The ZFY gene family in humans and mice

EIEVIEWS 17 Zalokar, M., Audit, C. and Erk, I. (1975) Dev. Biol. 47, 419-432 18 Edgar, B.A., Kiehle, C.R and Schubiger, G. (1986) Cell44, 365-372 19 Lin, H. and Wolfner, M.F. Cell (in press) 20 Raft, J.W. and Glover, D.M. (1988) J. CellBiol. 107, 2009-2019 21 Gonzalez, C. et al. (1990) J. Cell Sci. 96, 605-616 22 Nurse, R (1990) Nature 344, 503-508 23 Lee, M.G. and Nurse, R (1987) Nature 327, 31-35 24 Jimenez, J. et al. (1990) EMBOJ. 9, 3565-3571 25 Lehner, C. and O'Farrell, P (1990) ~ B O J . 9, 3573-3581 26 Masui,Y. and Markert, C.L. (1971)J. Exp. Zool. 177, 129-146 27 Gautier, J. et al. (1990) Cell 60, 487-494 28 Evans, T. et al. (1983) Cell 33, 389-396 29 Whitfield, W.G.E, Gonzalez, C., Sanchez-Herrero, E. and Glover, D.M. (1989) Nature 338, 337-340 30 Lehner, C.F. and O'Farrell, RH. (1990) Cell61, 535-547

T h e discovery of ZFY in 1987 ~ was widely publicized. Identified as a result of a systematic search for the testis-determining factor (TDF) gene on the human Y chromosome, it had several characteristics that made it a good candidate for that gene. However, from the outset it was clear that the story was not going to be a simple one, as a virtually identical gene (ZFX) was found on the X chromosome of many eutherian mammals. The situation in mice is even more complex 24, with a total of four genes, two of which map to the Y chromosome (Zfy-1 and Zfy-2), one X-linked (Zfx), and an autosomal homologue on chromosome 10 (Zfa; see Fig. 1). This family of genes has been intensively studied, but most of the results of this work have been interpreted in the light of the proposed role for ZFY in testis determination. Recent genetic evidence, which redefined the position of TDF, and the isolation of a far more promising candidate gene have now eliminated the possibility that ZFY is directly involved. In this review we re-evaluate the data, concentrating on aspects of the structure and expression of the ZFY gene family that may give an indication of the true function of these genes. Protein structure All the members of the ZFY gene family identified so far encode highly homologous proteins with the same overall structure. All have 13 zinc fingers of the Cys-Cys/His-His type (Fig. 2), encoded by a single exon. The zinc finger motif is one of the highly conserved motifs present in eukaryotic sequence-specific DNA-binding proteinsS. ZFY-related genes form a distinct subfamily of zinc finger proteins showing a twofinger repeat pattern of primary structure. This may reflect their origin from a primordial gene that had two fingers and might further imply an underlying repeat in the sequences to which they bind. Some of the ZFY genes - Zfa, Zfy-2 from the Mus musculus musculus subspecies 6 and the chicke~ Zfl) gene: - have mutations involving key residues in the third zinc finger, which could affect the coordination of Zn2+ and

31 Whitfield, W.G.F., Gonzalez, C., Maldonado-Codina, M.G. and Glover, D.M. (1990) ~ B O J . 9, 2563-2572 32 Raff, J.W. and Glover, D.M. (1989) Ce1157, 611-619 33 Anderson, K. (1989) in Genes and Embryos (Glover, D.M. and Hames, B.D., eds), pp. 1-37, IRL Press at Oxford University Press 34 Raft, J.W., Whiffield, W.G.F. and Glover, D.M. Development (in press) 35 Hartenstein, V. and Posakony, J.W. (1990) Dev. Biol. 138, 147-159 36 Edgar, B.A. and O'Farrell, RH. (1990) Cell 62, 469-480 37 O'Farrell, R, Edgar, B.A., Lakich, D. and Lehner, C. (1989) Science 246, 635-640 38 Foe, V. and Odell, G.M. (1989) Am.J. Zool. 29, 617 D.M. GLOVER IS IN THE CELL CYCLE GENETICS GROUP, CANCER RESEARCH CAMPAIGN LABORATORIES~ MEDICAL SCIENCES INSTITUT~ UNIVERSITY OF DUNDEE, DUNDEE D D I

The ZFYgenefamilyin humans and mice PETER KOOPMAN, ALAN ASHWORTH AND ROBIN LOVELL-BADGE

