PubMed Health⌕ Search

Biomedical subjects

Ali H Brivanlou

Publications and source records attributed to Ali H Brivanlou.

At least 19 recordsLinked to original sources
PubMed2006

Dephosphorylation of the linker regions of Smad1 and Smad2/3 by small C-terminal domain phosphatases has distinct outcomes for bone morphogenetic protein and transforming growth factor-beta pathways.

Smad proteins transduce bone morphogenetic protein (BMP) and transforming growth factor-beta (TGFbeta) signals upon phosphorylation of their C-terminal SXS motif by receptor kinases. The activity of Smad1 in the BMP pathway and Smad2/3 in the TGFbeta pathway is restricted by pathway cross-talk and feedback through protein kinases, including MAPK, CDK2/4, p38MAPK, JNK, and others. These kinases phosphorylate Smads 1-3 at the region that links the N-terminal DNA-binding domain and the C-terminal transcriptional domain. Phosphatases that dephosphorylate the linker region are therefore likely to play an integral part in the regulation of Smad activity. We reported previously that small C-terminal domain phosphatases 1, 2, and 3 (SCP1-3) dephosphorylate Smad1 C-terminal tail, thereby attenuating BMP signaling. Here we provide evidence that SCP1-3 also dephosphorylate the linker regions of Smad1 and Smad2/3 in vitro, in mammalian cells and in Xenopus embryos. Overexpression of SCP 1, 2, or 3 decreased linker phosphorylation of Smads 1, 2 and 3. Moreover, RNA interference-mediated knockdown of SCP1/2 increased the BMP-dependent phosphorylation of the Smad1 linker region as well as the C terminus. In contrast, SCP1/2 knockdown increased the TGFbeta-dependent linker phosphorylation of Smad2/3 but not the C-terminal phosphorylation. Consequently, SCP1/2 knockdown inhibited TGFbeta transcriptional responses, but it enhanced BMP transcriptional responses. Thus, by dephosphorylating Smad2/3 at the linker (inhibitory) but not the C-terminal (activating) site, the SCPs enhance TGFbeta signaling, and by dephosphorylating Smad1 at both sites, the SCPs reset Smad1 to the basal unphosphorylated state.

Animals↗
PubMed2006

Function of the two Xenopus smad4s in early frog development.

Signals from the transforming growth factor beta family members are transmitted in the cell through specific receptor-activated Smads and a common partner Smad4. Two Smad4 genes (alpha and beta/10, or smad4 and smad4.2) have been isolated from Xenopus, and conflicting data are reported for Smad4beta/10 actions in mesodermal and neural induction. To further understand the functions of the Smad4s in early frog development, we analyzed their activities in detail. We report that Smad10 is a mutant form of Smad4beta that harbors a missense mutation of a conserved arginine to histidine in the MH1 domain. The mutation results in enhanced association of Smad10 with the nuclear transcription corepressor Ski and leads to its neural inducing activity through inhibition of bone morphogenetic protein (BMP) signaling. In contrast to Smad10, both Smad4alpha and Smad4beta enhanced BMP signals in ectodermal explants. Using antisense morpholino oligonucleotides (MOs) to knockdown endogenous Smad4 protein levels, we discovered that Smad4beta was required for both activin- and BMP-mediated mesodermal induction in animal caps, whereas Smad4alpha affected only the BMP signals. Neither Smad4 was involved directly in neural induction. Expression of Smad4beta-MO in early frog embryos resulted in reduction of mesodermal markers and defects in axial structures, which were rescued by either Smad4alpha or Smad4beta. Smad4alpha-MO induced only minor deficiency at late stages. As Smad4beta, but not Smad4alpha, is expressed at high levels maternally and during early gastrulation, our data suggest that although Smad4alpha and Smad4beta may have similar activities, they are differentially utilized during frog embryogenesis, with only Smad4beta being essential for mesoderm induction.

Activins↗
PubMed2006

Unique players in the BMP pathway: small C-terminal domain phosphatases dephosphorylate Smad1 to attenuate BMP signaling.

