Zebrafish Embryo as a Developmental System.pdf

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Zebrafish Embryo as aDevelopmental SystemSharon L Amacher, University of California - Berkeley, Berkeley, California, USA

Zebrafish are simple, rapidly-developing animals that are amenable to detailed

developmental and genetic analyses. Despite their relatively simple morphology, they

share many developmental features with other vertebrates. By combining developmental

studies in zebrafish with those in other animals, we will begin to understand the complex

events that occur as an embryo develops from a one-celled zygote to a complex,

multicellular organism.

Introduction

In the last twenty years, the tropical freshwater zebrafish,Danio rerio, has emerged as an excellent model ofvertebrate development. Much of the pioneering workdone to advance zebrafish as a model system was initiatedin the laboratory of Dr George Streisinger, but nowlaboratories all over the world use zebrafish to studydevelopmental processes. This article will describe thefeatures of zebrafish that make them particularly tractableto embryological and genetic analyses.

Why Study the Zebrafish?

Zebrafish are simple, rapidly developing animals that are

amenable to detailed developmental analyses. Despitetheir relatively simple morphology, they share manydevelopmental features with other commonly-studiedvertebrates, such as frogs, chickens, and mice. Thus,studies of zebrafish will teach us about the development ofother organisms, including humans.

Several features make zebrafish an excellent animal touse in studies of genetics and development. First, zebrafishadults reach sexual maturity quickly and are very fecund.Thus, large numbersof embryos can be collected for study.Second, zebrafish embryos develop externally. Eggs thatare extruded into the surrounding water by adult femalesare immediately fertilized by sperm from adult males.

Embryos are thus easily accessible for observation andexperimentation. Third, zebrafish embryos are opticallyclear. Under a microscope, one can visualize tissues that liedeep within the embryo, follow individual cells duringdevelopment, and recognize developmental mutants.Fourth, zebrafish embryos develop rapidly, as illustratedin Figure1. To emphasize the rapid embryonic developmentand to review important developmental milestones, theseven periods of embryonic development are describedbelow.

Zebrafish embryonic development

Zygote period (034

h)

The newly fertilized egg, or zygote, is about 0.7 mm idiameter at the time of fertilization. The embryo developwithin a transparent eggshell, calledthe chorion, which cabe easily removed without harming the embryo. Durinthis period, cytoplasmic streaming separates the zygotinto two visibly different parts: a clear blastodisc at thanimal pole and a yolky cytoplasm at the vegetal pole. Thblastodisc is the part of the zygote that undergoes cleavagand gives rise to blastomeres, the cells that give rise to thembryo.

Cleavage period (3421

4h)

This period includes the first six regular, rapid, ansynchronous divisions of the embryo. At the end of thcleavage period, the developing embryo sits as a mound otop of the large yolk.

Blastula period (21451

4h, Figure 1a,b)

Many important processes occur during the blastulperiod. Prior to this period, the embryo has utilizeproteins and RNA that the mother deposited in the eggDuring midblastula stages, the embryo begins to transcribits own genes. Not only does transcription begin, bu

embryonic genes are regionally expressed, reflecting thfact that blastomeres are now becoming different from onanother. When embryonic transcription begins, cell divsion times gradually lengthen, so that blastomeres nlonger divide together. Another major event that beginduring the blastula period is epiboly, which describes ththinning and spreading of the cellular blastodisc over thyolk. Near the end of this period, the blastomeres arreferred to collectively as the blastoderm, a sheet-like arraof cells that sits like a cap on top of the large yolk.

Article Contents

Introductory article

. Introduction

. Why Study the Zebrafish?

