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Some time between five and six millionyears ago, the Mediterranean Seabecame isolated from the Atlantic

Ocean. In consequence some areas dried out— hence the title of Kenneth Hsü’s book TheMediterranean was a Desert 1 — and largesalty lakes recharged by rivers flowingthrough deep canyons replaced the previ-ously marine basins. During this time, theremaining bodies of water were either toosalty or not salty enough for normal marinefauna to flourish. This was the so-calledMessinian salinity crisis. Because Messiniansediments are essentially devoid of marinefossils this dramatic geological event hasbeen difficult to date or place accurately inthe global stratigraphic record, meaning inturn that its causes and environmentalimpact have been unclear.

Beginning in 1833 with Charles Lyell’s

formal subdivision of geological strata intorelative periods, based on the percentage offossil shells representing living specimensfound in the respective formations2, marinefossils in sedimentary sequences have beenthe fundamental tool used to help imposechronological order on the stratigraphicrecord. Without marine fossils, the precisecorrelation of the Messinian salinity crisiswith the global stratigraphic timescale hasbeen an elusive goal. The significance ofMessinian events can be judged from theestimate that as much as 6% of the ocean’ssalt was transformed into giant deposits lefton the Mediterranean sea floor by evapora-tion3.

Now, the application of the relatively newstratigraphic technique of astrochronology,as reported by Krijgsman et al. on page 652 ofthis issue4, has enabled accurate dating of

these fossil-poor sediments. Alterations inEarth’s obliquity and distance from the Sunoccur in regular cycles, and the resultingchanges in climate are reflected in the con-tents of sedimentary deposits. Astrochronol-ogy involves time-series analyses of a sedi-mentary sequence as related to these astro-nomically forced cycles. When supportedby biochronology and magnetostratigraphy,the approach allows for an unprecedentedprecision in the definition of geological time,and consequently for an equally high-preci-sion worldwide correlation of the Messinianevents with other sedimentary sequencesdeposited during this period of Earth’shistory.

In the 1950s, geologists working in south-ern Europe were already beginning to recog-nize the immensity of the Messinian salinitycrisis because of the unusual stratigraphicpositioning of the evaporitic deposits, whichare sandwiched between deep-water marinesediments, composed of clay and calciumcarbonate, known as marls. In contrast to thesalt minerals of the evaporites, which wereprecipitated from hypersaline waters, thesemarls represent deposition under normalopen-marine conditions. The upper transi-tion from continental evaporites to deep-seasediments is particularly abrupt and can beexplained only as the result of a huge floodin which Atlantic water poured across theGibraltar arc and into the Mediterraneanbasin (Fig. 1).

This transition to full marine conditionswas designated as the boundary between theMiocene and Pliocene epochs — which,dated at 5.33 million years (Myr) ago, is amajor marker in the geological calendar5.The evaporitic deposits of the ‘Gessoso-solfifera’ formation in Sicily were taken asdenoting the Messinian stage of geologicaltime6. But defining these stratigraphic unitsor stratotype localities in Sicily, or for thatmatter anywhere else in the Mediterranean,violated long-held stratigraphic principles,because a continuous marine sequencestraddling the Miocene–Pliocene boundaryexists nowhere in the region. A stratigraphiccontroversy was thus born7.

That controversy, however, has not beenlimited to the problems of stratigraphiccorrelation. With the initiation of scientificocean drilling, two expeditions of the DeepSea Drilling Program (DSDP) in theMediterranean showed that the Messinianevaporitic deposits were not restricted to theshallow marginal basins, where geologistshad studied land sections. Astonishingly,they were widespread in all of the deeperMediterranean basins, as imaged by basin-wide seismic profiles and sampled bydrilling8,9.

With the publication of these excitingresults, debate over the Messinian salinitycrisis erupted into a full-blown scientific fracas, with opposing camps developing

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From desert todeluge in theMediterranean Judith A. McKenzie

A time when the Mediterranean became cut off from, then reconnectedto, the Atlantic has been securely dated. An approach known asastrochronology lies behind this achievement.

