Endocytosis and Signaling A Relationship under Development

AbstractThe ability to internalize macromolecules by endocytosis is a property of all eukaryotic cells. Frontline research on endocytosis has been presented in a successful series of biannual meetings in Europe. This year's meeting on “Membrane Dynamics in Endocytosis” was held September 13–18 in Acquafredda di Maratea, on the coast of southern Italy. Four key questions were addressed: What are the molecular mechanisms of endocytic membrane trafficking? How does endocytosis modulate receptor signaling and vice versa? What is the importance of endocytosis during development? How do endocytic organelles contribute to immunity or susceptibility to pathogens

Precision of the Dpp gradient

Morphogen concentration gradients provide positional information by activating target genes in a concentration-dependent manner. Recent reports show that the gradient of the syncytial morphogen Bicoid seems to provide precise positional information to determine target gene domains. For secreted morphogenetic ligands, the precision of the gradients, the signal transduction and the reliability of target gene expression domains have not been studied. Here we investigate these issues for the TGF-β-type morphogen Dpp. We first studied theoretically how cell-to-cell variability in the source, the target tissue, or both, contribute to the variations of the gradient. Fluctuations in the source and target generate a local maximum of precision at a finite distance to the source. We then determined experimentally in the wing epithelium: (1) the precision of the Dpp concentration gradient; (2) the precision of the Dpp signaling activity profile; and (3) the precision of activation of the Dpp target gene spalt. As captured by our theoretical description, the Dpp gradient provides positional information with a maximal precision a few cells away from the source. This maximal precision corresponds to a positional uncertainly of about a single cell diameter. The precision of the Dpp gradient accounts for the precision of the spalt expression range, implying that Dpp can act as a morphogen to coarsely determine the expression pattern of target genes

Role of Drosophila Rab5 during endosomal trafficking at the synapse and evoked neurotransmitter release

During constitutive endocytosis, internalized membrane traffics through endosomal compartments. At synapses, endocytosis of vesicular membrane is temporally coupled to action potential–induced exocytosis of synaptic vesicles. Endocytosed membrane may immediately be reused for a new round of neurotransmitter release without trafficking through an endosomal compartment. Using GFP-tagged endosomal markers, we monitored an endosomal compartment in Drosophila neuromuscular synapses. We showed that in conditions in which the synaptic vesicles pool is depleted, the endosome is also drastically reduced and only recovers from membrane derived by dynamin-mediated endocytosis. This suggests that membrane exchange takes place between the vesicle pool and the synaptic endosome. We demonstrate that the small GTPase Rab5 is required for endosome integrity in the presynaptic terminal. Impaired Rab5 function affects endo- and exocytosis rates and decreases the evoked neurotransmitter release probability. Conversely, Rab5 overexpression increases the release efficacy. Therefore, the Rab5-dependent trafficking pathway plays an important role for synaptic performance

Critical Point in Self-Organized Tissue Growth

We present a theory of pattern formation in growing domains inspired by biological examples of tissue development. Gradients of signaling molecules regulate growth, while growth changes these graded chemical patterns by dilution and advection. We identify a critical point of this feedback dynamics, which is characterized by spatially homogeneous growth and proportional scaling of patterns with tissue length. We apply this theory to the biological model system of the developing wing of the fruit fly \textit{Drosophila melanogaster} and quantitatively identify signatures of the critical point.Comment: 5 pages, 3 figure

Kinetics of Morphogen Gradient Formation

In the developing fly wing, secreted morphogens such as Decapentaplegic (Dpp) and Wingless (Wg) form gradients of concentration providing positional information. Dpp forms a longer-range gradient than Wg. To understand how the range is controlled, we measured the four key kinetic parameters governing morphogen spreading: the production rate, the effective diffusion coefficient, the degradation rate, and the immobile fraction. The four parameters had different values for Dpp versus Wg. In addition, Dynamin-dependent endocytosis was required for spreading of Dpp, but not Wg. Thus, the cellular mechanisms of Dpp and Wingless spreading are different: Dpp spreading requires endocytic, intracellular trafficking

Patrones de proliferación en el ala de "Drosophila"

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 20-11-198

Morphogen Gradient Formation

How morphogen gradients are formed in target tissues is a key question for understanding the mechanisms of morphological patterning. Here, we review different mechanisms of morphogen gradient formation from theoretical and experimental points of view. First, a simple, comprehensive overview of the underlying biophysical principles of several mechanisms of gradient formation is provided. We then discuss the advantages and limitations of different experimental approaches to gradient formation analysis

Labelling cell structures and tracking cell lineage in zebrafish using SNAP-tag

We present a method for the specific labelling of fusion proteins with synthetic fluorophores in Zebrafish. The method uses the SNAP-tag technology and O-6-benzylguanine derivatives of various synthetic fluorophores. We demonstrate how the method can be used to label subcellular structures in Zebrafish such as the nucleus, cell membranes, and endosomal membranes. The stability of the synthetic fluorophores makes them attractive choices for long-term imaging and allows, unlike most of the autofluorescent proteins, the use of acid fixatives such as trichloroacetic acid. Furthermore, the use of O-6-benzylguanine derivatives bearing caged fluorescein allows cell lineage tracing through photo-deprotection of the fluorophore and its detection either through fluorescence microscopy or through immunohistochemistry after fixation using anti-fluorescein antibodies. Developmental Dynamics 240:820-827, 2011. (C) 2011 Wiley-Liss, Inc