Dynamics of TGF-β/Smad signaling

The physiological responses to TGF-β stimulation are diverse and vary amongst different cell types and environmental conditions. Even though the principal molecular components of the canonical and the non-canonical TGF-β signaling pathways have been largely identified, the mechanism that underlies the well-established context dependent physiological responses remains a mystery. Understanding how the components of TGF-β signaling function as a system and how this system functions in the context of the global cellular regulatory network requires a more quantitative and systematic approach. Here, we review the recent progress in understanding TGF-β biology using integration of mathematical modeling and quantitative experimental analysis. These studies reveal many interesting dynamics of TGF-β signaling and how cells quantitatively decode variable doses of TGF-β stimulation.     

Discovery of novel small molecule activators of β-catenin signaling

Wnt/β-catenin signaling plays a major role in embryonic development and adult stem cell maintenance. Reduced activation of the Wnt/β-catenin pathway underlies neurodegenerative disorders and aberrations in bone formation. Screening of a small molecule compound library with a β-galactosidase fragment complementation assay measuring β-catenin nuclear entry revealed bona fide activators of β-catenin signaling. The compounds stabilized cytoplasmic β-catenin and activated β-catenin-dependent reporter gene activity. Although the mechanism through which the compounds activate β-catenin signaling has yet to be determined, several key regulators of Wnt/β-catenin signaling, including glycogen synthase kinase 3 and Frizzled receptors, were excluded as the molecular target. The compounds displayed remarkable selectivity, as they only induced β-catenin signaling in a human osteosarcoma U2OS cell line and not in a variety of other cell lines examined. Our data indicate that differences in cellular Wnt/β-catenin signaling machinery can be exploited to identify cell type-specific activators of Wnt/β-catenin signaling.     

Characterization of the CaMKKβ-AMPK signaling complex

The AMP-activated protein kinase (AMPK) is a critical regulator of energy homeostasis, and is a potential target for treatment of metabolic diseases as well as cancer. AMPK can be phosphorylated and activated by the tumor suppressor LKB1 or the Ca²⁺/CaM-dependent protein kinase kinase β (CaMKKβ). We previously identified a physical complex between CaMKKβ and AMPK (Anderson, K. A., Ribar, T. J., Lin, F., Noeldner, P. K., Green, M. F., Muehlbauer, M. J., Witters, L. A., Kemp, B. E., and Means, A. R. (2008) Cell Metabolism 7, 377–388). Here we expand our analysis of the CaMKKβ–AMPK signaling complex and show that whereas CaMKKβ can form a complex with and activate AMPK, CaMKKα cannot. In addition, we show that CaMKKβ and AMPK associate through their kinase domains, and CaMKKβ must be in an active conformation in order to bind AMPK but not to associate with an alternative substrate, Ca²⁺/Calmodulin-dependent protein kinase IV (CaMKIV). Our results demonstrate that CaMKKβ and AMPK form a unique signaling complex. This raises the possibility that the CaMKKβ–AMPK complex can be specifically targeted by small molecule drugs to treat disease.     

Matrix-dependent perturbation of TGFβ signaling and disease

Transforming growth factor beta (TGFβ) is a multipotent cytokine that is sequestered in the extracellular matrix (ECM) through interactions with a number of ECM proteins. The ECM serves to concentrate latent TGFβ at sites of intended function, to influence the bioavailability and/or function of TGFβ activators, and perhaps to regulate the intrinsic performance of cell surface effectors of TGFβ signal propagation. The downstream consequences of TGFβ signaling cascades in turn provide feedback modulation of the ECM. This review covers recent examples of how genetic mutations in constituents of the ECM or TGFβ signaling cascade result in altered ECM homeostasis, cellular performance and ultimately disease, with an emphasis on emerging therapeutic strategies that seek to capitalize on this refined mechanistic understanding.     

Potentiation of μ-opioid receptor-mediated signaling by ketamine

Ketamine, a clinically relevant drug, has been shown to enhance opioid-induced analgesia and prevent hyperalgesia. However, the molecular mechanisms involved are not clearly understood. As previous studies found that activation of opioid receptors leads to the phosphorylation of mitogen-activated protein kinases, we investigated whether ketamine could modulate μ-opioid receptor (μOR)-mediated ERK1/2 phosphorylation. We find that acute treatment with ketamine enhances (～2- to 3-fold) the levels of opioid-induced ERK1/2 phosphorylation in recombinant as well as cells endogenously expressing μOR. Interestingly, we find that in the absence of ketamine ERK1/2 signaling is desensitized 10 min after opioid exposure whereas in its presence significant levels (～3-fold over basal) are detected. In addition, ketamine increases the rate of resensitization of opioid-mediated ERK1/2 signaling (15 min in its presence vs. 30 min in its absence). These results suggest that ketamine increases the effectiveness of opiate-induced signaling by affecting multiple mechanisms. In addition, these effects are observed in heterologous cells expressing μOR suggesting a non-NMDA receptor-mediated action of ketamine. Together this could, in part, account for the observed effects of ketamine on the enhancement of the analgesic effects of opiates as well as in the duration of opiate-induced analgesia.     