. Genetic Analysis of Zebrafish Development

. Summary

ENCYCLOPEDIA OF LIFE SCIENCES / & 2001 Nature Publishing Group / www.els.net


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Gastrula period (51410h, Figure 1c,d)

Epiboly continues during the gastrula period. During thisperiod, cell movements dramatically reorganize the em-bryo,so that bythe end ofthisperiod, threeembryonic axesare clearly visible. That is, the anterior head of the embryois clearly distinguished from the posterior tail bud, dorsaltissues are distinct fromventral ones, andmedial tissues areeasily discernible from lateralones. Many embryonic genes

are expressed in region-specific patterns. Cell movementsduring gastrulation also produce the germ layers (ecto-derm, mesoderm and endoderm) of the embryo. Finally,near the end of the gastrula period, the neural plate forms;this is the first morphological sign of central nervoussystem development.

Segmentation period (1024h, Figure 1eg)

The embryo elongates considerably during this period. Acomplete complement of about 30 bilateral somite pairs

forms in an anterior to posterior sequence, giving thsegmentation period its name. The rudiments of primarorgans, such as notochord, kidney, and blood, becomvisible. The central nervous system undergoes dramatichanges; the neural plate forms a solid neural keel, whicthen hollows out to form a neural tube. Additionally, thbrain becomes regionalized, with forebrain, midbrain anhindbrain subdivisions becoming distinct from one another. Finally, muscular contractions occur, and th

embryo moves within the chorion. To contrast, a mousembryo at one day after fertilization has only juscompleted the first cell division to form two cells.

Pharyngula period (2448h, Figure 1g)

This period is named for the seven pharyngeal arches tharapidly form during this period. Pharyngeal arch development is common to all vertebrates, and in zebrafishpharyngeal arches later form the jaw and gills. Also durin

Figure 1 Zebrafishdevelopmental series. Hourspost-fertilization (h)are shownin thelower rightof eachpanel.(a) Earlyblastula.(b) Midblastula. (c)Eargastrula. (d) Midgastrula. The arrows in (c) and (d) mark the position of the blastoderm margin and indicate the process of epiboly; epiboly is the

movement of the blastoderm as it crawls over the yolk toward the vegetal pole. (e) Early segmentation stage embryo. Arrowheads in (e) markdeveloping somites. (f)Midsegmentation stageembryo.(g) Pharyngula. (h)Hatchingembryo.The darkcells scatteredover thehead, bodyand yolkandthe eyes are pigment cells.

Zebrafish Embryo as a Developmental System

2 ENCYCLOPEDIA OF LIFE SCIENCES / & 2001 Nature Publishing Group / www.els.net


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the second day, the head straightens, fins form, pigmentcells differentiate, the circulatory system forms and theheart beats, and coordinated swimming movements begin.Embryos become responsive to touch, revealing thedevelopment of sensory-motor reflexive circuits.

Hatching period (4872h, Figure 1h)

Embryos hatch from their chorions during the third day.By this time, many of the organ rudiments have formed;however, the gut and its associated organs are stilldeveloping. In addition, this period is marked by rapiddevelopment of pectoral fins, jaws and gills.During the twodays after hatching (72120 h), the zebrafish juvenile startsto swim and feed. The embryonic yolk, which has sustainedthe embryo until this time, is almost depleted.The zebrafishembryo has rapidly become a small version of the adult.

Experimental methods for studying zebrafishdevelopment

This section will review powerful experimental methodsavailable to study zebrafish, such as cell lineage analysesand cell transplantation, that elucidate when and whereimportant developmental events occur. When theseexperiments are coupled with genetic analyses (as will bereviewed later), we learn what genes are important fordevelopmental processes, as well as when and where genefunction is required for normal development.

Cell lineage analysis and the zebrafish fate map

The rapid development of the transparent zebrafishembryo allows researchers to do many kinds of experi-

ments quickly and easily. For example, zebrafish cells canbe injected with lineage tracer dyes, which are fluorescentdyes that permanently identify the injected cell and itsdescendants. Since zebrafish embryos are transparent, the

fates of injected cells can be followed in living embryos blooking forfluorescent cells at later times. We can use timelapse analysis to watch the movements, divisions andifferentiation of fluorescently-labelled cells. In otheanimals that are not optically clear, one would have tserially section the embryo in order to find labelled cells.

