogenesis (Fig. 1), Strict regulatory systems exist that prevent 'futile cycles' such as the simultaneous and unregulated glycolysis and gluconeogenesis pathways. Several regulatory mechanisms to be discussed in this review are involved in these switches and are summarized in Tables I and II.

growth substrate, there is little substrate available for these enzymes and the two enzymes are regulated by positive and negative allosteric effectors. PFK is deactivated by ATP and activated by both AMP and fructose 2.6-bisphosphate. Pyruvate kinase is deactivated by both ATP and citrate and depends on

Table I. Mechanisms regulating enzymic activity Kind of regulation

Physiologicalobservation

Enzymicmechanism

AIIosteric activation and deactivation

Reversibleloss of enzymicactivity

Change in affinity for substrate

Interconversionby covalent modification

ReversibLeloss of enzymicactivity within minutes

Usually phosphorylationof enzyme

Table II. Mechanisms regulating the amount of enzyme Kind of regulation

Physiologicalobservation

Molecularmechanism

Inactivation

Irreversible toss of enzymicactivity

Specific proteolysisof the enzyme

Induction

Increase in enzymicactivity in responseto oresence of inducer (substrateor structurallysimilar compound)

Inducerevokes activation of transcription

Derepression

Increase in specific activity after removing repressing substrate

De-inhibitionOf transcription

Regulation by allosteric effectors

The key enzymes regulating glycolysis are 6-phosphofructokinase (PFK) and pyruvate kinase (Fig. 1). When a nonfermentable carbon source is the

fructose 1.6-bisphosphate (the product of PFK) for activation. The gluconeogenic counterpart of PFK, fructose-l,6-bisphosphatase (FBPase), is deactivated both by AMP and fructose 2,6-bisphosphate. On non-fermentable carbon sources, such as ethanol, fructose 2,6-bisphosphate concentrations are undetectable.

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If glucose becomes available, the concentration of fructose 2,6-bisphosphate increases within seconds, which activates PFK and deactivates FBPase (for review see Ref. 2).

Regulation by enzyme conversion (covalent modification) Phosphory]ation of Ser11 (Ref. 3) of FBPase halves its activity in one or two minutes 4. This covalent modification is reversible 4, and is a further regulatory response to the addition of glucose to the medium of S. cerevisiae cells growing on a non-fermentable carbon source.

Table III. Mutant genes involvedin glucose repression and derepression Mutant allelea

Regulation by glucose repression and derepression The glucose effect, originally defined as the absence of certain enzymic activities in the presence of ~exogenous glucose, is now known as glucose or carbon catabolite repression. It has a timecourse of many minutes, or even hours, and results from the failure of transcription. The many enzymes subject to

508

Physiological role of wild-type gene

Group I: Mutants affecting glucose repressible enzymes

hxk2 (hexl, girl)

c~-Glucosidase, invertase, enzymes of galactose utilization and TCA cycle, respiratory enzymes

Structural gene for hexokinase PII

hex2 (regl)

c¢-Glucosidase, invertase, enzymes of galactose utilization and respiratory enzymes

?Negative regulatory element 2°

cat80 (grrl)

c~-Glucosidase, invertase, enzymes of galactose utilization and TCA cycle, respiratory enzymes

Leucine-rich motifs indicate protein-protein interactions; ?primary response element21; enlarged cells

cidl

c(-Glucosidase, invertase, enzymes of galactose utilization

Not known

Regulation by enzyme inactivation The inactivation of gluconeogenic enzymes by glucose starts 5-10 min after the addition of glucose to cells growing on non-fermentable carbon sources and the activity of ' these enzymes is diminished within 1 h. The gluconeogenic enzymes that are subject to glucose inactivation include cytosolic malate dehydrogenase, FBPase, phosphoenolpyruvate carboxykinase and is0citrate lyase (for review see Ref. 5). This glucose inactivation is irreversible and immuno-precipitation experiments have shown that proteolysis occurs6; however, it is no longer thought that phosphorylation renders FBPase susceptible to proteolytic degradation 7. It is now clear that inactivation (of, for example, FBPase) caused by glucose occurs in the vacuole: mutants defective in proteinase A (S. cerevisiae aspartic proteinase), which regulates other vacuolar proteinases, are not subject to inactivation by glucose8. Moreover, the susceptibility of an enzyme to inactivation depends on the differential targeting of the enzymes to the vacuole and not on the specificity of the vacuolar proteases. Thus ]3-galactosidase, which is not normally inactivated in the presence of glucose, becomes subject to inactivation when fused to FBPase (K-D. Entian, K. Kbhn, M. Zweimfiller and M. Rose, submitted). It remains to be determined how this mechanism is triggered by glucose.

