marks by those studying the structure ofeggs.

Once the nuclei (pronuclei) of egg andsperm have combined, cell division in a fer-tilized egg or zygote is by mitosis — the divi-sion of single nuclei into two new nuclei,each containing a complete copy of theoriginal cell’s chromosomes. A fertilized eggdivides mitotically into many smaller cells, ina cleavage process that is marked early on bythe formation of a cleavage furrow at the sur-face of the egg.This furrow is usually precise-ly placed above where the genetic material isseparating, ensuring that each daughter cellreceives identical sets of chromosomes.

Gardner3 was the first to seriously ponderthe use of the mouse egg’s polar body as anavigational beacon. He and others8 typical-ly found the polar body to be located in thefirst cleavage furrow. With Hiiragi andSolter’s report, the reliability of polar-bodypositioning as a determinant of the cleavageplane is questioned. They report thatalthough many mouse eggs divided within30� of the second polar body, many did not.So they doubt whether the second polar bodyis a fiduciary mark of the cleavage plane, andwhether unfertilized mouse eggs even havean animal–vegetal axis.

Since first witnessing the frog egg’s ‘greycrescent’, which forms on the freshly fertil-ized egg’s pole opposite to the sperm-entrypoint, embryologists recognized that gastru-lation — the stage at which cell division andmigration start to form a complex structurewith a primordial central gut tube — laterstarts at this region.Piotrowska and Zernika-Goetz4 reported that in mice the meridiancreated by the sperm entry point specifiedthe second embryonic axis. Hiiragi andSolter, again using time-lapse ‘ovo-vision’ totrace the relations between the fertilizationcone (the structure around the sperm entrypoint) and first cleavage, report that thismeridian is a less accurate predictor of cleav-age than the orthogonal orientation of theapposed sperm and egg pronuclei beforethey fuse into a single nucleus followingfertilization.

For their ultimate experiment, Hiiragiand Solter scrambled mouse eggs by cloningearly-stage male or female pronuclei into fer-tilized eggs from which they had previouslyremoved the male pronucleus. Remember,the original question is whether an unfertil-ized egg has pre-existing asymmetry orwhether the sperm entry site defines a criticalmeridian. So the rationale here is that trans-ferring an early-stage male pronucleus mightcarry with it a signal for cleavage determina-tion. Again, the orientation of the apposedmale and female pronuclei was the mostaccurate predictor of the first cleavage plane,because this axis defines the axis of the firstmitotic apparatus, with cytokinesis — thedivision of cytoplasm that follows nucleardivision during mitosis — occurring

orthogonal to this structure.Early cell biologists appreciated the

importance of the pronuclei’s journey. Thestarting position of the egg’s meiotic spindle,the site of female-pronucleus formation andthe sperm entry point all influence the trajec-tories of pronuclear migrations and the ulti-mate axis of pronuclear apposition.Cleavageoccurs orthogonal to the mitotic spindle’saxis and pronuclear positioning, just beforebreakdown of the nuclear envelope depositsthe centrosomes, crucial intercellular archi-tecture that helps to organize chromosomes,at their ultimate positions when the mitoticspindle assembles. Consequently, the trajec-tories of the sperm’s entrance and the cyto-plasmic migration of the two pronuclei, cul-minating in their apposition near the eggcentre,will influence their final positions justbefore mitosis.

So perhaps the polar body is not alwaysthe precise marker of the egg’s ‘North Pole’, inthe way that magnetic north is off-axis fromgeographic north. And perhaps the spermentry point, especially if restricted from thearea over the meiotic spindle, provides onlytransient indications of directionality andnot long-term reliability in that respect.

It could of course be that the strategies forobserving this subtle, easily perturbed signalinfluence the experimental results and there-by undermine the conclusions drawn fromthem. A decade’s worth of evidence comesfrom skilled teams, each repeatedly havingperformed exacting experiments. But, atleast to us, each experimental set seems in-fluenced by the second polar body’s own

predilection to move into the furrow, as wellas the strength of its tether to the egg surface— all potentially influenced by the egg’scompression during imaging. Elegant stud-ies injecting oil or fluorescence markers intothe egg,or examining the movement of cyto-plasm, are questioned by non-believers asartefacts,as are results from studies involvingpronuclear extractions and subsequentnuclear fusions.