Smad transcription factors are key signal transducers for the TGF-beta/bone morphogenetic protein (BMP) family of cytokines and morphogens. C-terminal serine phosphorylation by TGF-beta and BMP membrane receptors drives Smads into the nucleus as transcriptional regulators. Dephosphorylation and recycling of activated Smads is an integral part of this process, which is critical for agonist sensing by the cell. However, the nuclear phosphatases involved have remained unknown. Here we provide functional, biochemical, and embryological evidence identifying the SCP (small C-terminal domain phosphatase) family of nuclear phosphatases as mediators of Smad1 dephosphorylation in the BMP signaling pathway in vertebrates. Xenopus SCP2/Os4 inhibits BMP activity in the presumptive ectoderm and leads to neuralization. In Xenopus embryos, SCP2/Os4 and human SCP1, 2, and 3 cause selective dephosphorylation of Smad1 compared with Smad2, inhibiting BMP- and Smad1-dependent transcription and leading to the induction of the secondary dorsal axis. In human cells, RNAi-mediated depletion of SCP1 and SCP2 increases the extent and duration of Smad1 phosphorylation in response to BMP, the transcriptional action of Smad1, and the strength of endogenous BMP gene responses. The present identification of the SCP family as Smad C-terminal phosphatases sheds light on the events that attenuate Smad signaling and reveals unexpected links to the essential phosphatases that control RNA polymerase II in eukaryotes.

Animals↗
PubMed2006

Contribution of human embryonic stem cells to mouse blastocysts.

In addition to their potential for cell-based therapies in the treatment of disease and injury, the broad developmental capacity of human embryonic stem cells (hESCs) offers potential for studying the origins of all human cell types. To date, the emergence of specialized cells from hESCs has commonly been studied in tissue culture or in teratomas, yet these methods have stopped short of demonstrating the ESC potential exhibited in the mouse (mESCs), which can give rise to every cell type when combined with blastocysts. Due to obvious barriers precluding the use of human embryos in similar cell mixing experiments with hESCs, human/non-human chimeras may need to be generated for this purpose. Our results show that hESCs can engraft into mouse blastocysts, where they proliferate and differentiate in vitro and persist in mouse/human embryonic chimeras that implant and develop in the uterus of pseudopregnant foster mice. Embryonic chimeras generated in this way offer the opportunity to study the behavior of specialized human cell types in a non-human animal model. Our data demonstrate the feasibility of this approach, using mouse embryos as a surrogate for hESC differentiation.

Animals↗
PubMed2006

GDF3 at the crossroads of TGF-beta signaling.

Embryonic stem (ES) cells present an excellent system for addressing the relevance of our current knowledge about how cell fate is determined and how cells integrate multiple signals into a single outcome as a function of time. Many of the factors that mediate these phenomena have been discovered through classical embryological experiments and are organized into several major signal transduction pathways including TGF-beta/BMP, Jak-STAT, Hedgehog, Wnt, Notch and FGF/MAPK. This review will summarize the current understanding of TGF-beta signaling in ES and focus on early embryological roles of the TGF-beta member, GDF-3. GDF-3 is associated with the undifferentiated state of ES cells and two recent and contradictory reports examined the function and mechanism of GDF-3 in the context of both stem cells and early embryonic differentiations. While Levine and Brivanlou found that GDF-3 inhibits its own subfamily members (the BMPs), Chen and colleagues found that GDF-3 acts as a nodal-like TGF-beta ligand. These combined findings raise the intriguing possibility that GDF-3 acts as a bi-functional protein, to regulate the balance between the two modes of TGF-beta signaling.

Animals↗
PubMed2006

The RNA-binding protein, Vg1RBP, is required for pancreatic fate specification.