Lineage tracing experiments have taught us a great dea

about zebrafish embryogenesis. When a zebrafish blastomere is injected with lineage tracer dye during cleavage anblastula stages, the labelled cell and its progeny contributlater to many different tissues. The labelled blastomerebegin to be fate-restricted during gastrula stages. Fatrestriction means that a labelled cell and its progeny wicontribute to only one tissue type, demonstrating that, athe time of labelling, the cell was specified to a particulafate. By injecting single cells at the gastrula stage anfollowing the fates of their progeny, a zebrafish fate mahas been constructed. This fate map is shown in Figure2antells us to what tissue a cell in a particular position of thgastrula is likely to contribute. Cells located near th

blastoderm margin will make mesodermal derivatives. Amarginal blastomere on the dorsal side will normally maknotochord, and a marginal blastomere on the ventral sidwill normally make blood. Cells located farther from thblastoderm margin and closer to the animal pole wicontribute to epidermis and neural tissue (brain and spinacord).When we look at the live gastrula embryoin Figure2ait looks deceptively simple, much like a knitted cap pulleover a large yolk cell. The fate map shows us that evethough the embryo may look simple, complicated cespecification events are occurring, and many cells aralready different from their neighbours. In fact, as showin Figure 2c, cells in one fate map domain often expres

different genes from cells in a neighbouring fate madomain, revealing that differential gene expression correlates well with cell fate choice.

Figure2 Zebrafish gastrula fate map andregionalgene expression. (a) Live early gastrula. (b)Schematic drawing of the fate map at a similar stage.Thembryonic blastoderm is shown in red. Mesodermal fates (notochord, muscle and blood) arise near the blastoderm margin, whereas neural tissue andepidermisarisefarther from theblastodermmargin.Onlya subsetof tissuefatesis shown.AP, animalpole;VP,vegetalpole;D, dorsal; V,ventral.(Redraw

from Kimmel CB et al. (1990) Development108: 581594.) (c) Early gastrula embryo fixed and processed to detect expression of the floating head(flhgene. The dorsal cells at the blastoderm margin that express flh (stained blue) are found within the notochord domain of the fate map.




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Cell transplantation analyses

The information from the fate map allows us to ask moresophisticated questions. We know from the gastrula fatemap that cells in certain positions are fated to give rise tospecific tissues. In addition, region-specific gene expressionpatterns are also apparent.Thus, duringearly gastrulation,cell fate specification is occurring; that is cells are becomingdistinct from their neighbours and begin to expresscharacteristics appropriate for their future fates. However,the fate map does not reveal whether those cells arecommitted to a particular fate or to express particulargenes. One can only learn about cell commitment, theirreversible decision to adopt a particular fate, if a cell ischallenged by moving it to a new environment. If a cell iscommitted, it will adopt the fate consistent with its originalposition, even though it is surrounded by neighbouringcells that adopt different fates. If a cell is not committed, itwill develop like its new neighbours and adopt the fateappropriate for its new environment. In zebrafish, singlecells can be aspirated into a micropipette and transplantedfrom their original positions to ectopic positions. When acell is transplanted during early gastrulation from oneregion of the fate map to another, it adopts the fateappropriate for its new location. This result shows thatearly gastrula cells are not committed to a particular fate.When single cells are transplanted at midgastrulation, theyadopt fates appropriate for their original position. Takentogether, these results show that cell fate specificationbegins at early gastrulation, but cell commitment (theirreversible decision to adopt a particular fate) does notbegin until midgastrulation, two hours later. Below, wediscuss how the powerful techniques of cell lineage analysis

and cell transplantation are used to characterize the defectsin developmental mutants.

