The Xenopus Lefty (xlefty) Prodomain Negatively Regulates Xlefty Activity And Is Necessary For Proper Xlefty Secretion

In vertebrates, the TGF #914; superfamily of secreted peptides are stringently regulated since they are responsible for multiple cell processes and behaviors that give rise to the proper patterning of embryonic tissues. Examples of members of the TGF #914; superfamily are Nodal and Lefty. Dysregulation of these proteins can lead to many diseases and developmental syndromes in humans. Lefty functions by antagonizing Nodal an essential organizer signal that patterns dorsal mesoderm and the embryonic axes. Loss of Lefty expression results in excess Nodal signaling which has been shown to cause several perturbations including metastatic cancer. Although studies have clearly shown that Lefty antagonizes Nodal signaling, there is a lack of understanding of the regulatory mechanism of Lefty itself. Previous studies have shown that proteolytic cleavage of the prodomain (PD) from mature Lefty (Mat-Lefty) is necessary for Lefty activity. Here we present studies carried out using the Xenopus laevis embryo that demonstrate that PD over-expression causes exogastrulation, a phenotype also resulting from a loss of Xenopus Lefty (Xlefty) function. Furthermore, when the PD and Xlefty are co-expressed, the effects of Xlefty over-expression are rescued. Our biochemical studies also showed that the PD interacts with Mat-Xlefty but a PD mutated (PDmut) molecule does not. The sites mutated in the PDmut are evolutionarily-conserved residues that mediate the interaction between the prodomain and the mature ligand in other TGF #914; proteins making our results consistent with previously observed behavior of TGF #914; prodomains but new to Lefty. Our study also shows that the PDmut is unable to rescue the effects of a Xlefty over-expression phenotype unlike the un-mutated PD. In addition we show here that mutations of the PD affect the secretion of these Xlefty-mutated derivatives. Taken together, these results suggest that the PD negatively regulates Xlefty activity by interacting with Mat-Xlefty and cleavage of the PD releases regulation allowing proper secretion and function. This new insight into the regulatory role of the Xlefty PD provides potential therapeutic value to address dysfunctional Nodal signaling. Furthermore, our secretion studies of the PD and Xlefty revealed that Xlefty is secreted but the PD is not. The Xlefty-mutated derivatives, including a PD Less, Xlefty, Xlefty cleavage mutants and the PDmut, resulted in no secreted products. These secretion results open a Pandora\u27s box and further studies are warranted to elucidate the mechanism of PD regulation of Xlefty and whether it occurs in an intracellular or extracellular context

Lefty Blocks a Subset of TGFβ 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 TGFβ signals Nodal and Vg1/GDF1, but not Activin, require EGF-CFC coreceptors to activate Activin receptors. We report that the TGFβ 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 TGFβ agonists and antagonists and suggest that subtle sequence variations in TGFβ signals result in greater ligand diversity

Role of Lefty in the anti tumor activity of human adult liver stem cells

Recent studies demonstrated that factors derived from embryonic stem cells inhibit the tumorigenicity of a variety of cancer cell lines. Embryonic stem cell-secreted Lefty, an inhibitor of Nodal-signalling pathway, was implicated in reprogramming cancer cells. Whether adult stem cells exhibited similar properties has not been explored. The aim of the present study was to investigate whether the conditioned medium (CM) derived from adult stem cells influence in vitro and in vivo tumor growth by a Nodal-dependent pathway. In particular we compared the anti-tumor effect of CM from human liver stem cells (HLSC) with that of bone marrow-derived mesenchymal stem cells (MSC). We found that HLSC-CM inhibited the in vitro growth and promoted apoptosis in HepG2 cells that expressed a deregulated Nodal pathway. The effect of HLSC-CM was related to the presence of Lefty A in the CM of HLSC. Silencing Lefty A in HLSC or Lefty A blockade with a blocking peptide abrogated the anti-proliferative and pro-apoptotic effect of HLSC-CM. Moreover, the administration of human recombinant Lefty A protein mimicked the effect of HLSC-CM indicating that Nodal pathway is critical for the growth of HepG2. At variance of HLSC, bone marrow-derived MSC did not express and release Lefty A and the MSC-CM did not exhibited an anti-tumor activity in vitro, but rather stimulated proliferation of HepG2. In addition, the intra-tumor administration of HLSC-CM was able to inhibit the in vivo growth of HepG2 hepatoma cells implanted subcutaneously in SCID mice. At variance, HLSC-CM derived from Lefty A silenced HLSC was unable to inhibit tumor growth. In conclusion, the results of present study suggest that Lefty A may account for the tumor suppressive activity of HLSC as a result of an inhibition of the Nodal-signalling pathway by a mechanism similar to that described for embryonic stem cells