Signaling mechanisms underlying the induction of the pre-pancreatic tissue within the endoderm remain poorly understood. Through an expression cloning strategy, we have identified a previously uncharacterized pancreatic factor that we named Shirin. Interestingly, the non-coding RNA regulatory sequence (3 UTR) of Shirin is sufficient to induce insulin expression in Xenopus embryos. Biochemical studies demonstrate that this RNA sequence is able to bind directly to a trans-acting factor, Vg1RBP, which was previously shown to be involved in the localization of endodermal determinant factors. Loss-of-function analysis indicates that Vg1RBP is required for establishment of pancreatic fate within the endoderm, suggesting a synergism between Vg1RBP and Shirin in the embryo. This study argues for a central role of post-transcriptional mechanisms in establishing pancreatic fate, where a 3 UTR may recruit factors necessary for pancreatic development, and highlights an unknown embryological activity of Vg1RBP.

3' Untranslated Regions↗
PubMed2006

An obligatory caravanserai stop on the silk road to neural induction: inhibition of BMP/GDF signaling.

Work in Xenopus laevis produced the first molecular explanation for neural specification, the default model, where inactivation of the BMP pathway in ectodermal cells changes fates from epidermal to neural. This review covers the present status of our understanding of neural specification, with emphasis on Xenopus, but including relevant facts in other model systems. While recent experiments have increased the complexity of the molecular picture, they have also provided additional support for the default model and the central position of the BMP pathway. We conclude that synergy between accumulated knowledge and technical progress will maintain Xenopus at the forefront of research in neural development.

Animals↗
PubMed2006

Manipulation of self-renewal in human embryonic stem cells through a novel pharmacological GSK-3 inhibitor.

This chapter introduces a new method of maintaining human embryonic stem cells (hESCs) in the undifferentiated state through treatment with a GSK-3 inhibitor, BIO, under a feeder-free condition. Additionally, methods are introduced that determine multidifferentiation potential of hESCs by differentiating into a specific type or random heterogenous cell populations in vitro. These approaches will become a fundamental platform to identify molecular networks regulating cell fate determination in hESCs and mouse embryonic stem cells.

Animals↗
PubMed2006

Microarray approach to identify the signaling network responsible for self-renewal of human embryonic stem cells.

This chapter introduces the representative method to culture human embryonic stem cells (hESCs) under the feeder and feeder-free conditions, the former of which is used to maintain or expand undifferentiated hESCs, and the latter can be used for preparation of pure hESCs RNA samples, or for screening factors influential on self-renewal of hESCs. Also described in this chapter: a protocol and tips for conducting gene chip analysis focusing on widely used Affymetrix microarrays. These techniques will provide an unprecedented scale of biological information to illuminate a key to decipher complex signaling networks controlling pluripotency.

Animals↗
PubMed2005

GDF3, a BMP inhibitor, regulates cell fate in stem cells and early embryos.

The TGFbeta superfamily of ligands plays key functions in development and disease. In both human and mouse embryonic stem cells, a member of this family, GDF3, is specifically expressed in the pluripotent state. We show that GDF3 is an inhibitor of its own subfamily, blocks classic BMP signaling in multiple contexts, interacts with BMP proteins and is expressed specifically in the node during gastrulation in a pattern consistent with BMP inhibition. Furthermore, we use gain- and reduction-of-function to show that in a species-specific manner, GDF3 regulates both of the two major characteristics of embryonic stem cells: the ability to maintain the undifferentiated state and the ability to differentiate into the full spectrum of cell types.

Animals↗
PubMed2005

XenDB: full length cDNA prediction and cross species mapping in Xenopus laevis.

BACKGROUND: Research using the model system Xenopus laevis has provided critical insights into the mechanisms of early vertebrate development and cell biology. Large scale sequencing efforts have provided an increasingly important resource for researchers. To provide full advantage of the available sequence, we have analyzed 350,468 Xenopus laevis Expressed Sequence Tags (ESTs) both to identify full length protein encoding sequences and to develop a unique database system to support comparative approaches between X. laevis and other model systems. DESCRIPTION: Using a suffix array based clustering approach, we have identified 25,971 clusters and 40,877 singleton sequences. Generation of a consensus sequence for each cluster resulted in 31,353 tentative contig and 4,801 singleton sequences. Using both BLASTX and FASTY comparison to five model organisms and the NR protein database, more than 15,000 sequences are predicted to encode full length proteins and these have been matched to publicly available IMAGE clones when available. Each sequence has been compared to the KOG database and approximately 67% of the sequences have been assigned a putative functional category. Based on sequence homology to mouse and human, putative GO annotations have been determined. CONCLUSION: The results of the analysis have been stored in a publicly available database XenDB http://bibiserv.techfak.uni-bielefeld.de/xendb/. A unique capability of the database is the ability to batch upload cross species queries to identify potential Xenopus homologues and their associated full length clones. Examples are provided including mapping of microarray results and application of 'in silico' analysis. The ability to quickly translate the results of various species into 'Xenopus-centric' information should greatly enhance comparative embryological approaches.