Genetic Analysis of ZebrafishDevelopment

Genetic analysis offers an important advantage to anydevelopmental system. By isolating mutations that disruptnormal development, we let the embryo tell us what genesare important and when gene function is required. If a generequired for normal development is destroyed by muta-

tion, the developmental consequences reveal the role of thegeneduring normaldevelopment.This sectionreviews howzebrafish mutants are analysed, explains how zebrafishmutants are isolated, and describes some of the interestingzebrafish genes that have been identified by mutationalanalyses.

Using cell lineage and transplantation toanalyse developmental mutants

One of the first zebrafish developmental mutationdescribed was the recessive lethal spadetail mutationFigure 3a shows how spadetail(spt) mutants got their namat the end of the first day of development, a distinctive ba

of cells has accumulated at the end of the spt mutant ta(arrow). The other prominent phenotype ofspt mutants seen in Figure 3b. In wild-type embryos, bilateral pairs osomites (which later give rise to muscle) form next to thnotochord. In contrast, spt mutants lack organized trunsomites, and later in development trunk muscle is severeldepleted. When spt mutant cells at the trunk musclposition of the gastrula fate map are labelled with lineagtracer,they adopt tail fates insteadof trunk fates. These celineage experiments suggest the defective migration osomitic precursors to the spt mutant trunk may explain thabnormal accumulation of cells in the spt mutant tail.

Lineage analyses in spt mutant embryos do not addres

whether the behaviour of mutant cells is intrinsic oextrinsic to the mutant cells. To explain these termconsider the following analogy. Suppose a car is travellinerratically down a road off in the distance. Just bwatching, we cannot tell whether the car is movinerratically because of extrinsic factors (e.g. because throad is in poor condition) or because of intrinsic factor(e.g. because the driver is blindfolded). How might wdiscriminate between these two possibilities? We coulobserve the car travel on a road we knew to be in goodcondition. If the car still drove erratically, we would deducthat the original road was in good condition and that thdriver was impaired. If the car now drove flawlessly, w

would deduce the opposite, that the driver was perfectlcompetent, but the original road was in need of repair. Thsame sort of experiment can be done with spt mutant celby creating a genetic mosaic embryo that contains botwild-type and mutant cells. When a few spt mutantgastrulcells are transplanted among trunk somitic precursors of wild-type host gastrula, they do not make somitic tissuebut instead they migrate abnormally during gastrulatioand end up at the tip of the wild-type hosts tail. Thubecause spt mutant cells behave abnormally even whethey are surrounded by wild-type cells, we say that spt actintrinsically, or cell-autonomously, in trunk somitprecursors. Even if the molecular identity of the spt gen

is not known, these transplantation experiments suggesthe type of molecule spt might encode. Examples ointrinsic (cell-autonomous) factors are transcription factors, intracellular structural or signalling molecules, or cesurface receptors. These types of molecules, if expressed athe expected time and place, would be good candidates fothe spt gene. Examples of extrinsic (non-cell-autonomousfactors are secreted signalling molecules or extracellulamatrix proteins.




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Figure 3 Wild-type and spadetail(spt) mutantzebrafish embryos. (a)Wild-typeand sptmutantembryosat 24 h. Arrowmarks theabnormalaccumulatioof cells inthe spttail bud. (b) High-magnification dorsalviews of wild-typeand sptmutantembryos at 14 h; sptmutants have a notochord, butclearly lasomites.




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It should be clear from this discussion of the spt gene thatstudying the defects in genetic mutants can teach us anenormous amount about normal development. Studies ofspt mutant embryos demonstrate that, during gastrulation,a gene called spadetail is required intrinsically in trunksomitic precursors to ensure that they migrate to thecorrect position and adopt the correct fate. The rest of the

article discusses in more detail how developmental muta-tions are found and highlights some of the interestingclasses of mutations that have recently been discovered.