The results of Hiiragi and Solter2 will forcerenewed attention on the natural forces thatgenerate the embryo’s three-dimensionalcomplexities,and perhaps less invasive exam-inations of the processes involved.Meantime,we remain fascinated by the polarized viewson polarity and polar bodies. ■

Gerald Schatten is in the Department of Obstetrics,Gynecology and Reproductive Sciences, University of Pittsburgh School of Medicine, 204 Craft Avenue,Pittsburgh, Pennsylvania 15213, USA.e-mail: gschatten@pdc.magee.eduPeter Donovan is in the Institute for CellEngineering, and the Department of Gynecologyand Obstetrics, Johns Hopkins University,Broadway Research Building, Suite 559,Baltimore, Maryland 21205, USA.e-mail: pdonova4@jhmi.edu1. Spemann, H. Forschung und Leben (Engelhorn, Stuttgart, 1943).

2. Hiiragi, T. & Solter, D. Nature 430, 360–364 (2004).

3. Gardner, R. L. Development 124, 289–301 (1997).

4. Gardner, R. L. Development 128, 839–847 (2001).

5. Piotrowska, K. & Zernicka-Goetz, M. Nature 409, 517–521

(2001).

6. Piotrowska, K. & Zernicka-Goetz, M. Development 129,

5803–5858 (2002).

7. Plusa, B. et al. Nature Cell Biol. 4, 811–815 (2002).

8. Weber, R. J., Pedersen, R. A., Wianny, F., Evans, M. J. &

Zernicka-Goetz, M. Development 126, 5591–5598 (1999).

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Plant biology

Good neighboursLaura Serna

Plants depend on structures called stomata to regulate gas exchangewith the air, and their positioning is crucial. A key factor controllingstomatal development and arrangement has now been found.

We all need good neighbours, butplant guard cells need them morethan most. Destined to remain

where they are produced, these cells cannotfunction efficiently without help from adja-cent cells. Guard cells are arranged in pairsto create stomata, and occur mainly on thesurfaces of leaves; they surround a porethrough which the plant exchanges gaseswith the atmosphere. The pore aperture isregulated by changes in the rigidity of theguard cell, and this in turn is controlled bythe flow of water and ions between theguard cells and their neighbouring non-guard cells. Now, writing in Science,Bergmann and co-workers1 report that arecently identified enzyme in the floweringplant Arabidopsis thaliana2, christened

YODA (YDA), is crucial to the formationand arrangement of stomata.

Mutations in the YDA gene result in aplant in which almost all the cells at theplant surface are guard cells. As might beexpected, this overproduction of stomatahas severe consequences, and many of themutant seedlings die, while those that sur-vive produce small plants with sterile flow-ers. Mutations in three other genes — thecatchily named TOO MANY MOUTHS(TMM)3, STOMATAL DENSITY AND DIS-TRIBUTION1 (SDD1)4 and FOUR LIPS(FLP)3 — also affect stomata, producingstomatal clusters, but also allowing the for-mation of single stomata. The single stoma-ta ensure correct gas exchange and so thesethree mutants develop normally. But how

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does YDA prevent such overproduction ofstomata?

In Arabidopsis, guard-cell formation isthe final step in a series, or lineage, of celldivisions that leads to the formation of theleaf surface5 (Fig.1a).This process starts witha precursor cell called a meristemoid mothercell (MMC), which divides to form a smallmeristemoid and a larger pavement cell. Themeristemoid is a self-renewing cell, but aftera few divisions it loses this stem-cell activityand becomes a guard mother cell (GMC).The GMC then divides to form a pair ofguard cells. In yda mutants, in which YDAactivity is lacking, increased numbers of cellsseem to become MMCs, which prematurelyform stomata because the meristemoids thatare produced assume a GMC identity. Thissuggests that YDA prevents stomata over-production by reducing the number of cells

entering the stomatal lineage and controllingthe balance between meristemoid renewaland stomata formation. Further studies mayconfirm these functions and might alsoidentify new ones.

