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G Struhl

Publications and source records attributed to G Struhl.

At least 19 recordsLinked to original sources
PubMed2001

Presenilin-mediated transmembrane cleavage is required for Notch signal transduction in Drosophila.

The cleavage model for signal transduction by receptors of the LIN-12/Notch family posits that ligand binding leads to cleavage within the transmembrane domain, so that the intracellular domain is released to translocate to the nucleus and activate target gene expression. The familial Alzheimer's disease-associated protein Presenilin is required for LIN-12/Notch signaling, and several lines of evidence suggest that Presenilin mediates the transmembrane cleavage event that releases the LIN-12/Notch intracellular domain. However, doubt was cast on this possibility by a report that Presenilin is not required for the transducing activity of N(ECN), a constitutively active transmembrane form of Notch, in Drosophila. Here, we have reassessed this finding and show instead that Presenilin is required for activity of N(ECN) for all cell fate decisions examined. Our results indicate that transmembrane cleavage and signal transduction are strictly correlated, supporting the cleavage model for signal transduction by LIN-12/Notch and a role for Presenilin in mediating the ligand-induced transmembrane cleavage.

Active Transport, Cell Nucleus↗
PubMed2001

Delta/Notch and Boss/Sevenless signals act combinatorially to specify the Drosophila R7 photoreceptor.

The R7 photoreceptor, a unique cell type within the Drosophila ommatidium, was initially proposed to be specified by two distinct signals from neighboring cells, one from the R8 photoreceptor and another from the R1/6 photoreceptor pair. The R8-to-R7 signal is the transmembrane ligand Bride of Sevenless (Boss), which is received by the receptor tyrosine kinase Sevenless (Sev) and transduced via Ras activation within the presumptive R7 cell. However, the identity of the R1/6-to-R7 signal has remained elusive. Here, we present evidence that the transmembrane ligand Delta (Dl), expressed by the R1/6 pair, activates the receptor Notch (N) in the presumptive R7 cell and constitutes the postulated R1/6-to-R7 signal required in combination with the Boss/Sev signal to specify the R7 fate.

Animals↗
PubMed2001

A hedgehog-insensitive form of patched provides evidence for direct long-range morphogen activity of sonic hedgehog in the neural tube.

Cell pattern in the ventral neural tube is organized by Sonic hedgehog (Shh) secreted by floor plate cells. To assay the range of direct Shh action, we developed a general method for blocking transduction of Hedgehog (Hh) signals through ectopic expression of a deleted form of the Hh receptor Patched (Ptc), termed Ptc(Deltaloop2). We validated this method in Drosophila and used mouse Ptc1(Deltaloop2) (mPtc1(Deltaloop2)) to block Shh transduction in the chick neural tube. mPtc1(Deltaloop2) expression caused cell-autonomous ventral-to-dorsal switches in progenitor identity and neuronal fate throughout the ventral neural tube, supporting a gradient mechanism whereby Shh acts directly and at long range. mPtc1(Deltaloop2) expression also caused the abnormal spread of Shh to more dorsal cells, indicating that Shh in the neural tube, like Hh in Drosophila, induces a feedback mechanism that limits its range of action.

Animals↗
PubMed2001

Nicastrin is required for Presenilin-mediated transmembrane cleavage in Drosophila.

The transmembrane glycoprotein Nicastrin was identified in a complex with the multipass membrane protein Presenilin. Presenilin mediates transmembrane cleavage of single-pass transmembrane proteins with short extracellular domains, including the ligand-activated form of the receptor Notch and beta-amyloid precursor protein (beta-APP). Transmembrane cleavage of Notch is essential for signal transduction, and transmembrane cleavage of beta-APP generates pathogenic amyloid peptides implicated in Alzheimer's disease. Here, we investigate the requirement for Nicastrin in Presenilin-mediated transmembrane cleavage. We show that, in Drosophila, loss of Nicastrin activity blocks the accumulation of Presenilin associated with the apical plasma membrane, abolishes Presenilin-dependent cleavage of the transmembrane domains of Notch and beta-APP, and abrogates Notch signal transduction.