Zebrafish genetic screens

How are developmental mutations isolated? Two large-scale zebrafish screens (hunts for mutants) have beenperformed, and both screens used a traditional two-generation breeding scheme, followed by screening ofembryos in the third generation. Figure 4a illustrates thetraditional screening strategy. Male parental (P) fish are

treated with a chemical mutagen to induce mutations itheir germ line. Each mutagenized male is crossedto a wildtype female to generate F1 lines; individual fish in each Fline carry different mutations inherited from their fatheFor simplicity, only a single mutation carried by a femalF1 fish is shown to be segregating. If these mutations arrecessive, no mutant phenotype will be observed since th

wild-type allele will mask the effect of the mutant recessivallele. Sibling matings between F1 fishgenerate F2 familiein the F2 generation, about half of all fish in each familcarry the same mutation. Now, crosses of F2 siblings wireveal the mutant phenotype when both the female anmale parents carry the same mutant allele. If the mutatiois recessive,then 25% of the F3 progeny of such a cross widisplay the mutant phenotype and 75% will be phenotypically normal.

A variation of this traditional approach utilizes genetitrickery to generate gynogenetic haploids during thscreening step. As the name implies, gynogenetic haploid

P

(a) Traditional screening strategy

F1

+/+

+/m +/+

F2 +/+ +/+

+/m +/+

+/+ +/m

+/m +/m

Random matings

F3

+/+ +/+ +/+ +/m +/m +/+ +/m +/m

(b) Haploid screening strategy

+/+

+/m

mutagen mutagen

UV-irradiatedsperm

Haploid embryos

Figure 4 Zebrafish mutagenesis screening strategies. (a) Traditional screening strategy. (b) Haploid screening strategy. +, Wild-type allele; m, mutanallele. Red embryos represent mutant diploids (m/m) or haploids (m).




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are embryos that have half the normal number ofchromosomes, all of which are derived from the mother.The haploid screening strategy, illustrated in Figure 4b,reduces the time and space required for screening. Howdoes haploid screening work?

As in the traditional strategy, male parental (P) fish aretreated with a chemical mutagen to induce mutations.

Mutagenized males are crossed to females to generate amutagenized F1 line. Now the procedure diverges from thetraditional approach. Instead of performing sibling crossesto generate F2 families, the haploid progeny of F1 femalesare examined directly for mutant phenotypes. Eggs aregently expressed from F1 females and mixed with spermthat has been treated with ultraviolet irradiation to destroyits genetic material. The irradiated sperm triggers devel-opment of the egg, but does not contribute any geneticmaterial to the developing haploid embryo. If the F1 femalecarries a recessive mutation that affects embryonic devel-opment, the mutant phenotype will be observed in about50% of her haploid F2 progeny. Many embryonic

structures, especially during early embryogenesis, can beobserved in haploids. However, haploid embryos live foronly about three days, and therefore haploid screens formutations that affect late embryogenesis are not feasible.Additionally, development of haploid embryos is notcompletely normal. The haploid body is short, the brain is

abnormally sculptured, and blood circulation is abnormaDespite these potential drawbacks, many interestindevelopmental mutants have been discovered using haploid screening strategy.

Zebrafish developmental mutants

What types of developmental mutations are discovered imorphological screens? Hundreds of mutations have beeidentified that affect virtually every aspect of earldevelopment. The phenotype caused by a mutation in thspadetailgene is shown in Figure 3. Figure 5 illustrates otheexamples of mutations that have been discovered imorphological screens. Embryos homozygous for a mutation inthe notailgene have no tail(as the name implies) analso lack the notochord, the embryonic backbone. Thgene disrupted by the no tail mutation is a transcriptiofactor that is conserved across animal phyla, and it turnout that mice homozygous fora mutation in a homologou

gene (called Brachyury) have a very similar phenotype. Socalled cyclops mutant embryos are cyclopic (having oncentrally located eye) and also lack the floor plate, distinct population of ventral spinal cord cells located jusabove the notochord. Like cyclops mutant embryoembryos homozgous for a mutation in the one-eyepinheadgene also are cyclopic and lack a floor plate. Th

Figure 5 Zebrafish mutants at one day of development. The notochord (arrowhead) and an eye (open arrowhead) are indicated on the wild-typeembryo. Theother eyein thewild-type embryois in another plane of focus. Seetext fordescriptions of mutants. Theboxedinset in thelowerright show

high-magnification views of wild-type(wt) and no isthmus(noi) mutantembryoheads, showing thelackof themidbrainhindbrainboundary(arrow)in nisthmusmutants.




