Positive and negative regulation of Easter, a member of the serine protease family that controls dorsal-ventral patterning in the Drosophila embryo

Development ◽  
1998 ◽  
Vol 125 (7) ◽  
pp. 1261-1267 ◽  
Author(s):  
S. Misra ◽  
P. Hecht ◽  
R. Maeda ◽  
K.V. Anderson
Keyword(s):  
Serine Proteases ◽  
Feedback Loop ◽  
Early Embryo ◽  
High Molecular Mass ◽  
Drosophila Embryo ◽  
Zymogen Activation ◽  
Cell Fates ◽  
Biochemical Processes ◽  
Serine Protease Family

The sequential activities of four members of the trypsin family of extracellular serine proteases are required for the production of the ventrally localized ligand that organizes the dorsal-ventral pattern of the Drosophila embryo. The last protease in this sequence is encoded by easter, which is a candidate to activate proteolytically the ligand encoded by spatzle. Here, we demonstrate biochemically that the zymogen form of Easter is processed in vivo by a proteolytic cleavage event that requires the three upstream proteases. Processed Easter is present in extremely low amounts in the early embryo because it is rapidly converted into a high molecular mass complex, which may contain a protease inhibitor. Easter zymogen activation is also controlled by a negative feedback loop from Dorsal, the transcription factor at the end of the signaling pathway. Each of these regulated biochemical processes is likely to be important in generating the ventral-to-dorsal gradient of Dorsal protein that organizes cell fates in the early embryo.

Spatial regulation of Drosophila snake protease activity in the generation of dorsal-ventral polarity

Development ◽  
1995 ◽  
Vol 121 (12) ◽  
pp. 4127-4135 ◽  
Author(s):  
C.L. Smith ◽  
H. Giordano ◽  
M. Schwartz ◽  
R. DeLotto
Keyword(s):  
Serine Proteases ◽  
Signal Transduction Pathway ◽  
Positional Information ◽  
Biochemical Properties ◽  
Site Directed Mutagenesis ◽  
Drosophila Embryo ◽  
Perivitelline Space ◽  
Zymogen Activation

Positional information along the dorsal-ventral axis of the Drosophila embryo is acquired through a signal transduction pathway which employs a extracellular protease cascade. The sequential activation of serine protease zymogens results in the ventrally localized production of a ligand in the perivitelline space of the embryo. Snake is one of several serine proteases which function in generating the ventralizing signal. Here, we investigate the biochemical properties of Snake in vivo and in vitro using recombinant forms of the protease. Wild-type Snake zymogen completely rescues embryos from snake null females when microinjected into the perivitelline space. Biochemical evidence for a covalently associated two-chain form of the activated protease is presented. The contribution of the activation peptide region to zymogen activation was addressed using site-directed mutagenesis. The phenotypic rescue properties of an autoactivated form of Snake reveal that the covalently associated proenzyme polypeptide chain suppresses a dominant effect associated with the activated catalytic chain alone. Recombinant active catalytic chain was produced and found to be short lived as a recombinant protein. These results suggest a model in which the proenzyme polypeptide both stabilizes and targets the Snake catalytic chain to a ventrally localized activation complex within the perivitelline space.

An activity gradient of decapentaplegic is necessary for the specification of dorsal pattern elements in the Drosophila embryo

Development ◽  
1993 ◽  
Vol 117 (2) ◽  
pp. 807-822 ◽  
Author(s):  
K.A. Wharton ◽  
R.P. Ray ◽  
W.M. Gelbart
Keyword(s):  
Cell Fate ◽  
Gene Dosage ◽  
Early Embryo ◽  
Drosophila Embryo ◽  
Cell Fates ◽  
Gene Encoding ◽  
In The Wild ◽  
Intercellular Signalling ◽  
Overlapping Domains ◽  
Dorsal Pattern

decapentaplegic (dpp) is a zygotically expressed gene encoding a TGF-beta-related ligand that is necessary for dorsal-ventral patterning in the Drosophila embryo. We show here that dpp is an integral part of a gradient that specifies many different cell fates via intercellular signalling. There is a graded requirement for dpp activity in the early embryo: high levels of dpp activity specify the amnioserosa, while progressively lower levels specify dorsal and lateral ectoderm. This potential for dpp to specify cell fate is highly dosage sensitive. In the wild-type embryo, increasing the gene dosage of dpp can shift cell fates along the dorsal-ventral axis. Furthermore, in mutant embryos, in which only a subset of the dorsal-ventral pattern elements are represented, increasing the gene dosage of dpp can specifically transform those pattern elements into more dorsal ones. We present evidence that the zygotic dpp gradient and the maternal dorsal gradient specify distinct, non-overlapping domains of the dorsal-ventral pattern.

