Tropomodulins are negative regulators of neurite outgrowth

Regulation of the actin cytoskeleton is critical for neurite formation. Tropomodulins (Tmods) regulate polymerization at actin filament pointed ends. Previous experiments using a mouse model deficient for the neuron specific isoform Tmod2 suggested a role for Tmods in neuronal function by impacting processes underlying learning and memory. However, the role of Tmods in neuronal function on the cellular level remains unknown. Immunofluorescence localization of the neuronal isoforms Tmod1 and Tmod2 in cultured rat primary hippocampal neurons revealed that Tmod1 is enriched along the proximal part of F-actin bundles in lamellipodia of spreading cells and in growth cones of extending neurites, while Tmod2 appears largely cytoplasmic. Functional analysis of these Tmod isoforms in a mouse neuroblastoma N2a cell line showed that knockdown of Tmod2 resulted in a significant increase in the number of neurite-forming cells and in neurite length. While N2a cells compensated for Tmod2 knockdown by increasing Tmod1 levels, over-expression of exogenous Tmod1 had no effect on neurite outgrowth. Moreover, knockdown of Tmod1 increased the number of neurites formed per cell, without effect on the number of neurite-forming cells or neurite length. Taken together, these results indicate that Tmod1 and Tmod2 have mechanistically distinct inhibitory roles in neurite formation, likely mediated via different effects on F-actin dynamics and via differential localizations during early neuritogenesis.     

Muscle development during vertebrate limb outgrowth

Limb development has become one of the model systems for studying vertebrate development. One crucial aspect in limb development is the origin, differentiation and patterning of muscle. Much progress has been made in recent years towards understanding this process. One of the general observations is that the genes involved in limb muscle development appear to be very similar to those involved in muscle development in other regions of the embryo. In this review, we summarize some of the genes and mechanisms that regulate limb muscle development and discuss various avenues along which a deeper understanding can be gained of how muscle cells originate and differentiate in different tissues during vertebrate development.     

Control of vertebrate limb outgrowth by the proximal factor Meis2 and distal antagonism of BMPs by Gremlin

The mechanisms controlling growth and patterning along the proximal-distal axis of the vertebrate limb are yet to be understood. We show that restriction of expression of the homeobox gene Meis2 to proximal regions of the limb bud is essential for limb development, since ectopic Meis2 severely disrupts limb outgrowth. We also uncover an antagonistic relationship between the secreted factors Gremlin and BMPs required to maintain the Shh/FGF loop that regulates distal outgrowth. These proximal and distal factors have coordinated activities: Meis2 can repress distal genes, and Bmps and Hoxd genes restrict Meis2 expression to the proximal limb bud. Moreover, combinations of BMPs and AER factors are sufficient to distalize proximal limb cells. Our results unveil a novel set of proximal-distal regulatory interactions that establish and maintain outgrowth of the vertebrate limb.     

Immunomodulatory mast cells: negative, as well as positive, regulators of immunity

Mast cells can promote inflammation and other tissue changes in IgE-associated allergic disorders, as well as in certain innate and adaptive immune responses that are thought to be independent of IgE. However, mast cells can also have anti-inflammatory and immunosuppressive functions. Here, we review the evidence that mast cells can have negative, as well as positive, immunomodulatory roles in vivo, and we propose that mast cells can both enhance and later suppress certain features of an immune response.     

Protein tyrosine phosphatases as negative regulators of mitogenic signaling

The regulation of tyrosine phosphorylation represents a key mechanism governing cell proliferation. In fibroblasts, inputs from both growth factor and extracellular matrix receptors are required for cell division. Triggering such receptors induces a wave of tyrosine phosphorylation on key signaling molecules, culminating in the activation of cyclin-dependent kinases and cell cycle progression. In general, protein tyrosine kinases stimulate, while protein tyrosine phosphatases inhibit, such cell proliferation pathways. The role of protein tyrosine kinases in mitogenesis has been extensively studied, but the identity and targets of the protein tyrosine phosphatases that regulate cell growth are not well described. In this review, I will survey recent advances in the identification and regulation of protein tyrosine phosphatases that downregulate cell proliferation. J. Cell. Physiol. 180:173–181, 1999. © 1999 Wiley-Liss, Inc.     

