Receptor tyrosine kinases: from biology to pathology

Receptor tyrosine kinases (RTKs) are transmembrane proteins involved in the control of fundamental cellular processes in metazoans. RTKs possess a general structure that includes an extracellular domain, a transmembrane domain and a highly conserved tyrosine kinase domain. RTKs are classified according to their variable extracellular ligand-binding domain. Studies of human RTK members have yielded a wealth of information elucidating their importance. Improper functioning of these enzymes due to mutations, mainly in the kinase domain, is often manifested in various human diseases and is known to be involved in several types of cancer. Here we summarize most of human RTKs, their cognate ligands, as well as related diseases and discuss the eventual use of certain RTKs as new therapeutic targets.     

Receptor tyrosine kinases: from biology to pathology

Receptor tyrosine kinases (RTKs) are transmembrane proteins involved in the control of fundamental cellular processes in metazoans. RTKs possess a general structure that includes an extracellular domain, a transmembrane domain and a highly conserved tyrosine kinase domain. RTKs are classified according to their variable extracellular ligand-binding domain. Studies of human RTK members have yielded a wealth of information elucidating their importance. Improper functioning of these enzymes due to mutations, mainly in the kinase domain, is often manifested in various human diseases and is known to be involved in several types of cancer. Here we summarize most of human RTKs, their cognate ligands, as well as related diseases and discuss the eventual use of certain RTKs as new therapeutic targets.     

Nuclear presence of adhesion-/growth-regulatory galectins in normal/malignant cells of squamous epithelial origin

Cellular activities in the regulation of growth or adhesion/migration involve protein (lectin)–carbohydrate recognition at the cell surface. Members of the galectin family of endogenous lectins additionally bind distinct intracellular ligands. These interactions with protein targets explain the relevance of their nuclear and cytoplasmic presence. Expression profiling for galectins and accessible binding sites is a histochemical approach to link localization with cellular growth properties. Non-cross-reactive antibodies for the homodimeric (proto-type) galectins-1, -2 and -7 and the chimera-type galectin-3 (Gal-3) as well as the biotinylated lectins were tested. This analysis was performed with the FaDu squamous carcinoma cell line and long-term cultured human and porcine epidermal cells as models for malignant and normal cells of squamous cell epithelial origin. A set of antibodies was added for phenotypic cell characterization. Strong nuclear and cytoplasmic signals of galectins and the differential reactivity of labeled galectins support the notion of their individual properties. The length of the period of culture was effective in modulating marker expression. Cytochemical expression profiling is a prerequisite for the selection of distinct proteins for targeted modulation of gene expression as a step toward functional analysis.     

Using phage display to select antibodies recognizing post-translational modifications independently of sequence context

Many cellular activities are controlled by post-translational modifications, the study of which is hampered by the lack of specific reagents due in large part to their ubiquitous and non-immunogenic nature. Although antibodies against specifically modified sequences are relatively easy to obtain, it is extremely difficult to derive reagents recognizing post-translational modifications independently of the sequence context surrounding the modification. In this study, we examined the possibility of selecting such antibodies from large phage antibody libraries using sulfotyrosine as a test case. Sulfotyrosine is a post-translational modification important in many extracellular protein-protein interactions, including human immunodeficiency virus infection. After screening almost 8000 selected clones, we were able to isolate a single specific single chain Fv using two different selection strategies, one of which included elution with tyrosine sulfate. This antibody was able to recognize sulfotyrosine independently of its sequence context in test peptides and a number of different natural proteins. Antibody reactivity was lost by antigen treatment with sulfatase or preincubation with soluble tyrosine sulfate, indicating its specificity. The isolation of this antibody signals the potential of phage antibody libraries in the derivation of reagents specific for post-translational modifications, although the extensive screening required indicates that such antibodies are extremely rare.     

Galectins as modulators of cell adhesion

The galectins are a family of carbohydrate-binding proteins that are distributed widely in metazoan organisms. Each galectin exhibits a specific pattern of expression in various cells and tissues, and expression is often closely regulated during development. Although these proteins are found mainly in the cell cytoplasm, some are secreted from cells and interact with appropriately glycosylated proteins at the cell surface or within the extracellular matrix. These receptors include cell-adhesion molecules such as integrins, and matrix glycoproteins such as laminin and fibronectin isoforms. Recent studies have increased understanding of the roles of the galectins in regulating cell-cell and cell-matrix adhesion. These interactions are critically involved in modulation of normal cellular motility and polarity and during tissue formation, and loss of adhesive function is implicated in several disease states including tumour progression, inflammation and cystic development in branching epithelia such as kidney tubules. This review discusses recent progress in defining the specificities and mechanisms of action of secreted galectins as multifunctional cell regulators.     

Reactive nitrogen species in cellular signaling

The transduction of cellular signals occurs through the modification of target molecules. Most of these modifications are transitory, thus the signal transduction pathways can be tightly regulated. Reactive nitrogen species are a group of compounds with different properties and reactivity. Some reactive nitrogen species are highly reactive and their interaction with macromolecules can lead to permanent modifications, which suggested they were lacking the specificity needed to participate in cell signaling events. However, the perception of reactive nitrogen species as oxidizers of macromolecules leading to general oxidative damage has recently evolved. The concept of redox signaling is now well established for a number of reactive oxygen and nitrogen species. In this context, the post-translational modifications introduced by reactive nitrogen species can be very specific and are active participants in signal transduction pathways. This review addresses the role of these oxidative modifications in the regulation of cell signaling events.     

