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Epithelia and mesenchyme interact during various physiologic and pathologic processes. Scatter factor is a mesenchyme-derived cytokine that stimulates motility, proliferation, and morphogenesis of epithelia. Recent studies suggest that scatter factor and its receptor (c-met) mediate mesenchyme/epithelia signalling and even interconversion. In this mini-review, we will discuss how scatter factor and c-met may mediate interactions between mesenchyme and epithelia during embryogenesis, organ repair, and neoplasia.

Conventional myosin has two different light chains bound to the neck region of the molecule. It has been suggested that the light chains contribute to myosin function by providing structural support to the neck region, therefore amplifying the conformational changes in the head following ATP hydrolysis (Rayment et al., 1993). The regulatory light chain is also believed to be important in regulating the actin-activated ATPase and myosin motor function as assayed by an in vitro motility assay (Griffith et al., 1987). Despite extensive in vitro biochemical study, little is known regarding RMLC function and its regulatory role in vivo. To better understand the importance and contribution of RMLC in vivo, we engineered Dictyostelium cell lines with a disrupted RMLC gene. Homologous recombination between the introduced gene disruption vector and the chromosomal RMLC locus (mlcR) resulted in disruption of the RMLC-coding region, leading to cells devoid of both the RMLC transcript and the 18-kD RMLC polypeptide. RMLC-deficient cells failed to divide in suspension, becoming large and multinucleate, and could not complete development following starvation. These results, similar to those from myosin heavy chain mutants (DeLozanne et al., 1987; Manstein et al., 1989), suggest the RMLC subunit is required for normal cytokinesis and cell motility. In contrast to the myosin heavy chain mutants, however, the mlcR cells are able to cap cell surface receptors following concanavilin A treatment. By immunofluorescence microscopy, RMLC null cells exhibited myosin localization patterns different from that of wild-type cells. The myosin localization in RMLC null cells also varied depending upon whether the cells were cultured in suspension or on a solid substrate. In vitro, purified RMLC- myosin assembled to form thick filaments comparable to wild-type myosin, but the filaments then exhibit abnormal disassembly properties. These results indicate that in vivo RMLC is necessary for myosin function.

Many proteins have been shown to cap the fast growing (barbed) ends of actin filaments, but none have been shown to block elongation and depolymerization at the slow growing (pointed) filament ends. Tropomodulin is a tropomyosin-binding protein originally isolated from red blood cells that has been localized by immunofluorescence staining to a site at or near the pointed ends of skeletal muscle thin filaments (Fowler, V. M., M. A., Sussman, P. G. Miller, B. E. Flucher, and M. P. Daniels. 1993. J. Cell Biol. 120: 411-420). Our experiments demonstrate that tropomodulin in conjunction with tropomyosin is a pointed end capping protein: it completely blocks both elongation and depolymerization at the pointed ends of tropomyosin-containing actin filaments in concentrations stoichiometric to the concentration of filament ends (Kd < or = 1 nM). In the absence of tropomyosin, tropomodulin acts as a "leaky" cap, partially inhibiting elongation and depolymerization at the pointed filament ends (Kd for inhibition of elongation = 0.1-0.4 microM). Thus, tropomodulin can bind directly to actin at the pointed filament end. Tropomodulin also doubles the critical concentration at the pointed ends of pure actin filaments without affecting either the rate of extent of polymerization at the barbed filament ends, indicating that tropomodulin does not sequester actin monomers. Our experiments provide direct biochemical evidence that tropomodulin binds to both the terminal tropomyosin and actin molecules at the pointed filament end, and is the long sought-after pointed end capping protein. We propose that tropomodulin plays a role in maintaining the narrow length distributions of the stable, tropomyosin-containing actin filaments in striated muscle and in red blood cells.

role of the cell body in generating the axonal microtubule array, whereas the other emphasizes the role of local mechanisms within the axon itself. Recent work suggests that events significant to the generation of the axonal microtubule array occur within both of these compartments, and that axon growth is dependent upon the coordinated efforts of several types of microtubule behavior. Our goal in this article is to evaluate the existing models in light of more recent data, and provide new perspectives on the manner by which the axonal microtubule array is generated.