For several years, ZFY (zinc finger gene on the Y chromosome) was considered the best candidate for the human testis-determining gene TDF. This gene and its close relatives have been intensely studied in the hope of understanding the molecular biology of sex determinatio~ particular~ in humans and mice. Now that there is overwhelming evidence that ZFY and TDF are distinct loci, we are left with a large body of dato. and a question: what do these genes really do? hence the folding of that finger (Fig. 2). As these genes have been maintained, for several million years in the case of Zfa, the mutations probably do not impair the function of the proteins. Adjacent to the zinc finger domain is a short basic region similar to the nuclear localization signal of SV40 large T antigen. This region of Zfy-1 can direct a heterologous protein to the nucleus of tissue culture cells (S. Swift and A. Ashworth, unpublished). Thus the ZFY family gene products are potentially nuclear proteins. The amino termini of the ZFY gene products are very acidic (approximately one in four residues are aspartic or glutamic acid over a 40 kDa domain). Acidic regions in transcription factors such as the yeast GAL4 protein have been shown to mediate transcriptional activation functions 8. Experiments performed in yeast support the notion that the acidic region in the ZFY gene products can serve a similar function9. Combining the structural information on the typical ZFY family gene product gives us a picture of a protein with all the hallmarks of an activating transcription factor (Fig. 3a). This model for the structure of ZFY-related proteins may not be the whole story. Several alternative

TIG APRIL1991 VOL. 7 NO. 4 l~)~)l t:lsc~k'r St i e n ( e P u N i s h e r s l i d (I:K) Ol{~

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H~EVIEWS Mouse

Human •• • / - -

' Zfy 1....

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,

ZFX 99%

Mouse Zfa is of recent origin (it is absent from the distantly related mouse species Mus pahari and Mus platythrix) and was derived by retroposition of an RNA transcript from Zfx 6. Almost all of the several hundred k n o w n retroposons are heavily mutated pseudogenes; Zfa is unusual in being an expressed gene. Like two other expressed retroposons - PGK2 (Ref. 12) and PDHA2(Ref. 13) - Zfa is expressed onl}, in the testis. Interestingly, these other retroposons were also derived from genes on the X chromosome. This c h r o m o s o m e is thought to be inactivated early in spermatogenesis, and so the presence of autosomal copies that could be expressed in a post-meiotic germ cell might have a selective advantage in evolution. Other vertebrates, such as marsupials, birds, reptiles and amphibians, also have genes homologous to ZFE but these are not present on the sex chromosomes ~.n,~s The Xenopus gene has been sequenced and shown to encode a protein similar in overall structure to those e n c o d e d by the mammalian genes (F. Connor and A. Ashworth, unpublished). The high sequence c(mservation of these genes confirms their importance.

79

Zfy-2 98%

'

_[78%

Zfx Zfa ', 97% b/GH The genes of the human and mouse ZFYfamily. Circles represent pairs of genes that are highly similar in their primary' structure. Figures show the degree of homology of the zinc finger regions, at the amino acid level, within and between these pairs. Mouse Zfy genes appear to be evolving differently from the other genes of the family.

Expression

transcripts have been identified, resulting from differential exon usage6.mJL (Fig. 3b). Some of these vary only within the 5' untranslated region of the mRNAs and their function is obscure. Others, however, would result in proteins with radically different properties. For example, a deletion of part of the acidic d o m a i n seen in some transcripts may interfere with the ability of the resulting protein to act as a transcription factor. In another spliced product, the putative nuclear localization signal would be absent from the protein. This protein could be located in the cytoplasm, where it could bind to RNA, as has been shown for the zinc finger protein TFIIIA.

Evolution

Expression of a gene in a certain tissue or at a particular stage of d e v e l o p m e n t can provide valuable clues to the possible role of the gene. The observ'atkm that both ZFXand ZFYare expressed at all times in all human tissues l < r p r o b a b l y indicates a general cellular functkm rather than a role in a specific developmental event. Similarly, mouse Zfx is ubiquitously expressedg. Stage or tissue specificity for any of the alternatively spliced transcripts has not been demonstrated, althc~ugh the testis contains transcripts of discrete ,;izc due to the use of alternative polyadenylatkm sites. Many models relating to ZF}TZPX interaction natu rally assumed that ZI~X is subject to X inactivation. In fact, ZFX is a rare e x a m p l e of a gene outside the pseudoautosomal region that escapes from X inacti vation 16. Unlike the human gene, mouse Zfx maps close to the putative X-inactivation centre (Xce) TM. It might be assumed that a gene so close to Xce would be subject to X inactivation, and prelimina U experiments support this view (A. Ashworth el aL, u n p u b l i s h e d ) As