Ah,sweet mystery of death! Galectins and control of cell fate

Control of cell death is critical in eukaryotic development, immune system homeostasis, and control of tumorigenesis. The galectin family of lectins is implicated in all of these processes. Other families of molecules function as death receptors or death effectors, but galectins are uniquely capable of acting both extracellularly and intracellularly to control cell death. Extracellularly, galectins cross-link glycan ligands to transduce signals that lead directly to death or that influence other signals regulating cell fate. Intracellular expression of galectins can modulate other signals controlling cell viability. Individual galectins can act on multiple cell types, and multiple galectins can act on the same cell. Understanding how galectins regulate cell viability and function will broaden our knowledge of the roles of galectins in basic biological processes and facilitate development of therapeutic applications for galectins in autoimmunity, transplant-related disease, and cancer.     

Caenorhabditis elegans galectins LEC-6 and LEC-10 interact with similar glycoconjugates in the intestine

Galectins are a family of metazoan proteins that show binding to various β-galactoside-containing glycans. Because of a lack of proper tools, the interaction of galectins with their specific glycan ligands in the cells and tissues are largely unknown. We have investigated the localization of galectin ligands in Caenorhabditis elegans using a novel technology that relies on the high binding specificity between galectins and their endogenous ligands. Fluorescently labeled recombinant galectin fusions are found to bind to ligands located in diverse tissues including the intestine, pharynx, and the rectal valve. Consistent with their role as galactoside-binding proteins, the interaction with their ligands is inhibited by galactose or lactose. Two of the galectins, LEC-6 and LEC-10, recognize ligands that co-localize along the intestinal lumen. The ligands for LEC-6 and LEC-10 are absent in three glycosylation mutants bre-1, fut-8, and galt-1, which have been shown to be required to synthesize the Gal-β1,4-Fuc modifications of the core N-glycans unique to C. elegans and several other invertebrates. Both galectins pull down the same set of glycoproteins in a manner dependent on the presence of these carbohydrate modifications. Endogenous LEC-6 and LEC-10 are expressed in the intestinal cells, but they are localized to different subcellular compartments that do not appear to overlap with each other or with the location of their glycan targets. An altered subcellular distribution of these ligands is found in mutants lacking both galectins. These results suggest a model where LEC-6 and LEC-10 interact with glycoproteins through specific glycans to regulate their cellular fate.     

Receptor tyrosine kinases: mechanisms of activation and signaling

Receptor tyrosine kinases (RTKs) are essential components of signal transduction pathways that mediate cell-to-cell communication. These single-pass transmembrane receptors, which bind polypeptide ligands - mainly growth factors - play key roles in processes such as cellular growth, differentiation, metabolism and motility. Recent progress has been achieved towards an understanding of the precise (and varied) mechanisms by which RTKs are activated by ligand binding and by which signals are propagated from the activated receptors to downstream targets in the cell.     

Phosphorylation, acetylation and ubiquitination: the molecular basis of RUNX regulation

The RUNX family members play pivotal roles in normal development and neoplasia. RUNX1 and RUNX2 are essential for hematopoiesis and osteogenesis, respectively, while RUNX3 is involved in neurogenesis, thymopoiesis and functions as a tumor suppressor. Inappropriate levels of RUNX activity are associated with leukemia, autoimmune disease, cleidocranial dysplasia, craniosynostosis and various solid tumors. Therefore, RUNX activity must be tightly regulated to prevent tumorigenesis and maintain normal cell differentiation. Recent work indicates that RUNX activity is controlled by various extracellular signaling pathways, and that phosphorylation, acetylation and ubiquitination are important post-translational modifications of RUNX that affect its stability and activity. Defining the precise roles, these modifications that play in the regulation of RUNX function may reveal not only how the RUNX proteins are regulated but also how they are assembled into other regulatory machineries.     

Regulation of protease-activated receptor signaling by post-translational modifications

Protease-activated receptors (PARs) are a unique family of G-protein-coupled receptors (GPCRs) that are irreversibly activated following proteolytic cleavage of their extracellular N-terminus. PARs play critical functions in hemostasis, thrombosis, inflammation, embryonic development, and cancer progression. Because of the irreversible proteolytic nature of PAR activation, signaling by the receptors is tightly regulated. Three distinct processes including desensitization, internalization, and lysosomal degradation, regulate the temporal and spatial aspects of activated PAR signaling. Post-translational modifications play a critical role in regulating each of these processes and here we review the nature of PAR post-translational modifications and their importance in signal regulation. The PARs are activated by numerous proteases, and some can elicit distinct cellular responses, how this biased agonism is determined is unknown. Further study of the function of post-translational modifications of the PARs will lead to a greater understanding of the physiological regulation of baised agonism and how PAR signaling is precisely controlled in different cellular contexts.