Activation of the easter zymogen is regulated by five other genes to define dorsal-ventral polarity in the Drosophila embryo

Development ◽  
1992 ◽  
Vol 115 (2) ◽  
pp. 607-616 ◽  
Author(s):  
R. Chasan ◽  
Y. Jin ◽  
K.V. Anderson
Keyword(s):  
Maternal Effect ◽  
Serine Proteases ◽  
Drosophila Embryo ◽  
Vitelline Membrane ◽  
Mutant Form ◽  
Wild Type ◽  
Zymogen Activation ◽  
Embryonic Polarity ◽  
Dominant Mutant ◽  
Maternal Effect Genes

The product of the Drosophila easter gene, a member of the trypsin family of serine proteases, must be more active ventrally than dorsally to promote normal embryonic polarity. The majority of the easter protein in the embryo is present in the unprocessed zymogen form and appears to be evenly distributed in the extracellular space, indicating that the asymmetric activity of wild-type easter must arise post-translationally. A dominant mutant form of easter that does not require cleavage of the zymogen for activity (ea delta N) is active both dorsally and ventrally. The ea delta N mutant bypasses the requirement for five other maternal effect genes, indicating that these five genes exert their effects on dorsal-ventral patterning solely by controlling the activation of the easter zymogen. We propose that dorsal-ventral asymmetry is initiated by a ventrally-localized molecule in the vitelline membrane that nucleates an easter zymogen activation complex, leading to the production of ventrally active easter enzyme.

Regulation of protein synthesis in germinating wheat embryos under polyethylene glycol and salt stress

1992 ◽  
Vol 2 (2) ◽  
pp. 75-80 ◽  
Author(s):  
A. Dell'Aquila ◽  
P. Spada
Keyword(s):  
Protein Synthesis ◽  
Salt Stress ◽  
Polyethylene Glycol ◽  
Stress Proteins ◽  
Early Embryo ◽  
Biochemical Processes ◽  
Radicle Emergence ◽  
Wheat Embryos ◽  
Emergence Phase

AbstractWheat seeds were imbibed in solutions of polyethylene glycol (PEG) and NaCl, and embryo water content and incorporation of [35S]methionine into proteins were determined. Both osmotica reduced water uptake and protein synthesis, though these were immediately resumed upon removal of stress. Results of in vivo labelling of newly synthesized proteins showed differences in the synthesis of many polypeptides when embryos that had imbibed in water passed from the radicle pre-emergence phase to early growth. The synthesis of a group of proteins associated with radicle emergence was reduced during imbibition in PEG or NaCl. All the major proteins of the pre-germination phase were produced in PEG-treated embryos, while some new polypeptides (‘salt stress’ proteins?) were actively synthesized in salt-treated embryos. Upon removal of PEG or NaCl, synthesis of proteins common to the phase of early embryo growth increased, while specific saltimbibition proteins disappeared. These findings are consistent with the hypothesis that biochemical processes leading to radicle emergence can be affected by osmotic stress and are likely to be damaged by severe stress.

Autonomous concentration-dependent activation and repression of Kruppel by hunchback in the Drosophila embryo

Development ◽  
1994 ◽  
Vol 120 (10) ◽  
pp. 3043-3049 ◽  
Author(s):  
C. Schulz ◽  
D. Tautz
Keyword(s):  
Target Gene ◽  
Target Genes ◽  
Early Embryo ◽  
Drosophila Embryo ◽  
Gap Gene ◽  
Low Concentrations ◽  
Localization Signal ◽  
Regulatory Effects ◽  
High Concentrations

The subdivision of the anterior-posterior axis in Drosophila is achieved by a cascade of spatially regulated transcription factors which form short-range gradients at the syncytial blastoderm stage. These factors are assumed to have concentration-dependent regulatory effects on their target genes. However, there is so far little direct in vivo evidence that a single factor can autonomously activate and repress a given target gene. We have analysed here the regulatory capabilities of the gap gene hunchback by creating an artificial gradient of hunchback in the early embryo. This was achieved by providing the maternally expressed mRNA of hunchback with the anterior localization signal of the bicoid RNA. The effects of this artificial hunchback gradient were then studied in different types of mutant background. We show that under these conditions hb is autonomously capable of activating the target gene Kruppel at low concentrations and repressing it at high concentrations. In addition, we show that the artificially created hunchback gradient can organize a large part of the segment pattern, although it is expressed at a different position and in a different shape than the wild-type gradient of hunchback.

