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Markus Affolter

Publications and source records attributed to Markus Affolter.

At least 19 recordsLinked to original sources
PubMed2006

DNA recognition by the brinker repressor--an extreme case of coupling between binding and folding.

The Brinker (Brk) nuclear repressor is a major element of the Drosophila Decapentaplegic morphogen signaling pathway. Its N-terminal part has weak homology to the Antennapedia homeodomain and binds to GC-rich DNA sequences. We have investigated the conformation and dynamics of the N-terminal 101 amino acid residues of Brk in the absence and in the presence of cognate DNA by solution NMR spectroscopy. In the absence of DNA, Brk is unfolded and highly flexible throughout the entire backbone. Addition of cognate DNA induces the formation of a well-folded structure for residues R46 to R95. This structure consists of four helices forming a helix-turn-helix motif that differs from homeodomains, but has similarities to the Tc3 transposase, the Pax-6 Paired domain, and the human centromere-binding protein. The GC-rich DNA recognition can be explained by specific major groove hydrogen bonds from the N-terminal end of helix alpha3. The transition from a highly flexible, completely unfolded conformation in the absence of DNA to a well-formed structure in the complex presents a very extreme case of the "coupling of binding and folding" phenomenon.

Amino Acid Sequence↗
PubMed2006

Polychaetoid/ZO-1 is required for cell specification and rearrangement during Drosophila tracheal morphogenesis.

The development of the complex network of epithelial tubes that ultimately forms the Drosophila tracheal system relies on cell migration, cell shape changes, cell rearrangements, cell differentiation, and branch fusion . Most of these events are controlled by a combination of distinct transcription factors and cell-cell signaling molecules, but few proteins that do not fall within these two functional classes have been associated with tracheal development. We show that the MAGUK protein Polychaetoid (Pyd/ZO-1), the Drosophila homolog of the junctional protein ZO-1 , plays a dual role in the formation of tracheal tubes. pyd/ZO-1 mutant embryos display branch fusion defects due to the lack of reliable determination of the fusion cell fate. In addition, pyd/ZO-1 mutant embryos show impaired cell intercalation in thin tracheal branches. Pyd/ZO-1 localizes to the adherens junctions (AJs) in tracheal cells and might thus play a direct role in the regulation of the dynamic state of the AJ during epithelial remodeling. Our study suggests that MAGUK proteins might play important roles during AJ remodeling in epithelial morphogenesis.

Adherens Junctions↗
PubMed2006

Remodelling epithelial tubes through cell rearrangements: from cells to molecules.

Epithelial cell movements, such as those that occur during cell intercalation, largely contribute to the formation of epithelial structures during the morphogenesis of multicellular organisms. As the architecture of epithelial tissues relies on strong adhesion between cells at adherens junctions (AJs), the intercalation or rearrangements of epithelial cells might be controlled by modulating the adhesion dynamics of the AJs by internal or external forces. In this review, we describe recent progress in understanding cell rearrangements during epithelial tube remodelling and discuss several models that might account for the developmental control of the spatial dynamics of AJs.

Adherens Junctions↗
PubMed2005

Renal tubule development in Drosophila: a closer look at the cellular level.

The function of excretion in insects is performed by the Malpighian tubules, a functional equivalent of the vertebrate kidney. Malpighian tubules are long, thin tubes connected to the hindgut. Upon the determination of the Malpighian tubule major cell type early in embryogenesis, the tubular architecture is achieved by extensive cell division and cell rearrangements. During the tube elongation process, cells exchange their neighbors, allowing the short and fat Malpighian tubule primordia to grow and become a thin tube. Cell rearrangement and intercalation underlie the morphogenesis of other epithelial tissues in Drosophila melanogaster, such as the embryonic epidermis. Recent work has provided insights in the cellular and molecular basis of cell intercalation. These advances are reviewed and discussed with regard to what is known about Malpighian tubule morphogenesis. Mature Malpighian tubules are composed of two cell types, each having a specific function in excretion: The principal cells and the stellate cells. Drosophila and mammalian kidney development show striking similarities, as the recruitment of the stellate cells to the Malpighian tubules, like the cells of the metanephric mesenchyme, requires that cells undergo a mesenchymal-to-epithelial transition. The molecular similarities between these two cases is reviewed here.

Animals↗
PubMed2005

Signaling to cytoskeletal dynamics during chemotaxis.

We review insights in signaling pathways controlling cell polarization and cytoskeletal organization during chemotactic movement in Dictyostelium amoebae and neutrophils. We compare and contrast these insights with our current understanding of pathways controlling chemotactic movements in more-complex multicellular developmental contexts.

Actins↗
PubMed2005

Distinct roles for two receptor tyrosine kinases in epithelial branching morphogenesis in Drosophila.