Flavonoids act as negative regulators of auxin transport in vivo in arabidopsis

Polar transport of the plant hormone auxin controls many aspects of plant growth and development. A number of synthetic compounds have been shown to block the process of auxin transport by inhibition of the auxin efflux carrier complex. These synthetic auxin transport inhibitors may act by mimicking endogenous molecules. Flavonoids, a class of secondary plant metabolic compounds, have been suggested to be auxin transport inhibitors based on their in vitro activity. The hypothesis that flavonoids regulate auxin transport in vivo was tested in Arabidopsis by comparing wild-type (WT) and transparent testa (tt4) plants with a mutation in the gene encoding the first enzyme in flavonoid biosynthesis, chalcone synthase. In a comparison between tt4 and WT plants, phenotypic differences were observed, including three times as many secondary inflorescence stems, reduced plant height, decreased stem diameter, and increased secondary root development. Growth of WT Arabidopsis plants on naringenin, a biosynthetic precursor to those flavonoids with auxin transport inhibitor activity in vitro, leads to a reduction in root growth and gravitropism, similar to the effects of synthetic auxin transport inhibitors. Analyses of auxin transport in the inflorescence and hypocotyl of independent tt4 alleles indicate that auxin transport is elevated in plants with a tt4 mutation. In hypocotyls of tt4, this elevated transport is reversed when flavonoids are synthesized by growth of plants on the flavonoid precursor, naringenin. These results are consistent with a role for flavonoids as endogenous regulators of auxin transport.     

The complex interplay between mosquito positive and negative regulators of Plasmodium development

The malaria parasite, Plasmodium, requires sexual development in the mosquito before it can be transmitted to the vertebrate host. Mosquito genes are able to substantially modulate this process, which can result in major decreases in parasite numbers. Even in susceptible mosquitoes, haemolymph proteins implicated in systemic immune reactions, together with local epithelial responses, cause lysis of more than 80% of the ookinetes that cross the mosquito midgut. In a refractory mosquito strain, immune responses lead to melanisation of virtually all parasites. Conversely, certain mosquito genes have an opposite effect: they are used by the parasite to evade defence reactions. Detailed understanding of the interplay between positive and negative regulators of parasite development could lead to the generation of novel approaches for malaria control through the vector.     

Schwann cells as regulators of nerve development.

Myelinating and non-myelinating Schwann cells of peripheral nerves are derived from the neural crest via an intermediate cell type, the Schwann cell precursor [K.R. Jessen, A. Brennan, L. Morgan, R. Mirsky, A. Kent, Y. Hashimoto, J. Gavrilovic. The Schwann cell precursor and its fate: a study of cell death and differentiation during gliogenesis in rat embryonic nerves, Neuron 12 (1994) 509-527]. The survival and maturation of Schwann cell precursors is controlled by a neuronally derived signal, beta neuregulin. Other factors, in particular endothelins, regulate the timing of precursor maturation and Schwann cell generation. In turn, signals derived from Schwann cell precursors or Schwann cells regulate neuronal numbers during development, and axonal calibre, distribution of ion channels and neurofilament phosphorylation in myelinated axons. Unlike Schwann cell precursors, Schwann cells in older nerves survive in the absence of axons, indicating that a significant change in survival regulation occurs. This is due primarily to the presence of autocrine growth factor loops in Schwann cells, present from embryo day 18 onwards, that are not functional in Schwann cell precursors. The most important components of the autocrine loop are insulin-like growth factors, platelet derived growth factor-BB and neurotrophin 3, which together with laminin support long-term Schwann cell survival. The paracrine dependence of precursors on axons for survival provides a mechanism for matching precursor cell number to axons in embryonic nerves, while the ability of Schwann cells to survive in the absence of axons is an absolute prerequisite for nerve repair following injury. In addition to providing survival factors to neurones and themselves, and signals that determine axonal architecture, Schwann cells also control the formation of peripheral nerve sheaths. This involves Schwann cell-derived Desert Hedgehog, which directs the transition of mesenchymal cells to form the epithelium-like structure of the perineurium. Schwann cells thus signal not only to themselves but also to the other cellular components within the nerve to act as major regulators of nerve development.     

Pioneer neurones use basal lamina as a substratum for outgrowth in the embryonic grasshopper limb

During axonogenesis, contacts made by the growth cone with its substratum are important in guiding the direction of neurone outgrowth. This study examines the contacts made by the growth cones of pioneer neurones in the embryonic grasshopper limb. Individual pioneer neurones at different stages of development were injected with horseradish peroxidase and the contacts made by the filopodia at the tip of their growth cones were examined by electron microscopy. Filopodia made few contacts with mesodermal cells, some contacts with ectodermal cells and very frequent contacts with basal lamina underlying the ectoderm. Components of the basal lamina may therefore play a role in guiding pioneer axon outgrowth.     

