Morphogens: experimental illusion or reality?

The main attention is paid to the critical analysis of experimental data on morphogenetically active substances, so called "morphogens". It is proposed to consider the morphogens as specific transmitters providing for definite phases of morphogenetic tissues interactions, rather than as vectors of "morphogenetic information". In the normal development, the most studied morphogenetic tissue interactions can be referred to as so called permissive inductions, since the cells of the vertebrate embryos (amphibians, avians) are early determined for development in the ectomeso--and endodermal directions. A slow progress in studying the morphogens can be due to the following causes. 1. Theoretical "inadequacy" of the former concepts on the essence and mechanisms of embryonic induction. The necessity to develop a new system of concepts in this area of developmental biology is stressed. 2. Incompleteness of knowledge about the properties of reacting tissues and the mechanisms of action of morphogens. The early gastrula ectoderm of amphibians, most frequently used for testing the morphogens, appears to be a heterogenous population of the cells with different properties and potencies. It is, therefore, impossible to standardize strictly the biotesting of morphogens. It is suggested that the use to this end of aggregates of cell "strains" from the gastrula ectoderm, rather than of the gastrula ectoderm itself, may be more adequate 3. Insufficiency of embryonic material for biochemical identification and isolation of natural morphogens. A study of so called heterogenous inductors might be of help; these latter can be considered as analogs of natural morphogens. But the similarity of natural and heterogenous inductors can be limited only by their final effect on target tissue. The data are provided on the chemical nature, properties and mechanisms of action for a number of natural and heterogenous inductors (vegetalizing, neuralizing, mesodermalizing, lens-inducing factors). A conclusion is drawn that specific antigens do exist normally but they should not be established as a special class of "informationally important" molecules. The information necessary for development is contained in target cells and the function of a morphogen consists in providing for a definite link in the chain of processes leading to the switching on or expression of one or another programme. Only syntheses of specific proteins can, apparently, be programmed, thus reflecting the "onset" of differentiation path for a cell.     

On the spread of morphogens

Pattern organisation of cells and tissue are specified during embryonic development by gradients of morphogens, substances that assign different fates of cells at different concentrations. Morphogen gradients form by transport from a localized site. Whether this occurs by diffusion or some other more elaborate mechanism is controversial. Here we provide an analysis of two models considered by Lander et al. [4] used in support of the diffusive transport hypothesis.     

Understanding morphogenetic growth control -- lessons from flies

Morphogens are secreted signalling molecules that control the patterning and growth of developing organs. How morphogens regulate patterning is fairly well understood; however, how they control growth is less clear. Four principal models have been proposed to explain how the morphogenetic protein Decapentaplegic (DPP) controls the growth of the wing imaginal disc in the fly. Recent studies in this model system have provided a wealth of experimental data on growth and DPP gradient properties, as well as on the interactions of DPP with other signalling pathways. These findings have allowed a more precise formulation and evaluation of morphogenetic growth models. The insights into growth control by the DPP gradient will also be useful for understanding other morphogenetic growth systems.     

BMP type I receptor complexes have distinct activities mediating cell fate and axon guidance decisions

The finding that morphogens, signalling molecules that specify cell identity, also act as axon guidance molecules has raised the possibility that the mechanisms that establish neural cell fate are also used to assemble neuronal circuits. It remains unresolved, however, how cells differentially transduce the cell fate specification and guidance activities of morphogens. To address this question, we have examined the mechanism by which the Bone morphogenetic proteins (BMPs) guide commissural axons in the developing spinal cord. In contrast to studies that have suggested that morphogens direct axon guidance decisions using non-canonical signal transduction factors, our results indicate that canonical components of the BMP signalling pathway, the type I BMP receptors (BMPRs), are both necessary and sufficient to specify the fate of commissural neurons and guide their axonal projections. However, whereas the induction of cell fate is a shared property of both type I BMPRs, axon guidance is chiefly mediated by only one of the type I BMPRs, BMPRIB. Taken together, these results indicate that the diverse activities of BMP morphogens can be accounted for by the differential use of distinct components of the canonical BMPR complex.     