Negative regulation of TGF-β signaling in development

The TGF-β superfamily members have important roles in controlling patterning and tissue formation in both invertebrates and vertebrates. Two types of signal transducers, receptors and Smads, mediate the signaling to regulate expression of their target genes. Despite of the relatively simple signal transduction pathway, many modulators have been found to contribute to a tight regulation of this pathway in a variety of mechanisms. This article reviews the negative regulation of TGF-β signaling with focus on its roles in vertebrate development.     

Molecular evolution of the NF-κB signaling system

The mechanisms of innate immunity in vertebrates show certain overall resemblances to immune mechanisms of insects. Two hypotheses have been proposed to explain these resemblances. (1) According to the evolutionary continuity hypothesis, innate immune mechanisms evolved in the common ancestor of vertebrates and insects and have been conserved since that time. (2) In the independent-evolution hypothesis, the mechanisms of innate immunity in vertebrates evolved independently from invertebrate immune mechanisms. Phylogenetic analysis of five gene families (Pelle, Rel, IkappaB, Toll, and TRAF) whose members are involved in NF-kappaB signaling in vertebrates and insects were used to decide between these hypotheses. The phylogenies of the Rel and TRAF families strongly supported independent evolution of immune functions in vertebrates and invertebrates, and, except for a possible case in the Pelle family, orthologous molecules having immune functions in both vertebrates and invertebrates were not found. The results suggest that NF-kappaB represents an ancient, generalized signaling system that has been co-opted for immune system roles independently in vertebrate and insect lineages.     

Expanding role of ubiquitination in NF-κB signaling

Best known for its role in targeting protein degradation by the proteasome, ubiquitin modification has also emerged as an important mechanism that regulates cell signaling through proteasome-independent mechanisms. The role of ubiquitin as a versatile signaling tag is characteristically illustrated in the NF-κB pathways, which regulate a variety of physiological and pathological processes in response to diverse stimuli. Here, we review the role of ubiquitination in different steps of the NF-κB signaling cascades, focusing on recent advances in understanding the mechanisms of protein kinase activation by polyubiquitin chains in different pathways that converge on NF-κB.     

Structure-based reassessment of the caveolin signaling model: do caveolae regulate signaling through caveolin-protein interactions?

Caveolin proteins drive formation of caveolae, specialized cell-surface microdomains that influence cell signaling. Signaling proteins are proposed to use conserved caveolin-binding motifs (CBMs) to associate with caveolae via the caveolin scaffolding domain (CSD). However, structural and bioinformatic analyses argue against such direct physical interactions: in the majority of signaling proteins, the CBM is buried and inaccessible. Putative CBMs do not form a common structure for caveolin recognition, are not enriched among caveolin-binding proteins, and are even more common in yeast, which lack caveolae. We propose that CBM/CSD-dependent interactions are unlikely to mediate caveolar signaling, and the basis for signaling effects should therefore be reassessed.     

New mechanisms of pulmonary arterial hypertension: role of Ca²⁺ signaling

Pulmonary arterial hypertension (PAH) is a severe and progressive disease that usually culminates in right heart failure and death if left untreated. Although there have been substantial improvements in our understanding and significant advances in the management of this disease, there is a grim prognosis for patients in the advanced stages of PAH. A major cause of PAH is increased pulmonary vascular resistance, which results from sustained vasoconstriction, excessive pulmonary vascular remodeling, in situ thrombosis, and increased pulmonary vascular stiffness. In addition to other signal transduction pathways, Ca(2+) signaling in pulmonary artery smooth muscle cells (PASMCs) plays a central role in the development and progression of PAH because of its involvement in both vasoconstriction, through its pivotal effect of PASMC contraction, and vascular remodeling, through its stimulatory effect on PASMC proliferation. Altered expression, function, and regulation of ion channels and transporters in PASMCs contribute to an increased cytosolic Ca(2+) concentration and enhanced Ca(2+) signaling in patients with PAH. This review will focus on the potential pathogenic role of Ca(2+) mobilization, regulation, and signaling in the development and progression of PAH.     

How costly is the honest signaling of need?

ESS models of biological signaling have shown that costly signals can provide honest information. In the context of parent-offspring conflict over the allocation of resources by parents to their young, the theory explains costly offspring solicitation behavior as an accurate signal of offspring need to parents who cannot assess offspring condition directly. In this paper, we provide a simple but general characterization of the honest signaling of need in models of parent-offspring conflict: the offspring's signaling cost is proportional to the parent's fitness loss from satisfying the offspring's resource requirement. The factor of proportionality is given by a measure of the extent of parent-offspring conflict that depends only on coefficients of relatedness. These results hold for interbrood conflict with uniparental investment even if the relationship between offspring condition and resource requirement is not monotonic, and extend to cases of biparental care, uncertainty concerning the parent's condition, and intra-brood conflict. Copyright 1999 Academic Press.