Figure 1 “The Filling of theMediterranean Sea” byGuy Billout. Thiswhimsical cartoon, firstpublished in The AtlanticMonthly and mixingmodern promenaders atthe Rock of Gibraltar withan event that happened5.33 million years ago,depicts the catastrophicflood that ended theMessinian salinity crisiswhen Atlantic waters re-entered theMediterranean.

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radically contrasting views and modelsabout the environmental conditionssurrounding the evaporite deposition7.Now, almost 30 years after the first DSDPdiscovery, the combatants may be able toreach an accord. Krijgsman and colleagues’application4 of astrochronology permits anew reading of the complex stratigraphicrecord produced by the Messinian events.

In my view, this work resolves severalissues. First, as I have mentioned, themethod sidesteps the need for marine fossilsto date the Messinian sequences. Second, thenew chronology shows that the beginningand end of the Messinian salinity crisis weresynchronous throughout the Mediter-ranean; Krijgsman et al. date the beginningat 5.96 Myr and the end at 5.33 Myr ago. Thecauses and consequences of events can thusbe distinguished. It is now clear that a tecton-ic closure in the general region of the modernStraits of Gibraltar, possibly augmented bya fall in sea level, led to the synchronous,basin-wide onset of evaporative conditions.The initial evaporative draw-down of waterin the Mediterranean would have been grad-ual, with the development of both shallow-water and deep-water evaporitic sequencesdepending on the size and shape of individ-ual basins. Many of the arguments putforward by the proponents of the variousmodels can be accommodated within thispicture.

Regardless of that, the massive flood thatpermanently ended evaporative conditionsin the Mediterranean must have been causedby a dramatic rise in sea level. Such a risewould undoubtedly have had consequenceselsewhere in the world. One example mayhave been the major flooding of the BahamasPlatform at around the Miocene–Plioceneboundary, which led to a huge ‘back-

stepping’ of the reef system to a higher, morecentral location on the platform10.

But what can explain the large erosionalsurfaces and deep canyons cut into the seafloor that are observed in parts of theMediterranean8,9? The extraordinary exis-tence of very deep, dried-out basins duringthe Messinian remains an essential elementof any such explanation, and our under-standing will have a large gap in it as long asthe complete sedimentary sequences in thedeepest Mediterranean basins remain unex-plored. It is only in the very deepest basinsthat sedimentation could have continueduninterrupted during the salinity crisis.Securing the necessary samples to fill in themissing period between 5.59 and 5.50 Myrago will require advanced drilling technolo-gy to penetrate through the thick evaporitesequences to determine what lies below.Until this final ‘Messinian gap’ is closed, thegiant salt deposits of the Messinian salinitycrisis will remain a theme for speculationand argument. Judith A. McKenzie is at the Geological Institute,Swiss Federal Institute of Technology, ETH-Zentrum, CH-8092 Zürich, Switzerland. e-mail: sediment@erdw.ethz.ch 1. Hsü, K. J. The Mediterranean was a Desert (Princeton Univ.

Press, 1983).

2. Lyell, C. Principles of Geology (Murray, London, 1833).

3. Ryan, W. B. F. in Messinian Events in the Mediterranean (ed.

Drogger, C. W.) 26–38 (K. Ned. Akad. Wetensch., Amsterdam,

1973).

4. Krijgsman, W., Hilgen, F. J., Raffi, I., Sierro, F. J. & Wilson, D. S.

Nature 400, 652–655 (1999).

5. Cita, M. B. Micropaleontology Spec. Publ. 1, 1–30 (1975).

6. Selli, R. Giorn. Geol. 28, 1–33 (1960).

7. Cita, M. B. in Controversies in Modern Geology (eds Müller,

D. W., McKenzie, J. A. & Weissert, H.) 13–23 (Academic,

London, 1991).

8. Ryan, W. B. F. Init. Rep. DSDP Leg 13 (1973).

9. Hsü, K. J. et al. Init. Rep. DSDP Leg 42A (1978).

10.McKenzie, J. A., Spezzaferri, S. & Isern, A. Mem. Soc. Geol. It.