Morphogenetic messages are in the extracellular matrix: biotechnology from bench to bedside

The origin and evolution of multicellular metazoa was accompanied by the appearance of extracellular matrix. The demineralized extracellular matrix of bone is enriched in morphogenetic proteins that induce bone. Bone morphogenetic proteins (BMPs) are intimately bound to collagens. BMP-4 has high affinity for type-IV collagen, and other binding proteins such as noggin and chordin. Soluble morphogens are kept in the solid state by extracellular matrix. In this sense Nature used the principles of affinity matrices long before humans patented the principle of affinity chromatography.     

The EGF receptor and notch signaling pathways control the initiation of the morphogenetic furrow during Drosophila eye development

The onset of pattern formation in the developing Drosophila retina begins with the initiation of the morphogenetic furrow, the leading edge of a wave of retinal development that transforms a uniform epithelium, the eye imaginal disc into a near crystalline array of ommatidial elements. The initiation of this wave of morphogenesis is under the control of the secreted morphogens Hedgehog (Hh), Decapentaplegic (Dpp) and Wingless (Wg). We show that the Epidermal Growth Factor Receptor and Notch signaling cascades are crucial components that are also required to initiate retinal development. We also show that the initiation of the morphogenetic furrow is the sum of two genetically separable processes: (1) the 'birth' of pattern formation at the posterior margin of the eye imaginal disc; and (2) the subsequent 'reincarnation' of retinal development across the epithelium.     

Endocytosis of Fgf8 Is a Double-Stage Process and Regulates Spreading and Signaling

Tightly controlled concentration gradients of morphogens provide positional information and thus regulate tissue differentiation and morphogenesis in multicellular organisms. However, how such morphogenetic fields are formed and maintained remains debated. Here we show that fibroblast growth factor 8 (Fgf8) morphogen gradients in zebrafish embryos are established and maintained by two essential mechanisms. Firstly, Fgf8 is taken up into the cell by clathrin-mediated endocytosis. The speed of the uptake rate defines the range of the morphogenetic gradient of Fgf8. Secondly, our data demonstrate that after endocytosis the routing of Fgf8 from the early endosome to the late endosome shuts down signaling. Therefore, intracellular endocytic transport regulates the intensity and duration of Fgf8 signaling. We show that internalization of Fgf8 into the early endosome and subsequent transport towards the late endosome are two independent processes. Therefore, we hypothesize that Fgf8 receiving cells control both, the propagation width and the signal strength of the morphogen.     

Secreted Wnt "inhibitors" are not just inhibitors: regulation of extracellular Wnt by secreted Frizzled-related proteins

Gradient formation and signaling ranges of secreted proteins are crucial problems to understand how morphogens work for positional information and patterning in animal development. Yet, extracellular behaviors of secreted signaling molecules remain unexplored compared to their downstream pathways inside the cell. Recent advances in bioimaging make it possible to directly visualize morphogen molecules, and this simple strategy has, at least partly, succeeded in uncovering molecular behaviors of morphogens, such as Wnt (wingless-type MMTV integration site family member) and BMP (bone morphogenetic protein) as well as secreted Wnt binding proteins, sFRPs (secreted Frizzled-related proteins), in embryonic tissues. Here, we review the regulation of Wnt signaling by sFRPs, focusing on extracellular regulation of Wnt ligands in comparison with other morphogens. We also discuss evolutionary aspects with comprehensive syntenic and phylogenetic information about vertebrate sfrp genes. We newly annotated several sfrp genes including sfrp2-like 1 (sfrp2l1) in frogs and fishes and crescent in mammals.     

Unveiling the bmp13 enigma: redundant morphogen or crucial regulator?

Bone morphogenetic proteins are a diverse group of morphogens with influences not only on bone tissue, as the nomenclature suggests, but on multiple tissues in the body and often at crucial and influential periods in development.     