The protein encoded by YDA appears tobe a MAPKK kinase, an enzyme that addsphosphate groups to proteins, thus regulat-ing their activity.The putative YDA protein isvery similar to members of the STE11 familyof such kinases2. MAPKK kinases areupstream components of so-called kinasesignalling cascades, which link cell-surfacereceptors to critical regulatory targets withincells6. When not required, the enzymaticactivity of such kinases is suppressed by aregulatory region of the enzyme molecule —the amino-terminal domain6,7. Thus, YDAshould be continuously switched on inplants expressing constructs of the enzyme

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in which the N-terminal regulatory domainhas been deleted (�N-YDA plants).Striking-ly, Bergmann and colleagues1 find that �N-YDA plants totally suppress guard-cell for-mation. The authors infer from this thatentry into the stomatal lineage requires areduction in YDA activity. Deletion of thisregulatory region also causes a loss of sig-nalling specificity6, but the characteristics ofthe yda mutants indicate that the absence ofstomata in �N-YDA plants is due to the con-tinuous activation of the YDA pathway.

SDD1/TMM and YDA may share at leasttwo functions — regulating entry into thestomatal lineage and maintenance of themeristemoids’ stem-cell activity. Do they actin the same genetic pathway? Bergmann et al.show that a single copy of the �N-YDA gene,which has no effect in wild-type controlplants, can restore normal stomatal produc-tion in both tmm and sdd1 mutants. Theauthors argue that YDA acts either down-stream of SDD1/TMM or in a parallel path-way. The structure of these molecules seemsto suggest the former possibility,and that thereceptor protein TMM might react with amolecule processed by SDD15,8, thereby acti-vating YDA (Fig. 1b). Because TMM has nokinase domain linking it to the cell interior,it might need to dimerize with a co-receptorthat would provide such a domain5,8. In fact,the presence of paired cysteine amino acidsin TMM suggests that it forms disulphidebridges with a partner5,8.The resulting recep-tor complex would transduce the signal fromthe cell surface to the cell interior,where YDAis activated.

The ability to generate plants with con-trasting features — yda, in which all thesurface cells are guard cells, and �N-YDA,completely lacking guard cells — provides anopportunity to uncover genes that might reg-ulate stomatal development. Bergmann et al.take matters a step further by comparing allthe genes expressed in yda and �N-YDAseedlings. They find that the expression of111 genes is reduced in �N-YDA andincreased in yda.As might be expected,TMMand SDD1, which are expressed in stomatalprecursor cells or stomata8,9, fall into thisgroup. Bergmann et al. also find that theexpression of a further 109 genes is increasedin �N-YDA and reduced in yda mutants.Downstream targets of YDA should be in thisgroup. Because MAPKK kinases usually actin more than one signalling cascade, therebyregulating several plant features6, only someof these genes should control stomatalformation and arrangement.

But that’s not all. A recent study of Ara-bidopsis identified mutants for 74% of theplant’s genes10, which should greatly acceler-ate study of the function of the 220 genesidentified by Bergmann et al. The authorshave followed this course and find that muta-tions in a gene called FAMA produce clustersof early-stage stomata that are reminiscent of

Figure 1 YODA and the development of stomata. a, Stomatal development begins with a meristemoidmother cell (MMC), which divides to produce a meristemoid (M) and a pavement cell. Meristemoidsare self-renewing cells, producing new meristemoids. After a number of cell divisions they developinto guard mother cells (GMCs), which then divide to produce two guard cells (GCs). Bergmann etal.1 suggest that YODA (YDA) promotes the MMC identity and maintains the stem-cell activity of themeristemoids. TMM and SDD1 also regulate such functions. The authors propose that FAMAcontrols GMC identity and GC differentiation, functions shared with FLP. b, YDA acts downstreamof SDD1/TMM or in a parallel pathway. Left, an SDD1-processed molecule might activate a TMM/co-receptor complex. The hypothetical co-receptor would provide TMM with a cytoplasmic domain,which it lacks. Activation of this receptor complex would activate YDA, triggering a transductionpathway that initiates the stomatal-cell lineage and maintains the meristemoids’ stem-cell activity.Right, activation of YDA might depend on an as-yet-unidentified receptor, and an SDD1/TMM/co-receptor might trigger a signal-transduction pathway independently of YDA. Both pathways wouldcontrol the two functions above.