Amyloid Precursor Protein Secretases↗
PubMed2001

Polycomb group proteins and heritable silencing of Drosophila Hox genes.

Early in Drosophila embryogenesis, transcriptional repressors encoded by Gap genes prevent the expression of particular combinations of Hox genes in each segment. During subsequent development, those Hox genes that were initially repressed in each segment remain off in all the descendent cells, even though the Gap repressors are no longer present. This phenomenon of heritable silencing depends on proteins of the Polycomb Group (PcG) and on cis-acting Polycomb response elements (PREs) in the Hox gene loci. We have removed individual PcG proteins from proliferating cells and then resupplied these proteins after a few or several cell generations. We show that most PcG proteins are required throughout development: when these proteins are removed, Hox genes become derepressed. However, we find that resupply of at least some PcG proteins can cause re-repression of Hox genes, provided that it occurs within a few cell generations of the loss of repression. These results suggest a functional distinction between transcriptional repression and heritable silencing: in at least some contexts, Hox genes can retain the capacity to be heritably silenced, despite being transcribed and replicated. We propose that silenced Hox genes bear a heritable, molecular mark that targets them for transcriptional repression. Some PcG proteins may be required to define and propagate this mark; others may function to repress the transcription of Hox genes that bear the mark.

Animals↗
PubMed2000

Requirements for presenilin-dependent cleavage of notch and other transmembrane proteins.

Ligand binding to receptors of the LIN-12/Notch family causes at least two proteolytic cleavages: one between the extracellular and transmembrane domains, and the other within the transmembrane domain. The transmembrane cleavage depends on Presenilin, a protein also required for transmembrane cleavage of beta-APP. Here, we have assayed the substrate requirements for Presenilin-dependent processing of Notch and other type I transmembrane proteins in vivo. We find that the Presenilin-dependent cleavage does not depend critically on the recognition of particular sequences in these proteins but rather on the size of the extracellular domain: the smaller the size, the greater the efficiency of cleavage. Hence, Notch, beta-APP, and perhaps other proteins may be targeted for Presenilin-mediated transmembrane cleavage by upstream processing events that sever the extracellular domain from the rest of the protein.

Amino Acid Sequence↗
PubMed1999

Presenilin is required for activity and nuclear access of Notch in Drosophila.

Presenilins are membrane proteins with multiple transmembrane domains that are thought to contribute to the development of Alzheimer's disease by affecting the processing of beta-amyloid precursor protein. Presenilins also facilitate the activity of transmembrane receptors of the LIN-12/Notch family. After ligand-induced processing, the intracellular domain of LIN-12/Notch can enter the nucleus and participate in the transcriptional control of downstream target genes. Here we show that null mutations in the Drosophila Presenilin gene abolish Notch signal transduction and prevent its intracellular domain from entering the nucleus. Furthermore, we provide evidence that presenilin is required for the proteolytic release of the intracellular domain from the membrane following activation of Notch by ligand.

Animals↗
PubMed1999

hedgehog and engrailed: pattern formation and polarity in the Drosophila abdomen.

Like the Drosophila embryo, the abdomen of the adult consists of alternating anterior (A) and posterior (P) compartments. However the wing is made by only part of one A and part of one P compartment. The abdomen therefore offers an opportunity to compare two compartment borders (A/P is within the segment and P/A intervenes between two segments), and ask if they act differently in pattern formation. In the embryo, abdomen and wing P compartment cells express the selector gene engrailed and secrete Hedgehog protein whilst A compartment cells need the patched and smoothened genes in order to respond to Hedgehog. We made clones of cells with altered activities of the engrailed, patched and smoothened genes. Our results confirm (1) that the state of engrailed, whether 'off' or 'on', determines whether a cell is of A or P type and (2) that Hedgehog signalling, coming from the adjacent P compartments across both A/P and P/A boundaries, organises the pattern of all the A cells. We have uncovered four new aspects of compartments and engrailed in the abdomen. First, we show that engrailed acts in the A compartment: Hedgehog leaves the P cells and crosses the A/P boundary where it induces engrailed in a narrow band of A cells. engrailed causes these cells to form a special type of cuticle. No similar effect occurs when Hedgehog crosses the P/A border. Second, we look at the polarity changes induced by the clones, and build a working hypothesis that polarity is organised, in both compartments, by molecule(s) emanating from the A/P but not the P/A boundaries. Third, we show that both the A and P compartments are each divided into anterior and posterior subdomains. This additional stratification makes the A/P and the P/A boundaries fundamentally distinct from each other. Finally, we find that when engrailed is removed from P cells (of, say, segment A5) they transform not into A cells of the same segment, but into A cells of the same parasegment (segment A6).