Five structural features in mRNAs have been found to contribute to the fidelity and efficiency of initiation by eukaryotic ribosomes. Scrutiny of vertebrate cDNA sequences in light of these criteria reveals a set of transcripts--encoding oncoproteins, growth factors, transcription factors, and other regulatory proteins--that seem designed to be translated poorly. Thus, throttling at the level of translation may be a critical component of gene regulation in vertebrates. An alternative interpretation is that some (perhaps many) cDNAs with encumbered 5' noncoding sequences represent mRNA precursors, which would imply extensive regulation at a posttranscriptional step that precedes translation.

Article de synthese traitant des mecanismes biochimiques et genetiques regulant le phenomene de differenciation cellulaire pour 2 especes d'insectes presentant ou non des mutations dans leur genome

Heat shock factor Hsf in nonvertebrate animals and homologous heat shock factor Hsf1 in vertebrate animals are key transcriptional regulators of the stress protein response. Hsf/Hsf1 is constitutively present in cells but is, typically, only active during periods during which cells are experiencing a physical or chemical proteotoxic stress. It has become increasingly clear that regulation of Hsf/Hsf1 activity occurs at multiple levels: the oligomeric status of Hsf/Hsf1, its DNA-binding ability, posttranslational modification, transcriptional competence, nuclear/ subnuclear localization, as well as its interactions with regulatory cofactors or other transcription factors all appear to be carefully controlled. This review emphasizes work reported over the past several years suggesting that regulation at several of these levels is mediated by repressive interactions of Hsp90-containing multichaperone complexes and/or individual chaperones and Hsf/Hsf1.

The molecular chaperone machinery contains multiple protein components that have 1 or more structural domains composed of tetratricopeptide repeat (TPR) motifs. Many other proteins of separate or unknown function also have TPR domains, so this motif is not exclusive to molecular chaperones. A general function of TPR domains is to bind other polypeptides, but this otherwise prosaic function has been exploited in an assortment of ways that link chaperones and other protein systems into cooperative networks. Among the best-characterized TPR proteins are several cochaperones that participate in assembly and regulation of steroid receptor complexes. Steroid receptors, members of the nuclear receptor subfamily, are hormone-dependent transcription factors that regulate many vertebrate pathways of homeostasis, growth, differentiation, reproduction, and pathology and, as such, have been of great interest to biologists and clinicians. Moreover, the steroid receptors are among the first recognized native clients for chaperones and have been widely studied models for complex chaperone interactions. To provide a coherent, representative minireview of TPR protein function, the scope of this article has been narrowed down primarily to functions of steroid receptor-associated TPR cochaperones.

The dramatic transformations in nuclear content and cellular organization that occur during gametogenesis require unique regulation and execution of the mitotic and meiotic cell cycle, apoptotic cell death, DNA recombination and repair, and cellular differentiation. These processes are accompanied by the constitutive and developmentally regulated expression of a number of hsp70 genes encoding 70 kDa heat shock proteins (Hsp70), including several hsp70s whose expression is unique to male germ cells. Examining the expression and function of Hsp70s in germ cells has provided significant insights into mechanisms of hsp70 gene regulation and Hsp70 protein function, as well as the developmental processes of gametogenesis.

The prion encephalopathies are a group of transmissible and inherited neurodegenerative diseases of man and animals (for reviews see Prusiner et al 1989; Prusiner 1991, 1992). Sheep (or goat) scrapie is the most studied of the prion diseases and interest has increased more recently with the outbreak of 'mad cow disease' (bovine spongiform encephalopathy, BSE) in cattle in the UK as this may indicate a transfer of scrapie from sheep into another species through the use of rendered contaminated sheep offal in cattle feed. While scrapie has been present in sheep for over 200 years without causing any public health hazard, the fact that it has been transferred to another species by ingestion of infected material may indicate a risk to humans through consumption of food from BSE-infected animals.

