COMMENTARY

Lampbrush chromosomes

H. C. MACGREGOR

Department of'/.oology, Univeisity of Leicester, Leicester LEI 7RH, UK

1986 was a celebration year for lampbrush chromo-somes (LBCs), marked by the publication of Callan'scomprehensive and authoritative book on these struc-tures. My commentary begins where Callan's bookends, standing on tiptoe beside a large and ratherneat assembly of well-established facts, principles andhypotheses, and trying hard to catch a glimpse of whatlies ahead.

Several major principles can now be accepted withconfidence. LBCs are a feature of the growing oocytesof most animals, except mammals and certain insects.Callan (1986) has reviewed this principle in depth, andhas discussed the significance of exceptional cases. Themain defining feature of LBCs, their lateral loops, areregions of intense RNA synthesis, and together theseloops produce a large variety of transcription products,though not all coding sequences in the genome aretranscribed. Much of the transcribed polyadenylatedRNA is passed to the cytoplasm and its quantity ismaintained at a steady-state level by continuing syn-thesis and turnover. This RNA is stored in thecytoplasm and probably functions in the regulation andmaintenance of oocyte maturation and early embryonicdevelopment.

The relationship between a transcription unit (TU)and a lampbrush loop (Fig. 1) has been well defined byGall et al. (1983). Transcription initiates at structuralgene promoters at the ends of loops or TUs, fails toterminate and 'reads through' into adjacent non-codingsequences. The 'read through' hypothesis (Varley et al.1980; Gall et al. 1983) probably applies to all lamp-brush loops. The very large size of lampbrush TUssuggests that they must include transcripts of inter-spersed repetitive elements of the genome. Structuralgenes in large genomes are more widely spaced, inter-spersed with non-coding repetitive DNA, than in smallgenomes. One might therefore expect LBCs from largegenomes to have longer loops (TUs) than those fromsmall genomes, and this is precisely what has beenobserved (see Callan, 1986; Macgregor, 1980).

On the molecular front, with the main questionsregarding RNA synthesis resolved, there remain the

Journal of Cell Science 88, 7-9 (1987)Printed in Great Britain © The Company of Biologists Limited 1987

problems of the fate and significance of the massiveamounts of non-coding transcripts, and the manner inwhich certain initiation sites are 'selected' for high-leveltranscription on lateral loops, whereas others remainrelatively inactive on or within the chromomeres. Ofparticular interest are the recent studies of Epstein etal. (1986) and Epstein & Gall (1987). These investi-gators have focussed their attention on the nature andtranscription of the 330 base-pair (bp) satellite DNA ofNotophthalmus viridescens (Nv2). Nv2 occurs in tan-demly repeated clusters throughout the genome and itssequence is highly conserved among the salamandridspecies so far examined (Epstein et al. 1986). It istranscribed on lampbrush loops by read-through fromadjacent structural gene promoters, and homologousstrand-specific cytoplasmic transcripts are found in thecytoplasm of a variety of tissues. The transcriptscorrespond in size to the Nv2 repeat unit or to simplemultiples of this unit. The transcripts seem to beencoded by a specific subset of the genomic Nv2sequences, and they undergo site-specific self-catalysedcleavage in vitro (Epstein & Gall, 1987), a reaction thatresembles the self-cleavage of certain small infectiousRNAs found in plants. The cleavage of Nv2 occurs at asite that is homologous to the conserved cleavage site ofthese infectious RNAs. The significance of this remark-able discovery remains a mystery, but perhaps will notdo so for long!

Future directions for the study of LBC RNAs andtranscription processes are at present unclear. Anunequivocal demonstration of the location and patternof transcription of any well-defined low-copy-number'structural' gene would be welcome. The question ofco-selection of initiation sites on sister loops, especiallywith regard to multigene families, is challenging, and isrelated to the important questions of whether sisterloops, arising from the same chromomere, are indeedmolecularly identical, and if so, how this remarkablefeat is accomplished?