Animals↗
PubMed2005

WDR5 associates with histone H3 methylated at K4 and is essential for H3 K4 methylation and vertebrate development.

Histone H3 lysine 4 (K4) methylation has been linked to the transcriptional activation in a variety of eukaryotic species. Here we show that a common component of MLL1, MLL2, and hSet1 H3 K4 methyltransferase complexes, the WD40-repeat protein WDR5, directly associates with histone H3 di- and trimethylated at K4 and with H3-K4-dimethylated nucleosomes. WDR5 is required for binding of the methyltransferase complex to the K4-dimethylated H3 tail as well as for global H3 K4 trimethylation and HOX gene activation in human cells. WDR5 is essential for vertebrate development, in that WDR5-depleted X. laevis tadpoles exhibit a variety of developmental defects and abnormal spatial Hox gene expression. Our results are the first demonstration that a WD40-repeat protein acts as a module for recognition of a specific histone modification and suggest a mechanism for reading and writing an epigenetic mark for gene activation.

Animals↗
PubMed2005

DRAGON, a bone morphogenetic protein co-receptor.

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor (TGF)beta superfamily of ligands that regulate many crucial aspects of embryonic development and organogenesis. Unlike other TGFbeta ligands, co-receptors for BMP ligands have not been described. Here we show that DRAGON, a glycosylphosphatidylinositol-anchored member of the repulsive guidance molecule family, which is expressed early in the developing nervous system, enhances BMP but not TGFbeta signaling. DRAGON binds directly to BMP2 and BMP4 but not to BMP7 or other TGFbeta ligands. The enhancing action of DRAGON on BMP signaling is also reduced by administration of Noggin, a soluble BMP antagonist, indicating that the action of DRAGON is ligand-dependent. DRAGON associates directly with BMP type I (ALK2, ALK3, and ALK6) and type II (ActRII and ActRIIB) receptors, and its signaling is reduced by dominant negative Smad1 and ALK3 or -6 receptors. In the Xenopus embryo, DRAGON both reduces the threshold of the ability of Smad1 to induce mesodermal and endodermal markers and alters neuronal and neural crest patterning. The direct interaction of DRAGON with BMP ligands and receptors indicates that it is a BMP co-receptor that potentiates BMP signaling.

Animals↗
PubMed2005

Induction of ectopic olfactory structures and bone morphogenetic protein inhibition by Rossy, a group XII secreted phospholipase A2.

The secreted phospholipases A(2) (sPLA(2)s) comprise a family of small secreted proteins with the ability to catalyze the generation of bioactive lipids through glycophospholipid hydrolysis. Recently, a large number of receptor proteins and extracellular binding partners for the sPLA(2)s have been identified, suggesting that these secreted factors might exert a subset of their broad spectrum of biological activities independently of their enzymatic activity. Here, we describe an activity for the sPLA(2) group XII (sPLA(2)-gXII) gene during Xenopus laevis early development. In the ectoderm, sPLA(2)-gXII acts as a neural inducer by blocking bone morphogenetic protein (BMP) signaling. Gain of function in embryos leads to ectopic neurogenesis and to the specification of ectopic olfactory sensory structures, including olfactory bulb and sensory epithelia. This activity is conserved in the Drosophila melanogaster, Xenopus, and mammalian orthologs and appears to be independent of the lipid hydrolytic activity. Because of its effect on olfactory neurogenesis, we have renamed this gene Rossy, in homage to the Spanish actress Rossy de Palma. We present evidence that Rossy/sPLA(2)-gXII can inhibit the transcriptional activation of BMP direct-target gene reporters in Xenopus and mouse P19 embryonic carcinoma cells through the loss of DNA-binding activity of activated Smad1/4 complexes. Collectively, these data represent the first evidence for signaling cross talk between a secreted phospholipase A(2) and the BMP/transforming growth factor beta pathways and identify Rossy/sPLA(2)-gXII as the only factor thus far described which is sufficient to induce anterior sensory neural structures during vertebrate development.