The human Hsp70 family encompasses at least 11 genes which encode a group of highly related proteins. These proteins include both cognate and highly inducible members, at least some of which act as molecular chaperones. The location of cognate Hsp70s within all the major subcellular compartments is an indication of the importance of these proteins. The expression of several inducible Hsp70 genes is also an indication of the importance of these proteins in the stres response. The existence of multiple genes and protein isoforms has created confusion in the identification and naming of particular family members. We have compiled, from the literature, a list of genes and genetic loci and produced a two-dimensional protein map of the known human Hsp70 family members. This will enable researchers in the field to quickly and reliably identify human Hsp70s. We have also devised a more rational nomenclature for these genes and gene products which, subject to general acceptance, could be extended to Hsp70 families from other species.

We surveyed published reports on about 50 glycoproteins whose amino acid sequence, glycosylation sites, and type of glycosylation at a particular site have been established. We note that high-mannose substances were rarely found at the N-terminal side of a previously glycosylated complex site. There was a very definite distribution of complex sites about the N-terminal region. Furthermore, secreted glycoproteins usually contained only complex oligosaccharides whereas membrane proteins contained both types. We suggest that the position of the glycosylation site with respect to the N-terminus affects the extent of oligosaccharide processing and subsequent presentation of complex or high-mannose structures in the mature glycoprotein. This review relates glycosylation type to its position in the known sequence of given proteins and discusses these observations in light of known glycosylation processing reactions.

Revue sur les connaissances actuelles concernant la biologie des facteurs de croissance basiques: biosynthese, localisation, activites biologiques, proteines associees, potentiel transformant, recepteurs, liberation, interactions avec les matrices

The Rous sarcoma virus (RSV) transforming gene product has been identified and characterized as a phosphoprotein with a molecular weight of 60,000, denoted pp60src. Partially purified pp60src displays a closely associated phosphotransferase activity with the unusual specificity of phosphorylating tyrosine residues in a variety of proteins. That the enzymatic activity observed is actually encoded by the RSV-transforming gene is indicated by the comparison of the pp60src-protein kinase isolated from cells tranformed by a wild-type RSV or by a RSV temperature-sensitive transformation mutant; these experiments revealed that the latter enzyme had a half-life of 3 min at 41 degrees C, whereas that of the wild-type enzyme was 20 min. Evidence is now beginning to accumulate showing that viral pp60src expresses its protein kinase activity in transformed cells as well as in vitro because at least one cellular protein has been identified as a substrate for this activity of pp60src. Although the protein kinase activity associated with pp60src is itself cyclic AMP (cAMP) independent, the molecule contains at least one serine residue that is directly phosphorylated by the cellular cAMP-dependent protein kinase, thus suggesting that the viral transforming gene product may be regulated indirectly by the level of cAMP. The significance of this latter observation must be regarded from the point of view that the RSV src gene is apparently derived from a normal cellular gene that seemingly expresses in normal uninfected cells a phosphoprotein structurally and functionally closely related to pp60src. This celluar protein, found in all vertebrate species tested, also is a substrate for a cAMP-dependent protein kinase of normal cells, and, therefore, may be evolved to function in a regulatory circuit involving cAMP.