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one-eyed pinheadgene has recently been shown to encode anovel protein, illustrating that genetic screens in zebrafishwill uncover new developmental genes. Another exampleof a mutation affecting brain development is the no isthmusmutation. Embryos lacking a functional no isthmus genelack the isthmus, a structure that demarcates the divisionbetween the midbrain and the hindbrain and later becomes

the cerebellum (see Figure 5 inset).Embryos with the no isthmus mutation are more subtle

than the other mutants shown in Figure 5. How might onescreen easily for even more subtle defects? One way to dothis is to screen directly for disruptions in gene expressionpatterns. Figure6 shows wild-type andmutantembryos thatwere fixed and processed to detect the transcripts of threegenes that have nonoverlapping, region-specific expressionpatterns. Two of these genes are expressed in discreteregions of the midbrain and hindbrain; engrailed3 (eng3) isexpressed in the midbrainhindbrain boundary, andkrox20 is expressed in two hindbrain segments (3 and 5).Comparing a wild-type embryo with an embryo carrying a

mutation in the valentino gene shows that valentino mutantembryos have normal krox20 expression in hindbrainsegment 3, but virtually no krox20 expression in segment 5.This strongly suggests that the valentino mutation is not inthe krox20 gene itself, but instead is a mutation in a genethat regulates krox20 expression specifically in hindbrainsegment 5. Thus, one important feature of the expressionscreening approach is that it allows isolation of regulatorygenes that control expression of downstream genes.Another important feature is that it allows identificationof genes that control subtle patterning of the embryo. Livevalentino mutant embryos do have a morphologicallyidentifiable defect, but it is a subtle defect in hindbrain

segmentation that can be picked out only by an experi-enced eye. On the other hand, even an untrained eye cansort out valentino mutant embryos from their wild-typesiblings using gene expression patterns!

We have discussed only a few of the hundreds ofmutations that have been described that disrupt zebrafishembryonic development. It is beyond the scope of this

article to review them all, and instead we briefly mentiothe classes of mutations that affect virtually all aspects oembryonic development. These classes include mutationthat disrupt gastrulation movements, establishment of thbody axes, tissue and organ formation (including thnotochord, blood, heart, neural crest derivatives,placodeand central nervous system), formation of neural connec

tions, and behaviour. Genetic screens using novel approaches will identify additional developmental genesince it is clear that there are still many new genes to bidentified and studied. By characterizing mutant phenotypes using cell lineage and transplantation analyses anby examining the genetic interactions among differenmutations, we can begin to understand regulatory pathways that govern embryonic development.

The zebrafish genetic map

The challenge now is to identify the molecular nature o

genes identified in mutational screens. What kind oproteins do these genes encode? Among the manpossibilities are transcription factors, secreted signallinmolecules, and cell surface receptors. An important toofor identifying the molecular identity of genes known onlby mutation is a genetic linkage map. A genetic linkagmap is constructed by examining the inheritance patternof various markers relative to one another. If two markerare unlinked (for example on different chromosomes), thethey will assort independently of one another, just aGregor Mendel observed for the various traits he followein pea plants. On the other hand, if two markers are closellinked, they will almost always be inherited together. Onl

rare recombination events will separate these two markerand the frequency of recombination indicates the genetidistance between the markers. Recently, the 25 chromosomes of the zebrafish have been ordered into 25 linkaggroups. These 25 linkage groups are collectively called thzebrafish genetic map.

