DEVELOPMENT385Morphogens act as graded positional cues that control cell fatespecification in many developing tissues. This concept, in which asignalling gradient regulates differential gene expression in aconcentration-dependent manner, provides a basis forunderstanding many patterning processes. It also raises severalmechanistic issues, such as how responding cells perceive andinterpret the concentration-dependent information provided bya morphogen to generate precise patterns of gene expressionand cell differentiation in developing tissues. Here, we reviewrecent work on the molecular features of morphogen signallingthat facilitate the interpretation of graded signals and attemptto identify some emerging common principles.IntroductionThe transformation of the spatial distribution of naïve cells in adeveloping tissue into an organised arrangement of cell differentiationis fundamental to the development of multicellular organisms. Morethan a century ago, evidence began to accumulate that cells receive‘positional information’ that instructs them to develop in specificways, depending on their location within a tissue (Wolpert, 1996).Over the intervening decades, the potential for signalling gradients toprovide this positional information has become a much-investigatedand -debated subject, and the term ‘morphogen’ has been coined todescribe such signals. Today the morphogen concept continues toform the basis of many models of pattern formation (Lewis et al.,1977; Green and Smith, 1991; Gurdon and Bourillot, 2001; Tabata andTakei, 2004). Typically, in current models it is proposed that a signalproduced from a defined localised source forms a concentrationgradient as it spreads through surrounding tissue (Fig. 1A). The gradedsignal then acts directly on cells, in a concentration-dependent manner,to specify gene expression changes and cell fate selection. Thus, theconcentration of ligand provides cells with a measure of their positionrelative to the source of the signal and organises the pattern of celldifferentiation. Experimental evidence from tissues in both vertebratesand invertebrates indicates that several molecules appear to functionas graded signals. The roles of these signals range from theestablishment of the initial polarities of embryos to specification ofcell identity in specific tissues, notably limb appendages and thenervous system in both vertebrates and Drosophila. The examples wefocus on in this review are introduced in Fig. 1. Evidence in supportof these signals acting as graded morphogens has been summarised inrecent reviews (Gurdon and Bourillot, 2001; Tabata and Takei, 2004).Although the morphogen concept has provided an enduring andvalid framework for understanding pattern formation, it raises manymechanistic issues. Much attention has focused on how thedistribution of a morphogen through a tissue establishes andmaintains a gradient of activity (Vincent and Dubois, 2002; Tabataand Takei, 2004); however, how the signal is perceived andinterpreted in a graded manner by the receiving cells has received lessconsideration. Nonetheless, this represents an equally importantelement of the morphogen hypothesis. Crucial to understanding themechanism of morphogen activity is determining how a graded signalis transformed into alterations in gene expression programmes, suchthat the positional information supplied by the morphogen producesthe appropriate spatial pattern of cellular differentiation. Tounderstand how this is accomplished, several questions have to beaddressed. How does the signal transduction pathway transmit gradedinformation intracellularly to control concentration-dependentdifferential gene expression? How is a continuous gradienttransformed into discrete changes in gene expression that ultimatelydetermine the choice of cell fate from the available alternatives? Andhow does graded signalling accommodate fluctuations in biologicalconditions to achieve the necessary robustness required for accuratedevelopmental patterning? By focusing on specific examples, wereview recent work that addresses these questions and, wherepossible, we highlight some of the general principles that appear tobe shared between different morphogen gradients.Morphogen signal transduction pathways arelinear and transmit graded informationHow many thresholds does a morphogen control?At a minimum, to meet the definition of a morphogen, a gradedsignal must be able to direct the generation of at least two distinctcell types at different concentrations. Theoretical analysis has raisedthe possibility that graded signals can achieve up to 30 thresholds(Lewis et al., 1977); however, empirical evidence has typicallyidentified between three and seven distinct thresholds. For example,the Dorsal (Dl) gradient appears to specify at least four, and as manyas seven, distinct thresholds of gene expression along thedorsoventral (DV) axis of Drosophila embryos (Stathopoulos andLevine, 2002a). A concentration gradient of activin is able to inducefive cell states in Xenopus blastula cells (Green et al., 1992), and asimilar number of neuronal subtypes appears to be produced bygraded Sonic Hedgehog (Shh) signalling in the neural tube (Ericsonet al., 1997; Pierani et al., 1999). In each of these cases, additionalsignals are believed to promote or cooperate in the forming of someof the threshold responses, so whether a single morphogen actingalone produces each of the observed threshold responses remainsunknown. In other well-studied cases, fewer defined thresholds havebeen clearly identified, for example Wingless (Wg) signalling in theDrosophila wing imaginal disc promotes three thresholds of geneexpression (Tabata and Takei, 2004), whereas gradedDecapentaplegic (Dpp) signalling is responsible for at least threethreshold responses in Drosophila embryos and the wing disc (Asheet al., 2000; Affolter et al., 2001).Small morphogen concentration changes are sensedIn the case of the vertebrate morphogens activin, bonemorphogenetic protein (Bmp) 4 and Shh, the dose responses of cellshave been assayed (Green et al., 1992; Wilson et al., 1997; Ericsonet al., 1997). For activin and Shh, the full range of responses iselicited over a 25- to 50-fold concentration range with relativelyDevelopment 133, 385-394 doi:10.1242/dev.02238The interpretation of morphogen gradientsHilary L. Ashe1and James Briscoe21Faculty of Life Sciences, The University of Manchester, Oxford Road, ManchesterM13 9PT, UK. 2Developmental