ARASITIC, bacterial, and viral infections and neoplastic disease profoundly affect host metabolism, disrupting normal homeostatic mechanisms both locally and systemically. A large body of research has documented that many of the observed biological responses to invasive stimuli are mediated by host-secreted cytokines, in particular, the secretory products of activated macrophages. One such cytokine, cachectin/tumor necrosis factor (TNF),~ has emerged as a particularly important mediator of inflammatory responses. Among its pleiotropic effects, cachectin/TNF has been shown to play a major endocrine role in the pathogenesis of gram-negative endotoxic shock (14, 118) and to induce catabolic responses which could contribute to the profound wasting (cachexia) associated with many chronic diseases (24, 83, 104, 121). It has been implicated in a variety of disease states including meningococcal septicemia (126), cerebral malaria (43), graft vs. host disease (93), and cancer cachexia (2, 8). Systemic exposure to this potent mediator might elicit the pathologic and catabolic derangements associated with such disease states. Cachectin/TNF has also been shown to exert local, tissuespecific effects. Its wide range of bioactivities includes stimulation of collagenase activity and prostaglandin E2 production by synovial cells (33), stimulation of osteoclast activity and bone resorption (9), promotion of angiogenesis (41, 60), and stimulation of procoagulant and platelet-activating factor activity in endothelial tissue (19, 75). It has also been shown to stimulate proliferation of normal fibroblasts (40, 114, 125), and to induce the release of certain growth factors including interleukin 1 (IL- 1), granulocyte-macrophage colony-stimulating factor (GM-CSF), I]2-interferon, and platelet-derived growth factor (46, 55, 58, 73). The ability of cachectin/TNF to modulate such activities raises the intriguing possibility that it may play a paracrine role in the regulation of normal tissue homeostasis. Cachectin/TNF also exerts profound effects on its own principal cell of origin, the macrophage. It serves as an autocrine immunomodulator, activating macrophages and enhancing their cytotoxic potential in vitro (92). It has been shown to be chemotactic for monocytes, suggesting that its production at a site of injury might function both to recruit additional macrophages and to activate those macrophages already present. 1. Abbreviations used in this paper: IL-1, interleukin 1; LPL, lipoprotein lipase; LPS, lipopolysaccharide; TNF, tumor necrosis factor. The capacity of this potent cytokine to mediate the inflammatory response, to modulate the metabolic activities of diverse tissues, and to augment the function of other cytokines necessitates that its synthesis and release be closely controlled in vivo. Local production of cachectin/TNF at a site of injury might act to limit tissue damage and to promote wound healing and tissue remodeling. Moderate systemic levels of the hormone might confer a survival advantage with respect to bacterial or viral infection by providing a useful mobilization of energy reserves for the acute metabolic demands of inflammatory responses. In sharp contrast to these beneficial effects, prolonged exposure to even low levels of cachectin/TNF might contribute to the cachexia associated with many chronic disease states. Rapid uncontrolled production of cachectin/TNF, like that observed in response to endotoxemia or overwhelming gram-negative sepsis, could act systemically to induce the metabolic derangements of septic shock leading to cardiovascular collapse, acute organ failure, and death.

A chlorophyll-protein complex of chloroplast membranes, which simultaneously serves as light-harvesting antenna and membrane adhesion factor, undergoes reversible, lateral diffusion between appressed and nonappressed membrane regions under the control of a protein kinase. The phosphorylation-dependent migration process regulates the amount of light energy that is delivered to the reaction centers of photosystems I and II (PS I and PS II), and thereby regulates their rate of turnover. This regulatory mechanism provides a rationale for the finding that the two photosystems are physically separated in chloroplast membranes (PS II in appressed, grana membranes, and PS I in nonappressed, stroma membranes). The feedback system involves the following steps: a membrane-bound kinase senses the rate of PS II vs. PS I turnover via the oxidation-reduction state of the plastoquinone pool, which shuttles electrons from PS II via cytochrome f to PS I. If activated, the kinase adds negative charge (phosphate) to a grana-localized pigment-protein complex. The change in its surface charge at a site critical for promoting membrane adhesion results in increased electrostatic repulsion between the membranes, unstacking, the lateral movement of the complex to adjacent stroma membranes, which differ in their functional composition. The general significance of this type of membrane regulatory mechanism is discussed.

Eucaryotic messenger RNA precursors are processed in nuclear ribonucleoprotein particles (hnRNP). Here recent work on the structure of hnRNP is reviewed, with emphasis on function. Detailed analysis of a specific case, the altered assembly of hnRNP in heat-shocked Drosophila and mammalian cells, leads to a general hypothesis linking hnRNP structure and messenger RNA processing.