Abdomen↗
PubMed1999

The hedgehog morphogen and gradients of cell affinity in the abdomen of Drosophila.

The adult abdomen of Drosophila is a chain of anterior (A) and posterior (P) compartments. The engrailed gene is active in all P compartments and selects the P state. Hedgehog enters each A compartment across both its anterior and posterior edges; within A its concentration confers positional information. The A compartments are subdivided into an anterior and a posterior domain that each make different cell types in response to Hedgehog. We have studied the relationship between Hedgehog, engrailed and cell affinity. We made twin clones and measured the shape, size and displacement of the experimental clone, relative to its control twin. We varied the perceived level of Hedgehog in the experimental clone and find that, if this level is different from the surround, the clone fails to grow normally, rounds up and sometimes sorts out completely, becoming separated from the epithelium. Also, clones are displaced towards cells that are more like themselves: for example groups of cells in the middle of the A compartment that are persuaded to differentiate as if they were at the posterior limit of A, move posteriorly. Similarly, clones in the anterior domain of the A compartment that are forced to differentiate as if they were at the anterior limit of A, move anteriorly. Quantitation of these measures and the direction of displacement indicate that there is a U-shaped gradient of affinity in the A compartment that correlates with the U-shaped landscape of Hedgehog concentration. Since affinity changes are autonomous to the clone we believe that, normally, each cell's affinity is a direct response to Hedgehog. By removing engrailed in clones we show that A and P cells also differ in affinity from each other, in a manner that appears independent of Hedgehog. Within the P compartment we found some evidence for a U-shaped gradient of affinity, but this cannot be due to Hedgehog which does not act in the P compartment.

Abdomen↗
PubMed1999

Wingless transduction by the Frizzled and Frizzled2 proteins of Drosophila.

Wingless (Wg) protein is a founding member of the Wnt family of secreted proteins which have profound organizing roles in animal development. Two members of the Frizzled (Fz) family of seven-pass transmembrane proteins, Drosophila Fz and Fz2, can bind Wg and are candidate Wg receptors. However, null mutations of the fz gene have little effect on Wg signal transduction and the lack of mutations in the fz2 gene has thus far prevented a rigorous examination of its role in vivo. Here we describe the isolation of an amber mutation of fz2 which truncates the coding sequence just after the amino-terminal extracellular domain and behaves genetically as a loss-of-function allele. Using this mutation, we show that Wg signal transduction is abolished in virtually all cells lacking both Fz and Fz2 activity in embryos as well as in the wing imaginal disc. We also show that Fz and Fz2 are functionally redundant: the presence of either protein is sufficient to confer Wg transducing activity on most or all cells throughout development. These results extend prior evidence of a ligand-receptor relationship between Wnt and Frizzled proteins and suggest that Fz and Fz2 are the primary receptors for Wg in Drosophila.

Alleles↗
PubMed1999

Decoding vectorial information from a gradient: sequential roles of the receptors Frizzled and Notch in establishing planar polarity in the Drosophila eye.