Most placental mammals, including humans, cows, dogs, rabbits and rats, have two ZFg-related genes ~, one located on the Y c h r o m o s o m e (ZFY) and one on the X c h r o m o s o m e (ZFX). The extreme similarity of the proteins e n c o d e d by these loci (Fig. 1) has led to the suggestion that they can Mus musculus function interchangeably. The musculus Zfy-2 situation in mice is again quite 6 AMINO ACIDS different. Zfy-1 and Zfy-2 must DELETED have arisen by a gene duplication event since the Mus rnusculus Zfa divergence of rats and mice. TYR Although these genes are very similar to each other, they are distinct from the h u m a n ZFY genes, and from mouse Zfx. Chicken Zfb _.t' ',..._ The constraints on the evoluARG tion of these two genes a p p e a r to have b e e n relaxed. This is #TG~ almost certainly linked with Mutations in the third zinc finger in tile ZkTgene family. Tile schematic diagram (~f tic the fact that they are expressed finger shows only the key cysteine and histidine residues, which are thought t(~ ~ ~rdmat~ in a very different fashion Zn2* ions tetrahedrally, essential to the normal confi~rmation of the finger Arrow> indic ak the effects of three mutations seen in this gene family. from their human counterpart. TIG APRIL1991 VOL. 7 NO. 4

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[]~EVIEWS the mouse Zfy genes are not widely expressed, this leads to a curious situation where all human cells have two active ZFY-like genes (ZFY plus ZFX or two copies of ZFX) whereas mouse cells generally have only one (Zfx). In contrast to ZFYand the X-linked genes, Zfy-1 and Zfy-2 show a very restricted pattern of expression z,10,19 (Table 1). Male germ cells apparently express Zfy-1 at moderate levels at all stages of their development, accounting for the presence of Zfy-1 transcripts in both foetal and adult mouse testes. While a low level of Zfy-2 expression has been reported in foetal testes 19, Zfy-2 transcripts only become abundant in testes between 7 and 14 days postnatally, and are then present at a level about threefold higher than Zfy-1 (Ref. 20). Neither gene is expressed in testes lacking germ cells. The most obvious conclusion is that mouse Zfy expression is intimately linked to spermatogenesis. Two observations compromise this view. Zfy-1 is expressed in foetal oocytes in XY mutant female mice 20. It is not yet k n o w n whether this persists during later development of the oocyte; no Zfy expression has been detected in adult XY ovaries 19, but very few oocytes are likely to have survived in the material tested. Further, Zfy transcripts have been described in mouse foetal tissues other than testis at a low level, but their significance is unclear and they are absent from their adult counterparts 19. Zfy-1 but not Zfy-2 is expressed at moderate levels in embryonic stem (ES) cells 1°. The finding that the two genes are regulated differently in ES cells and during male germ cell differentiation may reflect individual roles for each, even though their structure would predict that they bind to similar nucleic acid sequences.