Enzymes

Group I1: Mutants affecting glucose derepressible enzymes

cat1 (ccrl,snfl)

c(-Glucosidase, invertase, enzymes of galactose utilization and TCA cycle, respiratory and gluconeogenic enzymes and those of the glyoxylate cycle

Protein kinase 22 associated with nuclear fraction 23

cat3 ( s n f 4 )

c¢-Glucosidase, invertase, enzymes of galactose utilization and TCA cycle, respiratory and gluconeogenic enzymes and those of the glyoxylate cycle

Subunit of CAT1 protein kinase24; positive gene dosage effect; associated with nuclear fraction

Group II1: Mutants epistatic over derepression mutants cat1 and cat3

cat2

Fast derepression of c¢-glucosidase, invertase, enzymes of galactose utilization, gluconeogenic enzymes and those of the glyoxylate cycle

Notknown

cat4 (migl)

c~-Glucosidase, invertase, enzymes of galactose utilization

Transcriptional repressor; zinc finger protein; negative gene dosage effect26; efficient derepression 15

aThe allele names that were published first have been given preference and are referenced; alternative designations are in brackets. Readers interested in tracing references that use the alternative names should contact the authors directly.

repression by glucose include glycoside hydrolases and those of galactose catabolism (the Leloir pathway), gluconeogenesis, the tricarboxylic acid cycle, the glyoxylate cycle and the respiratory chain (for reviews see Refs 9, 10). Genetic analysis of repression by glucose has revealed two kinds of mutant. First, 'glucose-repression mutants', in which enzymes that are glucoserepressible in the wild type have high activities, even when glucose is the carbon source; none of these mutants isolated so far affect gluconeogenic enzymes. Second, 'glucose derepression mutants', which stay in the glucoserepressed state even after exhaustion of glucose and affect all the glucoserepressible enzymes mentioned above. A number of different mutant alleles have been described for each of several

independent glucose-repression loci (see also Table III; for review see Ref. 11); the designation for each allele used here is the one that was published first. The following four groups of mutants can be distinguished phenotypically: (I) hxk2, hex2 (Ref. 12), cat80 (Ref. 12) and cidl (Ref. 13), which affect glucoserepressible enzymes; (Il) catl and cat3 (Ref. 14), which affect derepression; (Ill) cat2 and cat4 (Ref. 15), which are epistatic over glucose derepression mutants (see Table III for synonyms for groups I, II and Ill); (IV) certain mutants that also give defects in glucose repression and derepression, but have additional regulatory disorders, tupl (Ref. 16) (synonyms flkl, umr7, cyc9, amml, aarl, slf2, aer2), cyc8 (ssn6) and rgll. The genes specifically involved in glucose repression and derepression

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are represented by the mutant groups (I), (II) and (III) (see Table III). Hexokinase PII appears to have a major function in triggering glucose repression ~7.This function can be complemented by over-expression of isoenzyme hexokinase PI, but not glucokinase18,19, although all three enzymes (PI, PII and glucokinase) have similar apparent affinities for glucose. The roles of phosphorylation and regulation therefore appear to be quite separate. No further glycolytic steps beyond glucose phosphorylation are necessary for glucose repression TM and the actual signal for glucose repression is still unknown. The HEX2 protein is located in the nucleus ~° and contains strongly acidic regions, which may compete with transcriptional activators. The CAT80 protein has leucine-rich motifs, which is indicative of protein-protein interactions2~; genetic analysis suggests that CAT80 functions early in the pathway of glucose repression. The two genes of group If, catl and cat3, are necessary for glucose derepression. How they affect transcription is not known. The CAT1 protein appears to be a protein kinase 22and the CAT3 protein 23 corresponds to a subunit necessary for the function of CAT1 kinase24. Mutations within these genes give highly pleiotropic effects, including the inability to grow with C2 or C3 carbon sources, disaccharides or galactose 14. Transcription of nearly all glucoserepressible genes is prevented in catl and cat3 mutants. The CAT1-CAT3 heteromeric protein seems to be associated with the nuclear membrane 23. The phenotype of catl and cat3 derepression mutants can be suppressed epistatically, by cat2 and cat4 (Ref. 15) of group III. The cat2 mutant has no effect on glucose repression, whereas several enzymes in cat4 mutants are not repressed by glucose. The TUP1-CYC8 protein complex seems to be generally involved in cellular gene expression and is, perhaps, only concerned secondarily in the regulatory cascade for glucose repression and derepression. Group IV proteins are part of a large multimeric complex2S of 800 kDa. The mutants cyc8 and tupl (Ref. 16) do not repress the synthesis of cz-glucosidase or invertase; they are epistatic over catl and cat3 for invertase derepression, although they do not derepress gluconeogenic enzymes. Further cellular functions that are also greatly affected by these mutations