The Drosophila eye is composed of several hundred ommatidia that can exist in either of two chiral forms, depending on position: ommatidia in the dorsal half of the eye adopt one chiral form, whereas ommatidia in the ventral half adopt the other. Chirality appears to be specified by a polarizing signal with a high activity at the interface between the two halves (the 'equator'), which declines in opposite directions towards the dorsal and ventral poles. Here, using genetic mosaics, we show that this polarizing signal is decoded by the sequential use of two receptor systems. The first depends on the seven-transmembrane receptor Frizzled (Fz) and distinguishes between the two members of the R3/R4 pair of presumptive photoreceptor cells, predisposing the cell that is located closer to the equator and having higher Fz activity towards the R3 photoreceptor fate and the cell further away towards the R4 fate. This bias is then amplified by subsequent interactions between the two cells mediated by the receptor Notch (N) and its ligand Delta (Dl), ensuring that the equatorial cell becomes the R3 photoreceptor while the polar cell becomes the R4 photoreceptor. As a consequence of this reciprocal cell fate decision, the R4 cell moves asymmetrically relative to the R3 cell, initiating the appropriate chiral pattern of the remaining cells of the ommatidium.

Animals↗
PubMed1999

Progression of the morphogenetic furrow in the Drosophila eye: the roles of Hedgehog, Decapentaplegic and the Raf pathway.

During Drosophila eye development, Hedgehog (Hh) protein secreted by maturing photoreceptors directs a wave of differentiation that sweeps anteriorly across the retinal primordium. The crest of this wave is marked by the morphogenetic furrow, a visible indentation that demarcates the boundary between developing photoreceptors located posteriorly and undifferentiated cells located anteriorly. Here, we present evidence that Hh controls progression of the furrow by inducing the expression of two downstream signals. The first signal, Decapentaplegic (Dpp), acts at long range on undifferentiated cells anterior to the furrow, causing them to enter a 'pre-proneural' state marked by upregulated expression of the transcription factor Hairy. Acquisition of the pre-proneural state appears essential for all prospective retinal cells to enter the proneural pathway and differentiate as photoreceptors. The second signal, presently unknown, acts at short range and is transduced via activation of the Serine-Threonine kinase Raf. Activation of Raf is both necessary and sufficient to cause pre-proneural cells to become proneural, a transition marked by downregulation of Hairy and upregulation of the proneural activator, Atonal (Ato), which initiates differentiation of the R8 photoreceptor. The R8 photoreceptor then organizes the recruitment of the remaining photoreceptors (R1-R7) through additional rounds of Raf activation in neighboring pre-proneural cells. Finally, we show that Dpp signaling is not essential for establishing either the pre-proneural or proneural states, or for progression of the furrow. Instead, Dpp signaling appears to increase the rate of furrow progression by accelerating the transition to the pre-proneural state. In the abnormal situation in which Dpp signaling is blocked, Hh signaling can induce undifferentiated cells to become pre-proneural but does so less efficiently than Dpp, resulting in a retarded rate of furrow progression and the formation of a rudimentary eye.

Animals↗
PubMed1998

Nuclear access and action of notch in vivo.

The Drosophila Notch (N) gene encodes a conserved single-pass transmembrane receptor that transduces extracellular signals controlling cell fate. Here, we present evidence that the intracellular domain of Notch gains access to the nucleus in response to ligand, possibly through a mechanism involving proteolytic cleavage and release from the remainder of the protein. In addition, our results suggest that signal transduction by Notch depends on the ability of the intracellular domain, particularly the portion containing the CDC10 repeats, to reach the nucleus and to participate in the transcriptional activation of downstream target genes.

Animals↗
PubMed1998

Regulation of the Hedgehog and Wingless signalling pathways by the F-box/WD40-repeat protein Slimb.

Members of the Hedgehog (Hh) and Wnt/Wingless (Wg) families of secreted proteins control many aspects of growth and patterning during animal development. Hh signal transduction leads to increased stability of a transcription factor, Cubitus interruptus (Ci), whereas Wg signal transduction causes increased stability of Armadillo (Arm/beta-catenin), a possible co-factor for the transcriptional regulator Lef1/TCF. Here we describe a new gene, slimb (for supernumerary limbs), which negatively regulates both of these signal transduction pathways. Loss of function of slimb results in a cell-autonomous accumulation of high levels of both Ci and Arm, and the ectopic expression of both Hh- and Wg- responsive genes. The slimb gene encodes a conserved F-box/WD40-repeat protein related to Cdc4p, a protein in budding yeast that targets cell-cycle regulators for degradation by the ubiquitin/proteasome pathway. We propose that Slimb protein normally targets Ci and Arm for processing or degradation by the ubiquitin/proteasome pathway, and that Hh and Wg regulate gene expression at least in part by inducing changes in Ci and Arm, which protect them from Slimb-mediated proteolysis.