The Hox-gene research programme and the shortcomings of molecular preformationism.

The study of the so-called HOM/Hox genes has provided many important insights on the control, at the molecular level, of developmental processes in a variety of model systems such as the fly and the mouse. Yet, in the specialised literature on the subject abound the claims that such genes, and the products coded by them, are the true morphogens responsible for determining the actual form of a particular organism. According to this view, morphogenesis results from the expression of specific 'master control' genes and thus, organic form is somehow pre-established (i.e., preformed) as an assembly programme encoded within the genome. Or in other words, it is claimed that the complex spatio-temporal order that leads to the achievement and maintenance of organic form is implied in a two-dimensional organisation of the genome. Moreover, some authors have claimed that the success of the Hox-gene research programme strongly suggests that morphological evolution is a direct result of evolution at the genetic level. Hereunder I discuss recent evidence that falsifies the basic preformationist tenets of molecular developmental biology. Thus suggesting that the problem of the origin of organic form is left untouched by the Hox-gene research programme and therefore, there is a need to reconsider alternative approaches, such as the structuralist morphogenetic outlook, that are better suited to eventually explain the origin of organic form.     

ERK and MMPs sequentially regulate distinct stages of epithelial tubule development

Epithelial cells undergo tubulogenesis in response to morphogens such as hepatocyte growth factor (HGF). To organize into tubules, cells must execute a complex series of morphogenetic events; however, the mechanisms that underlie the timing and sequence of these events are poorly understood. Here, we show that downstream effectors of HGF coordinately regulate successive stages of tubulogenesis. Activation of extracellular-regulated kinase (ERK) is necessary and sufficient for the initial stage, during which cells depolarize and migrate. ERK becomes dispensable for the latter stage, during which cells repolarize and differentiate. Conversely, the activity of matrix metalloproteases (MMPs) is essential for the late stage but not the initial stage. Thus, ERK and MMPs define two regulatory subprograms that act in sequence. By inducing these reciprocal signals, HGF directs the morphogenetic progression of tubule development.     

A local activator-inhibitor model of vertebrate skin patterns

A model for vertebrate skin patterns is presented in which the differentiated (colored) pigment cells produce two diffusible morphogens, an activator and an inhibitor. The concentrations of these two substances at any point on the skin determine whether a pigment cell at that point will be colored or not. Computer simulations with this model show many realistic features of spot and stripe patterns found in vertebrates.     

Regulation of articular chondrocyte phenotype by bone morphogenetic protein 7, interleukin 1, and cellular context is dependent on the cytoskeleton.

Bone morphogenetic proteins (BMPs) induce cartilage differentiation and morphogenesis. There are profound changes in the cytoskeletal architecture during the morphogenesis of cartilage. To investigate the possibility that morphogenetic signals such as BMPs may regulate chondrocyte phenotype by modulation of cytoskeletal protein expression, we determined whether the expression and distribution of cytoskeletal proteins in chondrocytes are regulated by bone morphogenetic protein 7 (BMP 7), interleukin 1 (IL-1), and cellular context. Addition of BMP 7, a morphogen that induces chondrogenesis, to primary cultures of bovine and murine chondrocytes induced increased expression of four cytoskeletal proteins: tensin, talin, paxillin, and focal adhesion kinase (FAK). The expression of cytoskeletal proteins is dependent on cellular context; compared to monolayer, chondrocytes in suspension exhibited increased expression of cytoskeletal components. Conversely, addition of IL-1, a catabolic cytokine, induced loss of chondrocyte phenotype and decreased the expression of these cytoskeletal components. Treatment of chondrocytes with cytochalasin D (an agent that disrupts the actin cytoskeleton) inhibited BMP 7-induced upregulation of tensin, talin, paxillin, and FAK, and blocked the effect of BMP 7 on chondrocyte phenotype. Taken together these data demonstrate that cytoskeletal components play a critical role in the response to morphogens and cytokines in the regulation of chondrocyte phenotype. (c)2001 Elsevier Science.     