Branching morphogenesis is a widespread mechanism used to increase the surface area of epithelial organs. Many signaling systems steer development of branched organs, but it is still unclear which cellular processes are regulated by the different pathways. We have used the development of the air sacs of the dorsal thorax of Drosophila to study cellular events and their regulation via cell-cell signaling. We find that two receptor tyrosine kinases play important but distinct roles in air sac outgrowth. Fgf signaling directs cell migration at the tip of the structure, while Egf signaling is instrumental for cell division and cell survival in the growing epithelial structure. Interestingly, we find that Fgf signaling requires Ras, the Mapk pathway, and Pointed to direct migration, suggesting that both cytoskeletal and nuclear events are downstream of receptor activation. Ras and the Mapk pathway are also needed for Egf-regulated cell division/survival, but Pointed is dispensable.

Animals↗
PubMed2005

The Drosophila Extradenticle and Homothorax selector proteins control branchless/FGF expression in mesodermal bridge-cells.

The stereotyped outgrowth of tubular branches of the Drosophila tracheal system is orchestrated by the local and highly dynamic expression profile of branchless (bnl), which encodes a secreted fibroblast growth factor (FGF)-like molecule. Despite the importance of the spatial and temporal bnl regulation, little is known about the upstream mechanisms that establish its complex expression pattern. Here, we show that the Extradenticle and Homothorax selector proteins control bnl transcription in a single cell per segment, the mesodermal bridge-cell. In addition, we observed that a key determinant of bridge-cell specification, the transcription factor Hunchback, is also required for bnl expression. Therefore, we propose that one of the functions of the bridge-cell is to synthesize and secrete the chemoattractant Bnl. These findings provide a hitherto unknown and interesting link between combinatorial inputs of transcription factors, cell-specific ligand expression and organ morphogenesis.

Alleles↗
PubMed2005

Recognition of distinct target sites by a unique Labial/Extradenticle/Homothorax complex.

Hox genes encode evolutionarily conserved transcriptional regulators, which define regional identities along the anteroposterior axis of multicellular animals. In Drosophila, Hox proteins bind to target DNA sequences in association with the Extradenticle (Exd) and Homothorax (Hth) co-factors. The current model of Hox-binding selectivity proposes that the nucleotide sequence identity defines the Hox protein engaged in the trimeric complex, implying that distinct Hox/Exd/Hth complexes select different binding sites and that a given Hox/Exd/Hth complex recognizes a consensus DNA sequence. Here, we report that the regulation of a newly identified Lab target gene does not rely on the previously established consensus Lab/Exd/Hth-binding site, but on a strongly divergent sequence. Thus Lab, and most probably other Hox proteins, selects different DNA sequences in regulating downstream target genes. These observations have implications with regard to the current model of Hox-binding selectivity.

Animals↗
PubMed2004

Genetic control of cell intercalation during tracheal morphogenesis in Drosophila.

BACKGROUND: Branching morphogenesis transforms an epithelial sheet into a tubular network with distinct features regarding the length and diameter of individual tubes. Branching is controlled by several signaling pathways, but the molecular consequences of these pathways in the responding cells are poorly understood. RESULTS: We have undertaken a detailed characterization of cell rearrangements during tracheal branching morphogenesis in Drosophila embryos with a GFP fusion protein labeling the adherens junctions (AJs) and high-resolution live imaging. To analyze the branching process at the cellular level, we further developed an imaging approach that allows us to follow single cells during the branching process. We find that controlled cell intercalation, which requires extensive AJ remodeling, is key to the formation of tracheal branches of different cellular complexities. In particular, most branches consist of tubes with individual cells wrapped around the lumen. These branches form through cell intercalation, which requires the transformation of most of the initial intercellular AJs into autocellular AJs. We propose a step-wise model explaining how this AJ remodeling occurs and use this model to better understand defects in various mutants. We find that Dpp and Wnt signaling control cell intercalation by regulating the expression of Spalt, a zinc finger transcription factor; Spalt inhibits intercalation, leading to the formation of large, multicellular tubes. CONCLUSION: Tracheal morphogenesis is regulated by an interplay of different signaling systems that control cell migration and cell intercalation, respectively. Only the combined action of these signaling systems allows efficient branch elongation and the formation of morphologically distinct branches.

Adherens Junctions↗
PubMed2004

Drosophila gastrulation: identification of a missing link.

Drosophila gastrulation serves as a model system to elucidate the genetic and molecular mechanisms involved in morphogenetic movements. The ligand of the FGF receptor Heartless, which is involved in mesoderm movement, has now been isolated and shown to be a link between a morphogen gradient and cell behavior.

Animals↗
PubMed2004

A simple molecular complex mediates widespread BMP-induced repression during Drosophila development.