those with a more general role in transcription, for example Spl, to those that are likely to be of specific developmental importance, such as Krox-20. The distribution of transcripts of these genes is taken to reflect their function. By analogy, it is likely that human ZFX and ZFY, and mouse Zfx, have a general cellular function. The restricted pattern of expression shown by the mouse Zfy genes can be interpreted in different ways. Initially there seems to be some correlation with cell types that in females would not show X inactivation, such as XX ES cells (and therefore, perhaps, preimplantation embryos), and germ cells in the foetal ()vary. Perhaps Zfy-1 is expressed at these stages to compensate dosage between males and females. Zfv-1 and Zfy-2 are both expressed at later stages of male germ cell development. Here again there is some correlation with X chromosome activity, as the X is shut down during spermatogenesis. The activity of the Ylinked genes, as well as Zfa, in testis may simply reflect the need to compensate for the lack of Zfx gene product. (It is also formally possible that Zfy expression in the testis may be entirely fortuitous, as many genes are activated de novo in the testis with no apparent function21.) On the one hand, then, all members of the ZFY family in mice could have essentially the same function, and interact with the same downstream genes in all tissues. Alternatively, selective pressure to maintain these genes on the Y chromosome during mammalian evolution suggests a male-specific function. For many years, sex determination was thought to be the only role of the Y chromosome. It is n o w known that several gene functions map to the Y chromosome for which the ZFY genes could be considered candidates. Potential functions of Z/~ genes First, there is testis determination itself. Although There are probably more than 300 zinc finger ZFY was the best candidate for 7DF for some time, it genes in the mammalian genome. These vary from is n o w clear that ZFY and TDF are two separate loci. Early evidence for this conclusion has been reviewed elsewhere=, 23. More recently, (a) four XX males and intersexes were identified w h o possess Y sequences that do not include Acidic domain: Basic domain: 13 Zinc fingers: ZFF 24. These Y sequences trans-activation? nuclear targeting? nucleic acid binding? map to within 35 kb of the pseudoautosomal boundary (b) (ZFY is some 200 kb further on), redefining the minimum Mouse Zfx type II ~ ~ region of the Y able to specify male development. A new TDF candidate from this region has been described25.26; MouseZfxtypelll~ ~ VVVVVVVVVVVVI the properties of this gene, Mouse Zfa SRE are so far entirely consistent with its being TDF 2L28. The varying degrees of divergence of the four SRYHuman ZFX type 1 positive, ZFY-negative, XX individuals from a normal male phenotype could be interFIG[] preted as a requirement for an I)omain composition of ZF}Zrelated g~he products. (a) Canonical three-domain structure. additional gene on the Y for (b) Predicted products of alternative splicing. The origin of each type of product is shown on the left. complete masculinization. This TIC, APRn.1991 VOL. 7 NO. 4

m

[]~EVIEWS gene could be ZFY or any closely linked gene. However, as these individuals carry SRY within just a small piece of Y unique sequence on an X chromosome, which will be subject to X inactivation and/or position effects, it seems to us more likely that the range of phenotypes depends on the proportion of cells expressing SRY, or the level of expression per cell, in the developing genital ridge. Another gene mapping to the Y chromosome directs the expression of H-Y antigen. However, this gene maps to the long arm of the human Y chromosome and must therefore be distinct from ZFY, which maps to the short ar,m. A number of fairly subtle growth differences have been found between male and female embryos. Recently, a preimplantation effect has been attributed to a gene on the mouse Y chromosome 29. This effect depends on the strain of origin of the Y Chromosome. The finding that Zf),-1 is expressed in ES cells which are derived from, and approximate to, cells of the inner cell mass of the blastocyst would be consistent with its being responsible for this effect; however, a difference has yet to be found between the Zfy genes on the different types of Y chromosome. In humans, X0 embryos frequently die in utero, and if born suffer from Turner syndrome. At least part of the Turner phenotype has been attributed to a gene for which there must be an X-linked copy that escapes X inactivation, and a Y-linked copy that maps to the short arm, close to the pseudoautosomal boundary. This gene could be similar to the mouse preimplantation growth gene described above and hence could be ZFY (see Ref. 23). Evidence that it is not comes from analysis of a ZFY-negative XY female w h o does not show the short stature associated with Turner syndromeS.30. Recently, a Y-linked homologue of the X-chromosomal gene encoding the ribosomal protein $4 has been found3L This gene maps close to ZFY, and the X-linked copy apparently escapes X inactivation. This gene is present in this ZFY-negative XY female, and so may be a more valid candidate for the gene that prevents the Turner phenotype. Expression of mouse Zfl: genes in male germ cells is often taken to reflect a role in spermatogenesis. This cannot be a role in early spermatogenesis, as X0 germ cells in X0/XY mosaics develop normally to stem cell spermatogonia in the absence of Zfy genes; however, very few of these X0 cells complete the differentiating spermatogonial divisions32. A dramatic block in spermatogonial proliferation at a similar stage has been observed in X Sxr'/0 but not X Sxr/0 male mice3~. Sxr (sex reversed) is a mutation resulting in transfer of short arm material from the Y to an X chromosome, denoted X Sxr. A variant, Sxr', has resulted from deletion of part of the Sxr region. The block to spermatogenesis has been attributed to the loss of a gene, termed Spy, in the Sxr' deletion34. This deletion is now thought to have involved a recombination event that has resulted in fusion of the Zfv-1 structural region to the Zfv-2 promoter (EM. Simpson and D.C. Page, unpublished). While it is possible that Spy is another gene within the deletion, the coincidence of the disruption of the ZJj, genes, and the fact that both TIG APRIL

TAm~ 1. ZFY-related genes expressed in different mature cell types in h u m a n s and mice Human

Mouse

Somatic cells

ZFX, ZPT(male) ZFX a (female)

Zfx

Male germ cells

ZFY

Zfa, Zfy-1, Zfy-2

aDouble dosage.

are normally expressed at significant levels during spermatogenesis, strongly suggest that one or both Zfy genes is Spy.