How can one use the genetic map to identify thmolecular nature of a gene known only by mutationFirst, the mutation is placed on the genetic map using thsame recombinational analysis described above. Thmutation will be linked only to markers that are on thsame chromosome and will be unlinked to markers oother chromosomes. Recombinational frequency wi

indicate which markers are most closely linked to thmutation. Sometimes, the most closely linked marker wibe a gene that is a good candidate for the one that idisrupted by the mutation. More frequently, the mutatiowill map to a chromosomal region in which no candidatgenes have yet been mapped. In this case, the most closellinked marker can be used to walk along the chromosomtowards the gene of interest. Once a good candidate geneidentified, either by serendipity or by chromosomawalking, several criteria are used to determine whethe

Figure 6 Mutagenesis screening using gene expression patterns. Wild-

type and valentino mutant embryos fixed at 22 h and processed to detectexpression of three different genes. Comparing head views of wild-type

and mutant embryos shows that engrailed3 (eng3) expression at themidbrainhindbrain boundary and krox20 expression in hindbrainsegment 3 is normalin valentino mutantembryos,but krox20 expressionin

hindbrain segment 5 is severely diminished (*). (Photographs courtesy ofDr Cecilia B. Moens.)




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the candidate gene is indeed the gene disrupted by themutation. First, recombinational analysis should showthat the candidate gene and the mutation always segregatetogether and are never separated by a recombinationalevent. Second, the gene should be expressed at the righttime and place to explain the defects observed in mutantembryos. Third, sequencing of the candidate gene from

mutant embryos should reveal a molecular lesion in thegene that disrupts its function. Finally, the mutantphenotype may be rescued by providing mutant embryoswith a wild-type copy of the candidate gene. When thesecriteria are satisfied, one can say with confidence that thecandidate gene is indeed the same gene identified bymutation. To date, the zebrafish genetic map has beenutilized to identify the molecular nature of several genesoriginally identified by mutation, and the genetic map willcontinue to be an important tool in the future.

SummaryWe have reviewed the features of the zebrafish embryo thatmake it an excellent vertebrate model system. Zebrafishembryos are small, virtually transparent creatures thatdevelop rapidly and synchronously outside the mother.Cell lineage andtransplantation analyses allow us to followthe behaviour of individual cells over time in both wild-type and mutant embryos. The relative ease of geneticscreens in zebrafish has allowed the identification ofhundreds of genes that regulate embryonic development,only a few of which were described here. Finally, the

establishment of molecular and genetic tools such as thzebrafish genetic map has allowed us to identify thmolecular nature of genes known only by mutationTogether, these attributes of the zebrafish allow us tdissect embryonic development at cellular, genetic anmolecular levels. By combining developmental studies izebrafish with those in other vertebrate model systems

developmental biologists will begin to unravel the extraordinary events that occur as an embryo develops from one-celled zygote to a complex, multicellular organism.

Further Reading

Development (1996) vol. 123. [Many articles in this issue contain initi

phenotypic descriptions of zebrafish developmental mutants found

two large-scale screens.]

Driever W, Stemple D, Schier A and Solnica-Krezel L (1994) Zebrafis

genetic tools for studying vertebrate development. Trends in Geneti

10: 152159.

Eisen J S (1996) Zebrafish make a big splash. Cell87: 969977.Kimmel CB, Kane DA and Ho RK (1991) Lineage specification durin

earlyembryonic development of the zebrafish.In: Gerhart J (ed.)Cel

Cell Interactions in Early Development, pp. 203225. New Yor

Wiley-Liss.

Kimmel CB, Ballard WW, Kimmel SR, Ullmann B and Schilling T

(1995) Stages of embryonic development of the zebrafish. Develop

mental Dynamics 203: 253310.

Postlethwait JH and Talbot WS (1997) Zebrafish genomics: fro

mutants to genes. Trends in Genetics 13: 183190.

Solnica-Krezel L, Stemple D and Driever W (1995) Transpare

things}cell fates and cell movements during early embryogenesis

zebrafish. BioEssays 17: 931939.



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Zebrafish Embryo as a Developmental System.pdf