Amino Acid Sequence↗
PubMed1998

In vivo evidence that Patched and Smoothened constitute distinct binding and transducing components of a Hedgehog receptor complex.

During Drosophila development, cells belonging to the posterior compartment of each segment organize growth and patterning by secreting Hedgehog (Hh), a protein which induces a thin strip of adjacent cells in the anterior compartment to express the morphogens Decapentaplegic (Dpp) and Wingless (Wg). Hedgehog is bound and transduced by a receptor complex that includes Smoothened (Smo), a member of the Frizzled (Fz) family of seven-pass transmembrane receptors, as well as the multiple-pass transmembrane protein Patched (Ptc). Ptc is required for the binding of Hh to the complex as well as for the Hh-dependent activation of Smo within the complex. Here, we identify a likely null allele of the smo gene and use it to determine whether Hh is bound by Ptc alone, or by Smo in concert with Ptc. We find that cells devoid of Smo can sequester Hh, but that their ability to do so depends, as in wild-type cells, on the expression of high levels of Ptc protein. These results suggest that Ptc normally binds Hh without any help from Smo and hence favor a mechanism of signal transduction in which Hh binds specifically to Ptc and induces a conformational change leading to the release of latent Smo activity.

Animals↗
PubMed1997

Hedgehog organises the pattern and polarity of epidermal cells in the Drosophila abdomen.

The abdomen of adult Drosophila, like that of other insects, is formed by a continuous epithelium spanning several segments. Each segment is subdivided into an anterior (A) and posterior (P) compartment, distinguished by activity of the selector gene engrailed (en) in P but not A compartment cells. Here we provide evidence that Hedgehog (Hh), a protein secreted by P compartment cells, spreads into each A compartment across the anterior and the posterior boundaries to form opposing concentration gradients that organize cell pattern and polarity. We find that anteriorly and posteriorly situated cells within the A compartment respond in distinct ways to Hh: they express different combinations of genes and form different cell types. They also form polarised structures that, in the anterior part, point down the Hh gradient and, in the posterior part, point up the gradient - therefore all structures point posteriorly. Finally, we show that ectopic Hh can induce cells in the middle of each A compartment to activate en. Where this happens, A compartment cells are transformed into an ectopic P compartment and reorganise pattern and polarity both within and around the transformed tissue. Many of these results are unexpected and lead us to reassess the role of gradients and compartments in patterning insect segments.

Abdomen↗
PubMed1997

Hedgehog acts by distinct gradient and signal relay mechanisms to organise cell type and cell polarity in the Drosophila abdomen.

The epidermis of the adult Drosophila abdomen is formed by a chain of anterior (A) and posterior (P) compartments, each segment comprising one A and one P compartment. In the accompanying paper (Struhl et al., 1997), we provide evidence that Hedgehog protein (Hh), being secreted from P compartment cells, organises the pattern and polarity of A compartment cells. Here we test whether Hh acts directly or by a signal relay mechanism. We use mutations in Protein Kinase A (PKA) or smoothened (smo) to activate or to block Hh signal transduction in clones of A compartment cells. For cell type, a scalar property, both manipulations cause strictly autonomous transformations: the cells affected are exactly those and only those that are mutant. Hence, we infer that Hh acts directly on A compartment cells to specify the various types of cuticular structures that they differentiate. By contrast, these same manipulations cause non-autonomous effects on cell polarity, a vectorial property. Consequently, we surmise that Hh influences cell polarity indirectly, possibly by inducing other signalling factors. Finally, we present evidence that Hh does not polarise abdominal cells by utilising either Decapentaplegic (Dpp) or Wingless (Wg), the two morphogens through which Hh acts during limb development. We conclude that, in the abdomen, cell type and cell polarity reflect distinct outputs of Hh signalling and propose that these outputs are controlled by separable gradient and signal relay mechanisms.

Abdomen↗