M GMC

YDASDD1TMM

M GMC GC

MMC

FLPFAMA

FLPFAMA

YDASDD1TMM

MMC identityStem-cell fate of M

MMC identityStem-cell fate of M

SDD1-processed molecule SDD1-processed molecule

Extracellular

YDA YDA

b

a

TMM TMM

Nucleus Nucleus

or

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coherent scattering of the Sun’s radiation atlast became fully accessible with the intro-duction a decade ago of the Zurich ImagingPolarimeter, ZIMPOL7, with which thepolarimetric noise could be dramaticallyreduced. Using ZIMPOL, the electro-opti-cally modulated polarization signal is‘demodulated’ into four image planes,whichcorrespond to the different polarizationstates needed to form the full Stokes vector(which contains the complete polarizationinformation).

With the polarimetric accuracy of onepart in 105 that is routinely reached withZIMPOL, combined with high spectralresolution, an astounding variety of spectralstructures can be seen throughout the wholesolar spectrum. This linearly polarizedspectrum — which has been called the‘second solar spectrum’, because it bears solittle resemblance to the ordinary intensityspectrum — is a veritable treasure trove ofall kinds of coherence phenomena8. Thedifferent polarized structures in the secondsolar spectrum are affected to variousdegrees, through the Hanle effect, by the‘hidden’ magnetic fields in the solar atmos-phere. Differential effects can then be usedas diagnostics.

However, the proper quantitative inter-pretation of the Hanle signatures in the solarspectrum is a tricky business. The polariza-tion amplitudes observed unfortunatelydepend on details of how the spectral line isformed — including the details of the struc-ture of both the atom/molecule and of thesolar atmosphere, as well as the way in whichthe polarized radiation is transportedthrough the atmosphere. Trujillo Bueno etal.1 have brought the theory of line forma-tion to a new level: they have modelled theHanle effect for spectral lines from atomsand molecules using a three-dimensionalradiative-transfer code and a model of theSun’s atmosphere obtained from simula-tions of the star’s surface convection. Theyfind much higher magnetic-energy densitiesthan in previous investigations, bothbecause of their more realistic three-dimen-sional approach and because they introducea more realistic probability distributionfunction for the turbulent field strengths,rather than using a single-value field.

This work is a pleasing example of howdifferent areas of astrophysics can bebrought together to achieve a scientificobjective: Trujillo Bueno et al. have com-bined high-precision spectro-polarimetry,coherency effects in atomic physics,advanced techniques in radiative-transfertheory and simulations of magnetoconvec-tion. The information that can be retrievedabout the Sun’s magnetism is, however,model-dependent,and will remain so for theforeseeable future, because the small-scalemagnetic structures in question will not beresolved even with the next generation of

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the stomatal clusters seen in the flp mutant3.One exciting consideration,however,has notbeen emphasized here. FAMA belongs to afamily of gene-transcription factors thathave a ‘basic helix–loop–helix’ structure1

and act with MYB proteins to regulate sever-al processes in Arabidopsis development11.Will the long-awaited FLP gene encode (orregulate) an MYB protein?

Bergmann and colleagues’ study not onlyuncovers a new function for YDA,it also pro-vides a powerful tool for identifying genesthat control stomatal development and pat-tern formation.Further studies are needed todefine the gene’s function and to identity thecomponents of the YDA cascade. Undoubt-

edly, we have a long but exciting way to go,but also the tools to take us there. ■

Laura Serna is in the Facultad de Ciencias delMedio Ambiente, Universidad de Castilla-LaMancha, E-45071 Toledo, Spain.e-mail: laura.serna@uclm.es 1. Bergmann, D. C., Lukowitz, W. & Somerville, C. R. Science 304,

1494–1497 (2004).