dent of growth conditions. However, the specific activities of enolase II (Refs 29, 30) and pyruvate kinase 12,3° are increased threefold during growth with glucose as the carbon source, compared with that observed with C2 and C3 carbon sources. Similar findings also apply to both pyruvate decarboxylase 12,3°and alcohol dehydrogenase I and are the result of increased transcription. The phenomenon is sometimes referred to as 'glucose induction', although it differs physiologically from induction proper, such as that by galactose, as these enzymes are not specific for glucose utilization. A new definition is necessary to distinguish precisely between the terms 'constitutive' and 'inducible'. Physiological observations have suggested that the rate of expression of a constitutive gene is constant and independent of environmental changes. Molecular analysis, however, has revealed a system of highly sophisticated regulation, even for largely constitutive genes. Scanning the promoters of many constitutive Regulationby sugar induction genes has shown that there are several In S. cerevisiae, the enzymes involved positive and negative regulatory sites specifically in galactose or maltose (UAS and URS). Each regulatory elcatabolism are synthesized in response ement reacts positively or negatively to to the presence of these sugars (for environmental changes, such as the review see Ref. 1). The transcription of available carbon sources. Altogether, the genes encoding the appropriate interaction of these elements results transport carriers and enzymes (a- physiologically in constitutive gene exglucosidase and enzymes of the Leloir pression. Evidence for this interpretpathway) need specific induction. ation has come from a study on the gcrl Induction by galactose needs the GAL4 mutant 31, which pleiotropically reduces protein with a cysteine zinc finger, as a transcription of several glycolytic final gene activator. The negative regu- enzymes that have been considered latory protein GAL80 forms a heteromer constitutive. Thus, a constitutive enzyme with GAL4, which prevents its function. is not without transcriptional control The molecular mechanism of galactose and, hence, the term is more useful in regulation (reviewed in Ref. 28) illus- describing physiological observations trates the difference between induction than transcriptional regulation. and derepression. Derepression depends only on the absence of the repressing General considerations compound, whereas induction depends Changes in the specific activities of on the presence of an inducer. In addi- enzymes or transport carriers are usution to induction, galactose and maltose ally given in terms of amount of subutilization are also subject to glucose strate converted per mg protein. Thus, repression and derepression. Glucose when measuring a regulatory phenomrepression overcomes induction; that enon as the result of gene expression, is, the genes are not transcribed as long an activity should be considered relaas glucose is present. GAL4, which is tive to changes in total protein synthe activator for induction, appears to thesis; the measured activity is the be the molecular target for glucose combined result of both transcriptional repression, as the transcription of GAL4 efficiency and the rate of protein synis also subject to glucose repression. thesis. Nearly all physiological and genetic estimations ignore the fact that Constitutiveenzymesand other factors increased or decreased protein synregulatinggene expression thesis, together with a constant tranThe specific activities of most glyco- scriptional rate, would also decrease lytic enzymes are relatively indepen- or increase measurements of specific include a-specific mating defects, flocculence and the inability of homozygous diploids to produce ascospores. Deletion analysis of the invertase promoter identified DNA sequences at which gene activators (UAS elements) or gene repressors (URS elements) can bind. The invertase URS element was used to characterize its gene repressor CAT4 (MIG1)26. The gene activator for invertase has not yet been found. However, an activator protein (Dapl) has been reported for FBPase27; Dapl binding occurs when cells are grown on ethanol. The physiology of glucose repression, in terms of molecular biology and genetics, therefore involves two regulatory mechanisms, namely, glucose repression and glucose derepression. Glucose repression is the result of the binding of a transcriptional repressor protein that prevents transcription and glucose derepression is the result of the binding of a transcriptional activator protein that promotes transcription.

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activity. Imbalances between transcriptional activity and growth rate may therefore also affect the interpretation of gene expression. The regulatory changes discussed above are necessary for survival in rapidly changing environments. They are observed not only in sugar utilization of eukaryotic microbes, but also in prokaryotes and even in differentiated cells of plants and animals and for systems involved in the catabolism of nitrogen compounds. For the sake of clarity, it is hoped that the various terms for regulatory mechanisms will be used rigorously in the future.