Future prospects To understand more precisely the role of the ZFY gene family, it will be necessary to identify their binding sites, the genes they control, and any other transcription factors with which they might interact. These may not be the same in all cell types. Also, although many of the genes appear to be expressed ubiquitously at the RNA level, this may not reflect the distribution of protein. The generation of specific antibodies will therefore be important. The changes associated with the ZFY gene family during evolution are especially intriguing. As ZFX is highly conserved among eutherian mammals and is most closely related to the homologues found in other vertebrates, it is likely to represent the archetypal gene of the family. We can assume that this archetypal gene was autosomal, and became located on the X and Y chromosomes of eutherian mammals as part of the general evolution of the sex chromosomes. As the X chromosome appears to be made up of segments from a number of autosomes, were there other genes syntenic with ZFX that moved at the same time, and what happened to their Y-linked homologues? It will be interesting to examine the association of ZFX/ZFY with the ribosomal protein $4 genes during evolution, and to look at the effect of X inactivation on the mouse $4 homologue. The difference between the human and mouse ZPX genes with respect to X inactivation deserves further study. It may be possible to test how the human gene escapes X inactivation by introducing it onto the mouse X chromosome with different lengths of flanking sequence. For example, this could be done ,~ia insertion into the Hprt locus in ES cells. The apparent change in control of Zfv x~as accc~mpanied by the rapid divergence of Zfv in terms of its primary sequence, its duplication and its regulation Does this divergence reflect acquisition of novel functions or the loss of its ubiquitous role and subsequent relaxation of structural constraints? Substituting, for example, the human ZFY zinc finger domain for that of the mouse, via homologous rccombin,tti,)n. should make no difference if the latter is correct. Finally, there is the duplication of the mouse Z/i' genes and the re-acquisition of an aut()somal c()py in mice. It should be possible t() test whether any of these genes is redundant, or whether they haxe

1951 VOL. 7

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I'~EVIEWS

significant roles, by gene targeting experiments to inactivate each in turn. Although the Z F Y gene family does not have the role originally proposed for it, it is clearly an important set of genes, with a fundamental cellular role. It may be that any part they may play in spermatogenesis is an extension of this role, necessitated by the peculiar biology of germ cells. Further work on these genes will help to resolve not only their function, but may also throw light on the evolution of the X and Y chromosomes and on the process of X inactivation.

Acknowledgements We thank Paul Burgoyne, David Page and Claude Nagamine for helpful comments and for access to unpublished results.

References 1 Page, D.C. etal. (1987) Cell51, 1091-1104 2 Nagamine, CM., Chan, K., Kozak, C.A. and Lau, Y-F. (1989) Science 243, 80-83 3 Mardon, G. el al. (1989) Science 243, 78-80 4 Mitchell, M. et al. (1989) Genetics 121,803-809 5 Miller, J., McLachlan, A.D. and Klug, A. (1985) EMBOJ. 4, 1609-1614 6 Ashworth, A., Skene, B., Swift, S. and Lovell-Badge, R. (1990) P214BOJ. 9, 1529-1534 7 DiLella, A.G., Page, D.C. and Smith, R.G. (1990) Neu, Biol. 2, 49-56 8 Ptashne, M. (1988) Nature 335, 683-689 9 Mardon, G. et al. (1990) Mol. Cell. Biol. 10, 681-688 10 Koopman, R, Gubbay, J., Collignon, J. and Lovell-Badge, R. (1989) Nature 342, 940-942 11 Schneider-G~idicke, A. et al. (1989) Nature 342, 708-711 12 McCarrey, J.R. and Thomas, K. (1987) Nature 326, 501-505 13 Dahl, H-H.M. el al. (1990) Genomics 8, 225-232