Do Lefty and Righty Matter More Than Lefty Alone?

Derek Parfit argues that fission is prudentially better for you than ordinary death. But is having more fission products with good lives prudentially better for you than having just one? In this paper, we argue that it is. We argue that, if your brain is split and the halves are transplanted into two recipients (who both have good lives), then it is prudentially better for you if both transplants succeed than if only one of them does (other things being equal). This upshot rules out, among other things, that the prudential value of standing in the relation that matters in survival to multiple people is equal to their average well-being

Interplay between Lefty and Nodal signaling is essential for the organizer and axial formation in amphioxus embryos

Abstract(#br)The organizer is an essential signaling center required for axial formation during vertebrate embryonic development. In the basal chordate amphioxus, the dorsal blastopore lip of the gastrula has been proposed to be homologous to the vertebrate organizer. Lefty is one of the first genes to be expressed in the organizer. The present results show that Lefty overexpression abolishes the organizer; the embryos were severely ventralized and posteriorized, and failed to develop anterior and dorsal structures. In Lefty knockouts the organizer is enlarged, and anterior and dorsal structures are expanded. Different from Lefty morphants in vertebrates, amphioxus Lefty mutants also exhibited left-right defects. Inhibition of Nodal with SB505124 partially rescued the effects of Lefty loss-of-function on morphology. In addition, while SB505124 treatment blocked Lefty expression in the cleavage stages of amphioxus embryos, activation of Nodal signaling with Activin protein induced ectopic Lefty expression at these stages. These results show that the interplay between Lefty and Nodal signaling plays an essential role in the specification of the amphioxus organizer and axes

October 25, 1974

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Temporal Control of the TGF-β Signaling Network by Mouse ESC MicroRNA Targets of Different Affinities.

Although microRNAs (miRNAs) function in the control of embryonic stem cell (ESC) pluripotency, a systems-level understanding is still being developed. Through the analysis of progressive Argonaute (Ago)-miRNA depletion and rescue, including stable Ago knockout mouse ESCs, we uncover transforming growth factor beta (TGF-β) pathway activation as a direct and early response to ESC miRNA reduction. Mechanistically, we link the derepression of weaker miRNA targets, including TGF-β receptor 1 (Tgfbr1), to the sensitive TGF-β pathway activation. In contrast, stronger miRNA targets impart a more robust repression, which dampens concurrent transcriptional activation. We verify such dampened induction for TGF-β antagonist Lefty. We find that TGF-β pathway activation contributes to the G1 cell-cycle accumulation of miRNA-deficient ESCs. We propose that miRNA target affinity is a determinant of the temporal response to miRNA changes, which enables the coordination of gene network responses

Baker's First-person Perspectives: They Are Not What They Seem

Lynne Baker's concept of a first-person perspective is not as clear and straightforward as it might seem at first glance. There is a discrepancy between her argumentation that we have first-person perspectives and some characteristics she takes first-person perspectives to have, namely, that the instances of this capacity necessarily persist through time and are indivisible and unduplicable. Moreover, these characteristics cause serious problems concerning personal identity