References 1 Barnett. J. A. (1976) Adv. Carbohydrate Chem. Biochem. 32. 125-234 2 Gancedo, C, and Serrano R. (1989) in The Yeasts 4.Vo/.3. 2nd ednl (Rose. A. H. and Harrison, J. S., eds), pp. 205-259, Academic

Press 3 Rittenhouse, J.. Moberly, L. and Marcus. F. (1987) J. Biol. Chem. 262.10114-10119 4 Lenz A-G. and Holzer, H. (1980) FEBS Lett. 109, 271-274 5 Holzer. H. (1976) Trends Biochem. Sci. 1. 178-181 6 Neeff. J. et al. (19781 Biochem. Biophys. Res, Commun. 80. 276-282 7 Rose. M. et al. (1988) FEBS Lett. 241.55-59 8 Chiang, H-L. and Schekman. R. (1991) Nature 350. 313-318 9 Entian, K-D, (1986) Microbiol. Sci. 3. 366-371 10 Gancedo. J. M. and Gancedo, C. (1986) FEMS Microbiol. Rev. 32. 179-187 11 Gancedo. J. M. 11992) Eur. J. Biochem. 206 297-313 12 Entian. K-D. and Zimmermann. F. K, I1980) Mol. Gen. Genet. 177,345-350 13 Neigebom. L. and Carlson M. (1987) Genetics 115.247-253 14 Entian. K-D. and Zimmermann. F. K, (1982) J. Bacteriol. 151. 1123-1128 15 SchOIler, H-J. and Entian. K-D. (1991) J. Bacteriol. 173. 2045-2052 16 Wickner. R. B. (1974) J. Bacteflol. 117 252-260 17 Entian. K-D. (19801 Mol. Gen. Genet. 178.

633-637 18 M~ H., Bloom. L. M.. Walsh. C. T. and Botstein. D. (19891 Mol, Cell. Biol. 9. 5643-5649 19 Rose. M.. Albig, W. and Entian. K-D. (1991) Eur. J. Biochem. 199. 511-518 20 Niederacher. D. and Entian K-D. [1991) Eur. J. Biochem. 200 311-319 21 Flick. J. S. and Johnston, M. (1991) Mol. Cell Biol. 11, 5101-5112 22 Celenza, J. L. and Carlson. M. (1986) Science 233. 1175-1180 23 SchOIler. H-J. and Entian. K-D. (1988) Gene 67, 247-257 24 Celenza. J. L.. Eng, F. J. and Carlson. M (1989) Mol. Cell. Biol. 9. 5045-5054 25 Williams. F. E.. Varanasi. U. and Trumbly, R. J. (1991) Mol. Ceil, Biol. 11. 3307-3316 26 Nehlin. J. O. and Ronne, H. (1990) EMBO J. 9. 2891-2898 27 Niederacher. D. et al. Curr. Genet, (in press) 28 Johnston, M, (19871 Microbiol. Rev. 51. 458-476 29 Fr6hlich. K-U. and Entian K-D. (1982) FEBS Lett. 139. 164-166 30 Entian. K-D.. Fr6hlich K-U. and Mecke D. (19841 Biochim. Biophys. Acta 799, 181-186 31 Clifton. D. and Fraenkel. D. G, (19811J. Biol. Chem. 256. 13074-13078

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This is an excellent multi-author book. forming the third in the series concerned with the characterization of signaltransducing receptors. As the author points out in the introduction, binding studies using receptor-ligand interactions in tissue preparations appear deceptively simple to carry out but can be difficult to perform well and to analyse and interpret. The individual authors set out to provide detailed information about each step: the selection of a suitable ligand, generation of receptor preparations from tissues, the design and execution of in vitro binding assays, separating bound from free ligand and the correct analysis of the data obtained. The initial chapters describe the selection of conventional radiochemical ligands and the use of radiolabelled polypeptide neurotoxins to study some of the voltage-dependent K* channels. Two appendices contributed by the tnajor suppliers. Amersham International PLC and Du Pont-NEN provide a classification of commercially available radioligands with some brief comments about their selectivity and pharmacological action.

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Regulation of sugar utilization by Saccharomyces cerevisiae.

There are several kinds of regulation that enable microbes to cope with rapidly changing supplies of nutrients. This is exemplified by sugar metabolis...
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