ERV~ cells display a number of unusual properties which are generally viewed as adaptations to their extreme length and regional specialization. The dense and highly organized neurofilament (NF) ~ and microtubule (MT) arrays which serve to support nerve axons provide one example; another is the phenomenon of axonal transport, by which macromolecules synthesized in the cell body move outward along the axon in several relatively discrete components with widely different velocities (3, 22, 32, 95; Fig. 1). During the 1970s, a crucial connection was made between these two properties of nerve cells by the demonstration that the slowest moving component of axonal transport (called slow component a [SCa]) conveys primarily tubulin and the three proteins comprising NFs (18, 23, 27, 33, 41, 52, 78, 96). This was achieved by a synthesis of the biochemical and ultrastructural data available at that time; since then, intensive study of neuronal cytoskeletal proteins has generated a succession of models of their transport, assembly, and turnover (39, 43, 58, 83, 93, 94). Recently, a number of studies have indicated that the components and interactions of the neuronal cytoskeleton may be more complex than previously thought. This review will attempt to evaluate whether current models are sufficient to accommodate the results of radiolabeling studies of axonal transport, ultrastructural studies, and recent work on the dynamics of the MT and NF systems. The principal questions to be addressed are (a) in what state are cytoskeletal proteins transported in axons; (b) how are their assembly and interactions regulated; and (c) in what region or regions of the axon does assembly of the cytoskeleton occur?

T H E mammalian intestinal epithelium represents a unique model system for studying cellular differentiation since it undergoes continuous and rapid renewal. Perpetual differentiation results in the generation of its four principal cell types, each with a unique phenotype. The processes of proliferation and cellular differentiation are topologically very well organized. Gradients in gene expression are established and maintained in several spatial dimensions within this epithelium. Recent experiments involving mouse aggregation chimeras and transgenic mice have provided insights about the origins of intestinal stem cells, the migration pathways followed during cellular differentiation as well as the molecular mechanisms which produce complex geographic differences in gut gene expression. The purpose of this mini-review is to emphasize the usefulness of these powerful methods for examining questions related to enteric epithelial biology.

Mini-synthese des problemes de mesure de concentration de calcium intracellulaire dans des cellules en division chez des invertebres (oursin Lytechinus, Hemicentrotus), chez des mammiferes (rat-kangourou, cellules 3T3) et chez les vegetaux (Haemanthus et Tradescantia) par differentes techniques, les unes utilisant l'ester de fura-2, des indicateurs fluorescents comme le quine, l'aequorine, le BAPTA ou des indicateurs colores comme l'arsenazo III, et des mesures par iontophorese

Platelet adhesion to the subendothelial matrix and the subsequent aggregation of the activated platelets is a special case of intercellular adhesion that is highly regulated and for which multiple, possibly redundant adhesive systems exist. The large "adhesive glycoproteins" such as fibrinogen, von Willebrand factor, and fibronectin and their receptors, which participate in platelet adhesion and aggregation, are not unique to platelets, and probably serve analogous functions in a variety of cell types and locations. Thrombospondin (TS)/the most abundant protein of platelet alpha-granules, has been recognized as a new member of this class of adhesive glycoproteins. Whereas other adhesive glycoproteins are contained within alpha-granules and secreted upon platelet activation, they also exist in plasma at high concentrations. Hence TS is the only member of the group that is unique to alpha-granules and whose expression in quantity at sites of platelet aggregation is absolutely dependent on

In interspecific hybrids, it is often observed that the ribosomal genes of one species are transcriptionally dominant over the ribosomal genes of the other species. This phenomenon has been called "nucleolar dominance" and has been reported in such diverse organisms as frogs (Xenopus), Drosophila, many genera of plants, and mammalian somatic cell hybrids. Recent advances in our knowledge of the structure of ribosomal genes and their transcription machinery have led to proposals that at least two different molecular mechanisms can operate to cause nucleolar dominance and that the relative contribution of each mechanism is different for different types of crosses. One proposed mechanism involves competition between ribosomal genes which possess unequal numbers of enhancer elements. This mechanism can be abbreviated as the "enhancer imbalance mechanism." The second proposed mechanism involves the fact that the ribosomal gene (RNA polymerase I) transcription machinery evolves more rapidly between species than does the machinery for the other two classes of polymerase. This leads to dominance effects based on the apparent inability of a key transcription factor from one species to recognize the ribosomal gene promoter of the other species. This mechanism will be referred to as the "speciesspecific factor mechanism." The purpose of this review is to briefly summarize the evidence for these two molecular mechanisms and then to examine each of the known types of nucleolar dominance to assess how well the proposed mechanisms can account for each case.