Bone morphogenetic protein signaling in limb outgrowth and patterning

Bone morphogenetic proteins (BMPs) are multifunctional growth factors belonging to the transforming growth factor beta (TGFbeta) multigene family. Current evidence indicates that they may play different and even antagonistic roles at different stages of limb development. Refined studies of their function in these processes have been impeded in the mouse due to the early lethality of null mutants for several BMP ligands and their receptors. Recently, however, these questions have benefited from the very powerful Cre-loxP technology. In this review, I intend to summarize what has been learned from this conditional mutagenesis approach in the mouse limb, focusing on Bmp2, Bmp4 and Bmp7 while restricting my analysis to the initial phases of limb formation and patterning. Two major aspects are discussed, the role of BMPs in dorsal-ventral polarization of the limb bud, together with their relation to apical ectodermal ridge (AER) induction, and their role in controlling digit number and identity. Particular attention is paid to the methodology, its power and its limits.     

A perspective on mammalian caspases as positive and negative regulators of inflammation

Members of the caspase family of cysteine proteases coordinate the morphological and biochemical events that typify apoptosis. However, neutralization of caspase activity in mammals fails to block death in response to most proapoptotic stimuli. This is because many cell death triggers provoke mitochondrial dysfunction upstream of caspase activation as a consequence of BAX/BAK channel opening. Although genetic or pharmacological inactivation of caspases fails to block cell death in most instances, it does convert the phenotype from apoptosis to necrosis. This has important implications for how the immune system responds to such cells, as necrotic cells provoke inflammation whereas apoptotic cells typically do not. Here, we propose an alternative perspective on apoptosis-associated caspase function by suggesting that these proteases are activated, not to kill, but to extinguish the proinflammatory properties of dying cells. This perspective unifies the mammalian caspase family as either positive or negative regulators of inflammation.     

PTEN and SHIP2 phosphoinositide phosphatases as negative regulators of insulin signalling

Insulin resistance in peripheral tissues is the primary cause responsible for onset of type II diabetes mellitus. Recently, the genetic and biochemical dissection of intracellular signalling pathways transducing the metabolic and mitogenic effects of insulin has contributed to the understanding of the molecular causes of this insulin resistance. In particular, important efforts have been developed to comprehend the role of negative regulators of insulin signalling, since they might represent future therapeutical targets to reduce insulin resistance in peripheral tissues. Herein, we will briefly review major intracellular signalling pathways activated by insulin and how they are negatively regulated by distinct mechanisms. In particular, the role of PTEN and SHIP2, two phosphoinositide phosphatases recently implicated as negative modulators of insulin signalling, is in focus. Current knowledge on the role of PTEN and SHIP2 in insulin resistance, type II diabetes and related disorders will also be discussed.     

Ski and SnoN: negative regulators of TGF-beta signaling

Ski and SnoN are unique proto-oncoproteins in that they can induce both oncogenic transformation and terminal muscle differentiation when expressed at high levels. Recent studies using in vitro and in vivo approaches have begun to unravel the complex roles of Ski and SnoN in tumorigenesis and embryonic development. The identification of Ski and SnoN as important negative regulators of signal transduction by the transforming growth factor-beta superfamily of cytokines provides a valuable molecular basis for the complex functions of Ski and SnoN.     

Novel short isoforms of adenylyl cyclase as negative regulators of cAMP production

Here, we cloned a new family of four adenylyl cyclase (AC) splice variants from interleukin-1β (IL-1β)-transdifferentiated vascular smooth muscle cells (VSMCs) encoding short forms of AC8 that we have named “AC8E-H”. Using biosensor imaging and biochemical approaches, we showed that AC8E-H isoforms have no cyclase activity and act as dominant-negative regulators by forming heterodimers with other full-length ACs, impeding the traffic of functional units towards the plasma membrane. The existence of these dominant-negative isoforms may account for an unsuspected additional degree of cAMP signaling regulation. It also reconciles the induction of an AC in transdifferentiated VSMCs with the vasoprotective influence of cAMP. The generation of alternative splice variants of ACs may constitute a generalized strategy of adaptation to the cell's environment whose scope had so far been ignored in physiological and/or pathological contexts.