The Drosophila Medea gene is required downstream of dpp and encodes a functional homolog of human Smad4

Development ◽  
1998 ◽  
Vol 125 (8) ◽  
pp. 1407-1420 ◽  
Author(s):  
J.B. Hudson ◽  
S.D. Podos ◽  
K. Keith ◽  
S.L. Simpson ◽  
E.L. Ferguson
Keyword(s):  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Sequence Similarity ◽  
Early Embryo ◽  
Drosophila Embryo ◽  
Phenotypic Analysis ◽  
Cell Fates ◽  
Functional Homolog ◽  
Phenotypic Rescue ◽  
Embryonic Ectoderm

The Transforming Growth Factor-beta superfamily member decapentaplegic (dpp) acts as an extracellular morphogen to pattern the embryonic ectoderm of the Drosophila embryo. To identify components of the dpp signaling pathway, we screened for mutations that act as dominant maternal enhancers of a weak allele of the dpp target gene zerknLllt. In this screen, we recovered new alleles of the Mothers against dpp (Mad) and Medea genes. Phenotypic analysis of the new Medea mutations indicates that Medea, like Mad, is required for both embryonic and imaginal disc patterning. Genetic analysis suggests that Medea may have two independently mutable functions in patterning the embryonic ectoderm. Complete elimination of maternal and zygotic Medea activity in the early embryo results in a ventralized phenotype identical to that of null dpp mutants, indicating that Medea is required for all dpp-dependent signaling in embryonic dorsal-ventral patterning. Injection of mRNAs encoding DPP or a constitutively activated form of the DPP receptor, Thick veins, into embryos lacking all Medea activity failed to induce formation of any dorsal cell fates, demonstrating that Medea acts downstream of the thick veins receptor. We cloned Medea and found that it encodes a protein with striking sequence similarity to human SMAD4. Moreover, injection of human SMAD4 mRNA into embryos lacking all Medea activity conferred phenotypic rescue of the dorsal-ventral pattern, demonstrating conservation of function between the two gene products.

Laser scanning confocal microscopic analysis of cytoskeletal and nuclear reorganization in live Drosophila embryos

1993 ◽  
Vol 51 ◽  
pp. 174-175
Author(s):  
William Theurkauf
Keyword(s):  
Laser Scanning ◽  
Microscopic Analysis ◽  
Large Cell ◽  
Early Embryo ◽  
Drosophila Embryo ◽  
Cortical Actin ◽  
Nuclear Reorganization ◽  
Cytoskeletal Elements ◽  
Microtubule Structure ◽  
Drosophila Embryos

Cell division in eucaryotes depends on coordinated changes in nuclear and cytoskeletal components. In Drosophila melanogaster embryos, the first 13 nuclear divisions occur without cytokinesis. During the final four divisions, nuclei divide in a uniform monolayer at the surface of the embryo. These surface divisions are accompanied by dramatic changes in cortical actin and microtubule structure (Karr and Alberts, 1986), and inhibitor studies indicate that these changes are essential to orderly mitosis (Zalokar and Erk, 1976). Because the early embryo is syncytial, fluorescent probes introduced by microinjection are incorporated in structures associated with all of the nuclei in the blastoderm. In addition, the nuclei divide synchronously every 10 to 20 min. These characteristics make the syncytial blastoderm embryo an excellent system for the analysis of mitotic reorganization of both nuclear and cytoskeletal elements. However, the Drosophila embryo is a large cell, and resolution of cytoskeletal filaments and nuclear structure is hampered by out-of focus signal.

scholarly journals Phenotypic Analysis of Mice Lacking the Tmprss2-Encoded Protease

2006 ◽  
Vol 26 (3) ◽  
pp. 965-975 ◽  
Author(s):  
Tom S. Kim ◽  
Cynthia Heinlein ◽  
Robert C. Hackman ◽  
Peter S. Nelson
Keyword(s):  
Serine Protease ◽  
Serine Proteases ◽  
Biological Activities ◽  
Type Ii ◽  
Phenotypic Analysis ◽  
Normal Prostate ◽  
Prostate Hyperplasia ◽  
Transmembrane Serine Protease