2. Lukowitz, W. et al. Cell 116, 109–119 (2004).

3. Yang, M. & Sack, F. D. Plant Cell 7, 2227–2239 (1995).

4. Berger, D. & Altmann, T. Genes Dev. 14, 1119–1131 (2000).

5. Serna, L. & Fenoll, C. Trends Genet. 18, 597–600 (2002).

6. Hirt, H. Proc. Natl Acad. Sci.USA 97, 2405–2407 (2000).

7. Asai, T. et al. Nature 415, 977–983 (2002).

8. Nadeau, J. A. & Sack, F. D. Science 296, 1697–1700 (2002).

9. Groll, U. V. et al. Plant Cell 14, 1527–1539 (2002).

10.Alonso, J. M. et al. Science 301, 653–657 (2003).

11.Schiefelbein, J. Curr. Opin. Plant Biol. 6, 74–78 (2003).

Solar physics

Hidden magnetismJan Olof Stenflo

Observations of the Hanle effect have revealed the existence of small-scale ‘hidden’ magnetic flux on the quiet Sun. The magnetic-energydensity of this hidden flux is much larger than previously thought.

Magnetic fields have occupied centrestage in solar physics for the pastseveral decades, and have come to

be regarded as the key ingredient for aunified understanding of solar phenomena.It may therefore come as a surprise that,after all these years, the magnetic-energydensity in the solar atmosphere mighthave been seriously underestimated — asTrujillo Bueno et al.1 conclude on page 326of this issue.

The spectrum of radiation from the Suncan be resolved into a series of lines corre-sponding to different atomic transitions. Inthe presence of a magnetic field, these spec-tral lines can split into multiple polarizedcomponents — this is the Zeeman effect andit has been used to diagnose the magneticfields on the Sun since it was first intro-duced2 to astrophysics by George Ellery Halein 1908. Ever smaller magnetic structureshave been revealed, and there is no end insight. In fact, the magnetic flux seems tohave a fractal structure, with an almostscale-invariant, self-similar pattern3 (Fig.1).According to theoretical predictions basedon magnetoturbulence, the structuringshould continue for several orders of mag-nitude beyond the scales that have so farbeen resolved.

But the Zeeman effect as a diagnostic toolis ‘blind’ to magnetic fields that are tangledon scales too small to be resolved. Below thescale of the achievable angular resolution,the contributions to the Zeeman-effectpolarization from opposite-polarity compo-nents within a tangled magnetic field canceleach other; so an unresolved mixed-polarityfield leaves no ‘footprint’in the Zeeman-split

spectral lines. For this reason, a vast amountof solar magnetic flux has possibly remainedhidden from view.

Trujillo Bueno et al.1 have used a differentapproach. The alternative tool is the Hanleeffect, a coherence phenomenon discovered4

by Wilhelm Hanle in 1924. This effect was ofgreat significance in the early developmentof quantum mechanics, because it demon-strated the principle of the coherent super-position of quantum states, and the natureof decoherence when the degeneracy of thequantum states is partially lifted as weakmagnetic fields are introduced5. In the con-text of solar physics, the Hanle effect refers tothe set of polarization effects that are causedby the coherent scattering of the radiation inthe Sun’s atmosphere under the influence ofan external magnetic field. The symmetryproperties of the Hanle effect with respect tothe orientation of the magnetic field areentirely different from those of the Zeemaneffect.Thus an unresolved, tangled magneticfield leaves a polarimetric footprint for theHanle effect, while being invisible to theZeeman effect.

Although the Hanle effect was intro-duced decades ago as a tool to gain informa-tion on the elusive, turbulent solar magneticfield6 (and a lower limit of 10 gauss on theturbulent field strength was determinedstraight away), further progress was limitedby the insufficient polarimetric precision ofthe instruments available. The polarizationamplitudes sought are small (typically atthe level of 0.1% or less), because theanisotropy of the radiation field in the solaratmosphere is so small. But the wealth ofpolarization phenomena caused by the

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