The Yin and Yang of bone morphogenetic proteins in cancer

Bone morphogenetic proteins (BMPs) were first studied as growth factors or morphogens of the transforming growth factor-beta superfamily. These growth molecules, originally associated with bone and cartilage development, are now known to play an important role in morphogenesis and homeostasis in many other tissues. More recently, significant contributions from BMPs, their receptors, and interacting molecules have been linked to carcinogenesis and tumor progression. On the other hand, BMPs can sometimes function as a tumor suppressor. Our report highlights these new roles in the pathogenesis of cancer that may suggest novel targets for therapeutic intervention.     

Understanding morphogen gradients: a problem of dispersion and containment

Protein morphogens are instructive signals that regulate growth and patterning of tissues and organs. They form long-range, dynamic gradients by moving from regions of high concentration (producing cells) to regions of low concentration (the adjacent, nonproducing developmental field). Since morphogen activity must be limited to the adjacent target field, we want to understand both how signaling proteins move and how their dispersion is restricted. We consider the variety of settings for long-range morphogen systems in Drosophila. In the early embryo, morphogens appear to disperse by free diffusion, and impermeable membranes physically constrain them. However, at later stages, containment is achieved without physical barriers. We argue that in the absence of constraining barriers, gradient-generating dispersion of morphogens cannot be achieved by passive diffusion and that other mechanisms for distribution must be considered.     

Mechanical stresses as factors in the autoregulation of morphogenesis

Since morphogenetic processes are nonlinear, feedback must be essential for their regulation. Two concepts of feedback in morphogenesis are developed: 1) inhibition by diffusing morphogenetic substances and 2) action of mechanical strain resulting from morphogenetic movements. The data on the role of mechanical strain in formation of integral structure of embryonic tissues, primary demarcation of embryonic epithelia and mesenchymal rudiments and bending of epithelial layers are presented. Evolutionary aspects of morphogenetic role of mechanical strain and its possible use in applied biotechnology are discussed.     

Multiple morphogens and rapid elongation promote segmental patterning during development

The vertebrate hindbrain is segmented into rhombomeres (r) initially defined by distinct domains of gene expression. Previous studies have shown that noise-induced gene regulation and cell sorting are critical for the sharpening of rhombomere boundaries, which start out rough in the forming neural plate (NP) and sharpen over time. However, the mechanisms controlling simultaneous formation of multiple rhombomeres and accuracy in their sizes are unclear. We have developed a stochastic multiscale cell-based model that explicitly incorporates dynamic morphogenetic changes (i.e. convergent-extension of the NP), multiple morphogens, and gene regulatory networks to investigate the formation of rhombomeres and their corresponding boundaries in the zebrafish hindbrain. During pattern initiation, the short-range signal, fibroblast growth factor (FGF), works together with the longer-range morphogen, retinoic acid (RA), to specify all of these boundaries and maintain accurately sized segments with sharp boundaries. At later stages of patterning, we show a nonlinear change in the shape of rhombomeres with rapid left-right narrowing of the NP followed by slower dynamics. Rapid initial convergence improves boundary sharpness and segment size by regulating cell sorting and cell fate both independently and coordinately. Overall, multiple morphogens and tissue dynamics synergize to regulate the sizes and boundaries of multiple segments during development.     

The problem of transition from the chemical to the biological evolution: some possible solutions

On the basis of evidence that several low-molecular-weight substances as well as enzymes are compartmentalised within the so-called soluble phase of the cell, and other considerations, it is argued that DNA may not contain information for certain types of organisation found in living cells. It may be necessary for a cell to possess the "non-DNA-controlled" organisation for performance of its minimum functions; such organisation would then also serve as a "template" for its appearance in the daughter cell. The problem of transition from chemical to biological evolution (that is, the formation of the "first cell") may be essentially the problem of emergence of such intracellular organisation for which information may not reside in DNA. Two possible mechanisms through which this may have happened are stated.