Amino Acid Sequence↗
PubMed2005

A molecular basis for human embryonic stem cell pluripotency.

Embryonic stem cells (ESCs) are able to generate a wide array of differentiated cell fates while maintaining self-renewal. Understanding the biology of these choices may be central to the use of human embryonic stem cells (HESCs), both as a model for early human development as well as a resource for cell based therapies. Efforts to dissect the molecular mechanisms that mediate stem cell identity are underway, and in this review we summarize recent progress in defining the markers and pathways involved in these decisions. We discuss recent efforts to assess the molecular signature of pluripotent HESCs and highlight work demonstrating a set of genes, including representatives from the FGF, TGFbeta, and Wnt signaling pathways, that consistently mark the undifferentiated state. In addition, we describe experiments in which signaling of HESCs is augmented by chemical probing with small molecule compounds. Using these compounds, we have demonstrated an important role for Wnt signaling in HESC pluripotency and shown a requirement for TGFbeta signaling in the maintenance of the undifferentiated state. These experiments have revealed some molecular aspects of the pluripotent state and demonstrated clear differences between mouse and human ESCs in the maintenance of this identity.

Animals↗
PubMed2004

FGF-20 and DKK1 are transcriptional targets of beta-catenin and FGF-20 is implicated in cancer and development.

beta-catenin is the major effector of the canonical Wnt signaling pathway. Mutations in components of the pathway that stabilize beta-catenin result in augmented gene transcription and play a major role in many human cancers. We employed microarrays to identify transcriptional targets of deregulated beta-catenin in a human epithelial cell line (293) engineered to produce mutant beta-catenin and in ovarian endometrioid adenocarcinomas characterized with respect to mutations affecting the Wnt/beta-catenin pathway. Two genes strongly induced in both systems-FGF20 and DKK1-were studied in detail. Elevated levels of FGF20 RNA were also observed in adenomas from mice carrying the Apc(Min)allele. Both XFGF20 and Xdkk-1 are expressed early in Xenopus embryogenesis under the control of the Wnt signaling pathway. Furthermore, FGF20 and DKK1 appear to be direct targets for beta-catenin/TCF transcriptional regulation via LEF/TCF-binding sites. Finally, by using small inhibitory RNAs specific for FGF20, we show that continued expression of FGF20 is necessary for maintenance of the anchorage-independent growth state in RK3E cells transformed by beta-catenin, implying that FGF-20 may be a critical element in oncogenesis induced by the Wnt signaling pathway.

Adenocarcinoma↗
PubMed2004

Lefty blocks a subset of TGFbeta signals by antagonizing EGF-CFC coreceptors.

Members of the EGF-CFC family play essential roles in embryonic development and have been implicated in tumorigenesis. The TGFbeta signals Nodal and Vg1/GDF1, but not Activin, require EGF-CFC coreceptors to activate Activin receptors. We report that the TGFbeta signaling antagonist Lefty also acts through an EGF-CFC-dependent mechanism. Lefty inhibits Nodal and Vg1 signaling, but not Activin signaling. Lefty genetically interacts with EGF-CFC proteins and competes with Nodal for binding to these coreceptors. Chimeras between Activin and Nodal or Vg1 identify a 14 amino acid region that confers independence from EGF-CFC coreceptors and resistance to Lefty. These results indicate that coreceptors are targets for both TGFbeta agonists and antagonists and suggest that subtle sequence variations in TGFbeta signals result in greater ligand diversity.

Animals↗