54, 93–108 (1999).

components of the V(D)J recombinationmachinery are encoded by the recombina-tion-activating genes, RAG1 and RAG2.These genes are expressed in pro- and pre-lymphocytes, coincident with the two periodsof V(D)J recombination. But if immature Bcells in the bone marrow recognize a self anti-gen, they can maintain expression of RAG1and RAG2, and alter their BCR through aprocess known as receptor editing5,6.

The RAG proteins initiate recombina-tion by recognizing and cleaving chromoso-mal DNA at specific recombination sites.Mature B and T cells do not express the genesthat encode these proteins — or so it wasthought, until RAG messenger RNA tran-scripts, protein and V(D)J recombinationactivity were detected in B-lineage cells inthe spleen, lymph nodes and tonsils7–13.The two basic aspects of these findings weredistinguished by their kinetics. First, withinone to two days, cultured B cells from thespleen or lymph nodes could be stimulated(by cytokines9,11, or by interaction withantigens13) to perform de novo immuno-globulin-gene recombination, replacing oneBCR with another. Second, RAG-expressingB cells and evidence of recombination couldbe found late in the immune response (after12–16 days) in splenic germinal centres7,10 —sites where mature B cells are activated byantigen and T cells to differentiate andsecrete immunoglobulins. These resultssupported the conclusion that mature B cellscan be induced by antigen to re-expressRAG1 and RAG2, and, by immunoglobulin-gene recombination, to generate new BCRspecificities. With them, a central pillarof the clonal-selection theory seemed tocollapse.

Much of this argument relied on theinference that expression of RAG1 and RAG2is reactivated in mature B cells after havingbeen turned off. But this had, in fact, neverbeen demonstrated, because methods fordetecting and quantifying expression ofRAG1 and RAG2 in individual, live cells werenot available. Yu et al.3 and Monroe et al.4

have now created strains of mice in which agreen fluorescent protein (GFP) is a markerfor cells that express RAG2 (and, by infer-ence, RAG1, because the two genes seem tobe coordinately expressed).

Yu et al. used a large reporter transgeneconsisting of the RAG chromosomal regionwith GFP coding sequences substituted forthose of RAG2. Monroe et al. created ‘knock-in’ mice, in which the endogenous RAG2locus directs expression of a RAG2–GFPfusion protein. Both approaches yield GFPexpression in the expected subsets of devel-oping B and T cells, but each has a limitation.Although the GFP protein encoded byYu and colleagues’ transgene is highlyexpressed, it seems to be more stable thanendogenous RAG2 protein. The result issome ‘green’ cells containing little or no

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614 NATURE | VOL 400 | 12 AUGUST 1999 | www.nature.com

Mark Twain once wrote, “The reportsof my death are greatly exaggerated”.Perhaps the same can now be said

for some aspects of ‘clonal selection’ in theadaptive immune response1,2. According tothis theory, B and T cells have a diverse arrayof antigen receptors before they encounterantigens (bacteria or viruses, for example).Those cells whose receptors recognize anti-gen are then selected to proliferate and dif-ferentiate into effector and memory cells.

The possibility that antigen-specificinteractions trigger cells to produce a newarray of receptors is specifically forbiddenby the clonal-selection theory. Yet a seriesof surprising studies has implied that, inmature B cells, assembly of antigen-receptorgenes can be reactivated in response to anti-

gen. Now, reports by Yu et al.3 (on page 682 ofthis issue) and Monroe et al.4 (in Immunity)describe a powerful new tool that shouldallow a more sophisticated analysis of B- andT-cell development, and indicate how theprevious findings might be reconciled withthe clonal-selection theory.

The development of B and T cells is orga-nized around V(D)J recombination — theassembly of immunoglobulin and T-cellreceptor (TCR) genes from component V(variable), D (diversity) and J (joining) genesegments. Two periods of V(D)J recombina-tion, at the pro- and pre-lymphocyte stages ofdevelopment, lead to expression of the B-cellreceptor (BCR) and TCR on the surface ofimmature B and T cells, respectively (Fig. 1,overleaf). The essential, lymphocyte-specific

Immunology

Developing B-cell theoriesDavid G. Schatz