ABSTRACT Tmprss2 encodes an androgen-regulated type II transmembrane serine protease (TTSP) expressed highly in normal prostate epithelium and has been implicated in prostate carcinogenesis. Although in vitro studies suggest protease-activated receptor 2 may be a substrate for TMPRSS2, the in vivo biological activities of TMPRSS2 remain unknown. We generated Tmprss2 −/− mice by disrupting the serine protease domain through homologous recombination. Compared to wild-type littermates, Tmprss2 −/− mice developed normally, survived to adulthood with no differences in protein levels of prostatic secretions, and exhibited no discernible abnormalities in organ histology or function. Loss of TMPRSS2 serine protease activity did not influence fertility, reduce survival, result in prostate hyperplasia or carcinoma, or alter prostatic luminal epithelial cell regrowth following castration and androgen replacement. Lack of an observable phenotype in Tmprss2 −/− mice was not due to transcriptional compensation by closely related Tmprss2 homologs. We conclude that the lack of a discernible phenotype in Tmprss2 −/− mice suggests functional redundancy involving one or more of the type II transmembrane serine protease family members or other serine proteases. Alternatively, TMPRSS2 may contribute a specialized but nonvital function that is apparent only in the context of stress, disease, or other systemic perturbation.

scholarly journals LncRNA MIR17HG promotes colorectal cancer liver metastasis by mediating a glycolysis-associated positive feedback circuit

Oncogene ◽  
2021 ◽  
Author(s):  
Senlin Zhao ◽  
Bingjie Guan ◽  
Yushuai Mi ◽  
Debing Shi ◽  
Ping Wei ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Liver Metastasis ◽  
Positive Feedback ◽  
Feedback Loop ◽  
Metastatic Tumor ◽  
Feedback Circuit ◽  
Colorectal Cancer Liver Metastasis ◽  
Positive Feedback Circuit

AbstractGlycolysis plays a crucial role in reprogramming the metastatic tumor microenvironment. A series of lncRNAs have been identified to function as oncogenic molecules by regulating glycolysis. However, the roles of glycolysis-related lncRNAs in regulating colorectal cancer liver metastasis (CRLM) remain poorly understood. In the present study, the expression of the glycolysis-related lncRNA MIR17HG gradually increased from adjacent normal to CRC to the paired liver metastatic tissues, and high MIR17HG expression predicted poor survival, especially in patients with liver metastasis. Functionally, MIR17HG promoted glycolysis in CRC cells and enhanced their invasion and liver metastasis in vitro and in vivo. Mechanistically, MIR17HG functioned as a ceRNA to regulate HK1 expression by sponging miR-138-5p, resulting in glycolysis in CRC cells and leading to their invasion and liver metastasis. More interestingly, lactate accumulated via glycolysis activated the p38/Elk-1 signaling pathway to promote the transcriptional expression of MIR17HG in CRC cells, forming a positive feedback loop, which eventually resulted in persistent glycolysis and the invasion and liver metastasis of CRC cells. In conclusion, the present study indicates that the lactate-responsive lncRNA MIR17HG, acting as a ceRNA, promotes CRLM through a glycolysis-mediated positive feedback circuit and might be a novel biomarker and therapeutic target for CRLM.

scholarly journals The AF-6 Homolog Canoe Acts as a Rap1 Effector During Dorsal Closure of the Drosophila Embryo

Genetics ◽  
2003 ◽  
Vol 165 (1) ◽  
pp. 159-169
Author(s):  
Benjamin Boettner ◽  
Phoebe Harjes ◽  
Satoshi Ishimaru ◽  
Michael Heke ◽  
Hong Qing Fan ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Genetic Interaction ◽  
Critical Role ◽  
Adherens Junction ◽  
Small Gtpases ◽  
Drosophila Embryo ◽  
Dorsal Closure ◽  
Cell Sheets ◽  
Jnk Pathway

Abstract Rap1 belongs to the highly conserved Ras subfamily of small GTPases. In Drosophila, Rap1 plays a critical role in many different morphogenetic processes, but the molecular mechanisms executing its function are unknown. Here, we demonstrate that Canoe (Cno), the Drosophila homolog of mammalian junctional protein AF-6, acts as an effector of Rap1 in vivo. Cno binds to the activated form of Rap1 in a yeast two-hybrid assay, the two molecules colocalize to the adherens junction, and they display very similar phenotypes in embryonic dorsal closure (DC), a process that relies on the elongation and migration of epithelial cell sheets. Genetic interaction experiments show that Rap1 and Cno act in the same molecular pathway during DC and that the function of both molecules in DC depends on their ability to interact. We further show that Rap1 acts upstream of Cno, but that Rap1, unlike Cno, is not involved in the stimulation of JNK pathway activity, indicating that Cno has both a Rap1-dependent and a Rap1-independent function in the DC process.

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