Intermediate filament proteins participate in signal transduction

How timely transport of chemical signals between the distal end of long axonal processes and the cell bodies of neurons occurs is an interesting and unresolved issue. Recently, Perlson et al. presented evidence that cleavage products of newly synthesized vimentin, an intermediate filament (IF) protein, interact with mitogen-activated protein (MAP) kinases at sites of axon injury. These IF fragments appear to be required for the transport of these kinases to the cell body along microtubule tracks. The truncated vimentin is instrumental in signal propagation as it provides a scaffold that brings together activated MAP kinases (such as Erk 1 and Erk2), as well as importin beta and cytoplasmic dynein. The authors propose that this all-in-one transport complex has the extraordinary ability to travel towards the cell body and enter the nucleus where the kinases activate and influence gene expression so that a neuron can generate a timely response to injury.     

Evidence that carnosine and anserine may participate in Wilson's disease

Carnosine, anserine and copper(II) ion all bind to specific sites on bovine serum albumin, and, in addition, both dipeptides chelate copper(II) ion in the absence of serum albumin. Thus a solution of dipeptide, copper(II) ion and serum albumin exhibits several complexes that arise from the competing binding reactions. Since a change in this complex equilibrium might occur in Wilson's disease, we have investigated the reactions between the various complexes with NMR and ESR spectroscopy. Serum albumin simultaneously binds the copper(II) ion and carnosine to separate sites rather than forming a mixed chelate, but carnosine still is capable of competing with serum albumin for subsaturating amounts of copper.     

A pertussis/choleratoxin-sensitive N protein may mediate chemoattractant receptor signal transduction

Chemoattractant receptors on phagocytic leukocytes utilize a guanine nucleotide regulatory (N) protein to activate phospholipase C and subsequent biological responses. Since pertussis toxin inhibits activation of leukocytes by chemoattractants and ribosylates a ca. 40 kD protein in these cells it had generally been assumed that chemoattractant receptors are coupled to Ni. We now report that human polymorphonuclear leukocytes (PMNs), monocytes, and the myeloid HL-60 and U937 cell lines, but not erythrocytes or bovine brain contain a ca. 40 kD protein which is a substrate for ADP ribosylation by choleratoxin (CT). This N protein, termed Nc for chemotaxis-related N protein, comigrates with the ca. 40 kD PT substrate during one-dimensional gel electrophoresis. In vivo treatment of PMNs with PT or CT reduced high affinity binding of chemoattractants to membrane preparations from the cells, implying that chemoattractant receptors are coupled to an N protein which is a substrate for both PT and CT. We suggest that Nc rather than Ni couples chemoattractant receptors to phospholipase C.     

Phytohormones participate in an S6 kinase signal transduction pathway in Arabidopsis

Addition of fresh medium to stationary cells of Arabidopsis suspension culture induces increased phosphorylation of the S6 ribosomal protein and activation of its cognate kinase, AtS6k. Analysis of the activation response revealed that medium constituents required for S6 kinase activation were the phytohormones 1-naphthylacetic acid (auxin) and kinetin. Pretreatment of cells with anti-auxin or PI3-kinase drugs inhibited this response. Consistent with these findings, LY294002, a PI3-kinase inhibitor, efficiently suppressed phytohormone-induced S6 phosphorylation and translational up-regulation of ribosomal protein S6 and S18A mRNAs without affecting global translation. These data indicate that (1) activation of AtS6k is regulated by phytohormones, at least in part, via a lipid kinase-dependent pathway, that (2) the translational regulation of ribosomal proteins appears to be conserved throughout the plant and animal kingdom, and that (3) these events are hallmarks of a growth-related signal transduction pathway novel in plants.     