The spatial and temporal control of gene expression during the development of multicellular organisms is regulated to a large degree by cell-cell signaling. We have uncovered a simple mechanism through which Dpp, a TGFbeta/BMP superfamily member in Drosophila, represses many key developmental genes in different tissues. A short DNA sequence, a Dpp-dependent silencer element, is sufficient to confer repression of gene transcription upon Dpp receptor activation and nuclear translocation of Mad and Medea. Transcriptional repression does not require the cooperative action of cell type-specific transcription factors but relies solely on the capacity of the silencer element to interact with Mad and Medea and to subsequently recruit the zinc finger-containing repressor protein Schnurri. Our findings demonstrate how the Dpp pathway can repress key targets in a simple and tissue-unrestricted manner in vivo and hence provide a paradigm for the inherent capacity of a signaling system to repress transcription upon pathway activation.

Alleles↗
PubMed2004

A family of genes encoding zona pellucida (ZP) domain proteins is expressed in various epithelial tissues during Drosophila embryogenesis.

Zona pellucida (ZP) domain proteins have been identified in various species from worms to humans. Most of the characterized ZP family members are secreted or remain anchored to the plasma membrane where they play a structural role and/or act as receptors. In humans, several ZP proteins attracted attention because of their abundant expression in certain organs and their relation to various diseases. Here, we compare the molecular architecture and embryonic expression pattern of the 18 genes encoding ZP proteins in Drosophila melanogaster. Only five of these genes have been genetically characterized. All ZP genes are expressed in the embryo in epithelial tissues, such as the foregut, the hindgut, the Malpighian tubules, the salivary glands, the tracheal system, sensory organs and epidermis. Five genes are expressed during oogenesis; two of them are transcribed in the follicular epithelium, but not in the germ line cells.

Animals↗
PubMed2004

Downstream-of-FGFR is a fibroblast growth factor-specific scaffolding protein and recruits Corkscrew upon receptor activation.

Fibroblast growth factor (FGF) receptor (FGFR) signaling controls the migration of glial, mesodermal, and tracheal cells in Drosophila melanogaster. Little is known about the molecular events linking receptor activation to cytoskeletal rearrangements during cell migration. We have performed a functional characterization of Downstream-of-FGFR (Dof), a putative adapter protein that acts specifically in FGFR signal transduction in Drosophila. By combining reverse genetic, cell culture, and biochemical approaches, we demonstrate that Dof is a specific substrate for the two Drosophila FGFRs. After defining a minimal Dof rescue protein, we identify two regions important for Dof function in mesodermal and tracheal cell migration. The N-terminal 484 amino acids are strictly required for the interaction of Dof with the FGFRs. Upon receptor activation, tyrosine residue 515 becomes phosphorylated and recruits the phosphatase Corkscrew (Csw). Csw recruitment represents an essential step in FGF-induced cell migration and in the activation of the Ras/MAPK pathway. However, our results also indicate that the activation of Ras is not sufficient to activate the migration machinery in tracheal and mesodermal cells. Additional proteins binding either to the FGFRs, to Dof, or to Csw appear to be crucial for a chemotactic response.

Animals↗
PubMed2004

Cellular and molecular mechanisms involved in branching morphogenesis of the Drosophila tracheal system.

Recent comparative studies have shown that, in many instances, the genetic network underlying the development of distinct organ systems is similar in invertebrate and vertebrate organisms. Genetically well-characterized, simple invertebrate model systems, such as Caenorhabditis elegans and Drosophila melanogaster, can thus provide useful insight for understanding more complex organ systems in vertebrates. Here, we summarize recent progress in the genetic analysis of tracheal development in Drosophila and compare the results to studies aimed at a better understanding of lung development in mouse and man. Clearly, both striking similarities and important differences are apparent, but it might still be too early to conclude whether the former or the latter prevail.

Animals↗
PubMed2003

Epithelial tube morphogenesis during Drosophila tracheal development requires Piopio, a luminal ZP protein.

The formation of branched epithelial networks is fundamental to the development of many organs, such as the lung, the kidney or the vasculature. Little is known about the mechanisms that control cell rearrangements during tubulogenesis and regulate the size of individual tubes. Recent studies indicate that whereas the basal surface of tube cells interacts with the surrounding tissues and helps to shape the ramification pattern of tubular organs, the apical surface has an important role in the regulation of tube diameter and tube growth. Here we report that two proteins, Piopio (Pio) and Dumpy (Dp), containing a zona pellucida (ZP) domain are essential for the generation of the interconnected tracheal network in Drosophila melanogaster. Pio is secreted apically, accumulates in the tracheal lumen and possibly interacts with Dp through the ZP domains. In the absence of Pio and Dp, multicellular tubes do not rearrange through cell elongation and cell intercalation to form narrow tubes with autocellular junctions; instead they are transformed into multicellular cysts, which leads to a severe disruption of the branched pattern. We propose that an extracellular matrix containing Pio and Dp provides a structural network in the luminal space, around which cell rearrangements can take place in an ordered fashion without losing interconnections. Our results suggest that a similar structural role might be attributed to other ZP-domain proteins in the formation of different branched organs.

Adherens Junctions↗