With regard to the proteins of LBCs, there isexciting progress, mainly through the applicationof monoclonal antibody technology. Antibodies have

Fig. 1. Phase-contrast micrograph of a long lampbrushloop that consists of three tandemly arranged TUs eachwith the same direction of polarization. The arrowsnumbered 2 and 3 indicate the starts of the second andthird TUs. X 1000. (Reproduced with the kind permissionof Van Nostrand Reinhold Company, New York, from:Macgregor, H. C. (1986). The lampbrush chromosomes ofanimal oocytes. In Chromosome Structure and Function(ed. M. S. Risley), pp. 152—186. New York: Van NostrandReinhold Company.)

been prepared that bind specifically to chromomeres(histones), hnRNP (heterogeneous nuclear RNP) coreproteins on loops, particular loops or sets of loops, andmorphologically distinct regions of certain loops. Mostof the loop-specific antibodies are almost certainlybinding to proteins that are associated with nascentRNA transcripts, and some of them are present in thenuclei of somatic cells and oocytes (Sommerville etal. 1978; Lacroix et al. 1985; Roth & Gall, 1987).Predictably, such proteins are absent from metaphase

chromosomes, but present in the cytoplasm surround-ing the chromosomes (Roth & Gall, 1987). Callan(1986) remarks that the use of monoclonal antibodiesfor investigation of the cytogenetics of LBCs and theirloops is potentially "on a par with in-situ hybridiz-ation". We shall see! In-situ hybridization has taken usa long way in a short time, the answers it has given havebeen highly and immediately significant, and there issurely more to come.

In this connection I wish to offer a comment on theobjects known as 'spheres' on LBCs (see Callan &Lloyd, 1960; Callan, 1986). In his book, Callanremarks that "intensive study of the relationship be-tween spheres and histone transcripts could be reward-ing". His view is based on the close proximity in severalamphibia of the spheres, whose antigenic specificity hasbeen established, and the loops that transcribe histonemRNA. The inference is that the spheres and histoneloops are in some way functionally linked. This may beso, and there is no doubt that the spheres shouldcontinue to be investigated vigorously; but it should beborne in mind that urodele karyotypes are conserva-tive, and the fact that two 'genes' have remained next toone another for more than 50 million years does notnecessarily mean that they are functionally related.

On the cytological front, there has been one majorbreakthrough in the past year. Callan et al. (1987) haveshown that it is possible to obtain excellent prep-arations of LBCs from Xenopus laevis and this papergives a detailed technical protocol and an excellentLBC map. With the considerable arsenal of molecularprobes from A', laevis and a wide knowledge of thegenetics and molecular biology of this species, the wayis now open to combine cytological and molecularapproaches on a scale that hitherto seemed impossible.

Persistent looking at LBCs continues to confirmexisting ideas and open up new questions. The giantloops on chromosome 2 of A', vtridescens, reported byGould et al. (1976) as uniquely unbreakable by therestriction enzyme Haelll, have been shown to bebreakable with this enzyme, but only on loops thatconsist of tandem TUs and only at points where there isan abrupt change from the end of one TU to the startof another: good confirmation of the principle behindthe read-through hypothesis (Macgregor & Fairchild,unpublished observation). Scanning and transmissionelectron microscopy have confirmed the doubleness ofthe interchrom6meric fibril (Bakken & Graves, 1975),shown that all loop matrices are composed of standard30 nm RNP particles (N'Da et al. 1986) and helped toexplain the specific morphologies of the matrices ofcertain loops and other parts of the chromosomes(Bonnanfant-JaiseJa/. 1985; Macgregor, 1986). One ofour own (unpublished) observations is that the inci-dence of sister loops that are of distinctly unequallength is much higher than might be supposed from a

8 //. V. Macgregor

reading of the early descriptive literature. There is noeasy answer to this point, and the observations aresuggestive of real molecular differences between sisterchromatids or discordant transcriptional activity onsister loops. Accordingly, I believe it may now bemisleading to say that sister loops are 'identical', andthe significance of non-identity needs attention.

Finally, let us not be so intensely occupied with thefunctioning of LBCs that we forget that they aremeiotic chromosomes, and their organization mayinclude features that are of long-term adaptive signifi-cance in relation to evolution and speciation. LBCs areunique in so far as they allow us to look at half bivalentsand at sister chromatids, and to examine DNA se-quence organization in a germ cell that has undergonemany rounds of mitosis with ample opportunities forsister chromatid exchanges, followed by all the eventsof early meiotic prophase. After all, oocytes are cellsthat work for evolution, and in their differentiationtheir chromosomes are subject to some of the mainmolecular driving forces of that process.

References

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Lampbrush clnvinosonies