How do ue-specific gene "! nction? Tissue Specific Gene Expression edited by Rainer Renkawitz,VCHVerlagsgesellschaft, 1989.DM198.00,£71.00 (xvii + 221 pages) ISBN3 527 27875 3 Tissue Specific G e n e Expression

comprises a dozen reviews and a short, but very informative introduction (by E.L. Winnacker) about various aspects of expression of tissue-specific genes in vertebrates. Apart from one chapter on the tissue-specific alternative splicing of troponin-T (R.E. Breitbart and B. Nadal-Ginard), all the other contributions are devoted to control at

14 Sinclair, A.H. et al. (1988) Nature 336, 780-783 15 Bull, J.J., Hillis, D.M. and O'Steen, S. (1988) Science 242,

567-569 16 Schneider-G/~dicke, A. et al. (1989) Cell 57, 1247-1258 17 Lau, Y-F.C. and Chan, K. (1989) Am.J. Hum. Genet. 45,

942-952 18 Keer, J.T. et al. (1990) Genomics 7, 566-572 19 Nagamine, C.M., Chan, K., Hake, L.E. and Lau, Y-EC. (1990) Genes Dev. 4, 63-74 20 Gubbay, J. et al. (1990) Development 109, 6474553 21 Willison, K. and Ashworth, A. (1987) Trends Genet. 3 ,

351-355 22 Erickson, R.P. and Verga, V. (1989) Am.J. Hum. Genet. 45, 6714574 23 Burgoyne, P.S. (1989) Nature 342, 8604"~62 24 Palmer, MS. el al. (1989) Nature 342, 937-939 25 Gubbay, J. el al. (1990) Nature 346, 245-250 26 Sinclair, A.H. et al. {.1990) Nature 346, 240-2£i/ 27 Berta, R et al. (1990) Nature 348, 448-450 28 Koopman, P. el al. (1990) Nature 348, 450--452 29 Burgoyne, P.S. in Advances in Developmental Biology (Vol. 1) (Wassarman, P.M., ed.), Greenwich (in press) 3 0 Page, D.C., Fisher, E.M.C., McGillivray, B. and Brown, L.G. (1990) Nature 346, 279-281 31 Fisher, E.M.C. etal. (1990) Cell63, 1205-1218 32 Levy, E. and Burgoyne, P.S. (1986) Qytogenet. Cell Genet. 42, 208-213 3 3 Burgoyne, P.S., Levy, E.R. and McLaren, A. (1986) Nature 320, 170-172 34 Sutcliffe, M.J. and Burgoyne, RS. (1989) Development 107, 373-380 P. KOOPMAN AND R. LOVELL-BADGEARE IN THE LABORATORY OF EUKARYOTIC MOLECULAR GENETIC~ M R C NATIONAL INSTITUTE FOR MEDICAL RESEARCH, THE RIDGEWAY, MILL HILL, LONDON N W 7 1AA, UK; A. ASHWORTH XS AT THE CXESTER B E A ~ LABORATOgXES, THE INSTITUTE FOR CANCER RESEARCH, FULHAMROA~ LONDONSW3 6BJ, UK.

the transcriptional level, in particular to trans-acting factors. It is a risky adventure to edit a book about an expanding field such as tissue-specific gene expression. A few years after the discovery of the first tissue-specific transcription factor, Oct-2, controlling the B-lymphocyte-specific transcription of immunoglobulin genes (L.M. Staudt et al., N a t u r e 323, 640-643, 1986), each month now sees a multitude of reports about the transcriptional control of tissuespecific genes, on the isolation of new transcription factors and their structural and functional properties. Hence several of the highlights of the past few years, such as the detection of muscle-specific transcription and differentiation factors (e.g. the 'master gene' M v o D by Weintraub and colleagues), could not be included. The editor, Rainer Renkawitz, T1G APRIl. 1991 VOL. 7 NO. 4

i 3(1

had the foresight to select a few 'model' systems, such as the immunoglobulin and globin genes, and he persuaded a number of outstanding groups to summarize their latest news in several excellent reviews. Four articles deal with the immunoglobulin genes: their chromatin structure during gene expression (W.T. Garrard); the 'prototype' of a tissue-specific trans-acting factor, Oct-2 (E. Schreiber et al.); the positive and negative factors contributing to the activity of heavy chain gene enhancer (B. Wasylyk et al.); and studies on the kappa light chain enhancer (K. Mocikat et aLL Two chapters each are devoted to the expression of globin genes (J.L. Gallarda et al.; F. Grosveld et al.) and of liver-specific genes (the {x-antitrypsin gene, P. Monaci et al.; the albumin gene, P. Herbomel and