In most eucaryotes the tubulin genes comprise small multigene families with approximately equal numbers of genes for alpha- and beta-tubulin, the structural proteins of microtubules. The recent isolation of tubulin mutations in several species is proving to be a powerful tool for examining the structure and function of specific sets of microtubules. In Drosophila melanogaster, genetic analysis of a testis-specific beta-tubulin gene has shown that a single tubulin gene product may fulfill a number of different microtubule functions. In addition to tubulin mutations, mutations in other genes whose products are involved in the regulation or structure of specific microtubule arrays have also been isolated. The combination of analysis of both classes of mutations is beginning to allow a molecular description of the construction and function of three-dimensional cellular structures. In addition, such studies may also shed light on the evolutionary pressures that gave rise to and serve to maintain small families of genes encoding very similar proteins.

Band 4.9 (a 48,000-mol-wt polypeptide) has been partially purified from human erythrocyte membranes. In solution, band 4.9 polypeptides exist as trimers with an apparent molecular weight of 145,000 and a Stokes radius of 50 A. Electron microscopy shows that the protein is a three-lobed structure with a radius slightly greater than 50 A. When gel-filtered rabbit muscle actin is polymerized in the presence of band 4.9, actin bundles are generated that are similar in appearance to those induced by "vinculin" or fimbrin. The bundles appear brittle and when they are centrifuged small pieces of filaments break off and remain in the supernatant. At low band 4.9 to actin molar ratios (1:30), band 4.9 lowers the apparent steady-state low-shear falling ball viscosity by sequestering filaments into thin bundles; at higher ratios, the bundles become thicker and obstruct the ball's movement leading to an apparent increase in steady-state viscosity. Band 4.9 increases the length of the lag phase and decreases the rate of elongation during actin polymerization as measured by high-shear Ostwald viscometry or by the increase in the fluorescence of pyrene-labeled actin. Band 4.9 does not alter the critical actin monomer concentration. We hypothesize that band 4.9, together with actin, erythrocyte tropomyosin, and spectrin, forms structures in erythroid precursor cells analogous to those formed by fimbrin, actin, tropomyosin, and TW 260/240 in epithelial brush borders. During erythroid development and enucleation, the actin filaments may depolymerize up to the membrane, leaving a membrane skeleton with short stubs of actin bundled by band 4.9 and cross-linked by spectrin.

Primary aldosteronism (PA) is a common form of endocrine hypertension that is characterized by the excessive production of aldosterone relative to suppressed plasma renin levels. PA is usually caused by either a unilateral aldosterone-producing adenoma or bilateral adrenal hyperplasia. Somatic mutations have been identified in several genes that encode ion pumps and channels that may explain the aldosterone excess in over half of aldosterone-producing adenomas, whereas the pathophysiology of bilateral adrenal hyperplasia is largely unknown. A number of mouse models of hyperaldosteronism have been described that recreate some features of the human disorder, although none replicate the genetic basis of human PA. Animal models that reproduce the genotype–phenotype associations of human PA are required to establish the functional mechanisms that underlie the endocrine autonomy and deregulated cell growth of the affected adrenal and for preclinical studies of novel therapeutics. Herein, we discuss the differences in adrenal physiology across species and describe the genetically modified mouse models of PA that have been developed to date.

Presentation et discussion des deux hypotheses sur le ou les genes des tubulines α et β, base des polymeres des microtubules

ADP-rlbosylation of regulatory proteins is an important mechanism by which bacterial toxins affect the eukaryotic organism.9,27,32 Diphtheria toxin and Pseudomonas aeruginosa exotoxin A modify elongation factor 2 thereby inhibiting protein synthesis.9,26,32 Cholera toxin, Escherichia coli heat-labile enterotoxin and pertussis toxin ADP-ribosylate regulatory G-proteins involved in a variety of transmembranous signaling processes in eukaryotic cells.8,9,28,32 These G-proteins (Gg, Gi, Go, transducin) comprise a family of highly homologous proteins of heterotrimeric structure,25 which function by utilizing a guanine nucleotide binding and GTPase cycle. The toxins ADP-ribosylate the α-subunit of the G-proteins thereby either enhancing (cholera toxin) or inhibiting (pertussis toxin) signal transduction.25