The sodium pump and cardiotonic steroids-induced signal transduction protein kinases and calcium-signaling microdomain in regulation of transporter trafficking

The Na/K-ATPase was discovered as an energy transducing ion pump. A major difference between the Na/K-ATPase and other P-type ATPases is its ability to bind a group of chemicals called cardiotonic steroids (CTS). The plant-derived CTS such as digoxin are valuable drugs for the management of cardiac diseases, whereas ouabain and marinobufagenin (MBG) have been identified as a new class of endogenous hormones. Recent studies have demonstrated that the endogenous CTS are important regulators of renal Na⁺ excretion and blood pressure. The Na/K-ATPase is not only an ion pump, but also an important receptor that can transduce the ligand-like effect of CTS on intracellular protein kinases and Ca²⁺ signaling. Significantly, these CTS-provoked signaling events are capable of reducing the surface expression of apical NHE3 (Na/H exchanger isoform 3) and basolateral Na/K-ATPase in renal proximal tubular cells. These findings suggest that endogenous CTS may play an important role in regulation of tubular Na⁺ excretion under physiological conditions; conversely, a defect at either the receptor level (Na/K-ATPase) or receptor–effector coupling would reduce the ability of renal proximal tubular cells to excrete Na⁺, thus culminating/resulting in salt-sensitive hypertension.     

G protein defects in signal transduction

G proteins couple receptors for many hormones to effectors that regulate second messenger metabolism. Several endocrine disorders have been shown to be caused by either loss- or gain-of-function mutations in G proteins or G protein-coupled receptors. In pseudohypoparathyroidism type Ia (PHP Ia), there are generalized hormone resistance (parathyroid hormone [PTH], thyroid-stimulating hormone, gonadotropins) and associated abnormal physical features, Albright hereditary osteodystrophy. Subjects with PHP Ib are normal in appearance and show renal resistance to PTH. In McCune-Albright syndrome (MAS), subjects show autonomous endocrine hyperfunction associated with fibrous dysplasia of bone and skin hyperpigmentation. Germline loss-of-function mutations have been identified in the G(s)-alpha gene in PHP Ia, and recent evidence suggests that the G(s)-alpha gene is paternally imprinted in a tissue-specific manner. Abnormal imprinting of the G(s)-alpha gene may be the cause of PHP Ib. MAS, in contrast, is caused by gain-of-function missense mutations of the G(s)-alpha gene.     

Prion protein and its role in signal transduction

Prion diseases are a class of fatal neurodegenerative disorders that can be sporadic, genetic or iatrogenic. They are characterized by the unique nature of their etiologic agent: prions (PrP^Sc). A prion is an infectious protein with the ability to convert the host-encoded cellular prion protein (PrP^C) into new prion molecules by acting as a template. Since Stanley B. Prusiner proposed the “protein-only” hypothesis for the first time, considerable effort has been put into defining the role played by PrP^C in neurons. However, its physiological function remains unclear. This review summarizes the major findings that support the involvement of PrP^C in signal transduction.     

Phosphoinositides and protein phosphorylation in plant signal transduction

A highly complex set of interactions are responsible for the perception and transduction of signals in living cells. It is likely that a number of fundamental principles of signalling mechanisms are of early evolutionary origin, have been highly conserved and are shared by apparently disparate organisms. Possible clues to the biochemical and molecular basis of plant signalling might thus be obtained from research carried out on other eukaryotes. Like mammalian cells, plant cells have been found to possess a phosphoinositide system and also make extensive use of phosphorylation and dephosphorylation cascades. The potential role of these mechanisms in plant cell signalling is reviewed.     

GLUT4--at the cross roads between membrane trafficking and signal transduction

GLUT4 is a mammalian facilitative glucose transporter that is highly expressed in adipose tissue and striated muscle. In response to insulin, GLUT4 moves from intracellular storage areas to the plasma membrane, thus increasing cellular glucose uptake. While the verification of this ‘translocation hypothesis’ (Cushman SW, Wardzala LJ. J Biol Chem 1980;255: 4758–4762 and Suzuki K, Kono T. Proc Natl Acad Sci 1980;77: 2542–2545) has increased our understanding of insulin-regulated glucose transport, a number of fundamental questions remain unanswered. Where is GLUT4 stored within the basal cell? How does GLUT4 move to the cell surface and what mechanism does insulin employ to accelerate this process? Ultimately we require a convergence of trafficking studies with research in signal transduction. However, despite more than 30 years of intensive research we have still not reached this point. The problem is complex, involving at least two separate signal transduction pathways which feed into what appears to be a very dynamic sorting process. Below we discuss some of these complexities and highlight new data that are bringing us closer to the resolution of these questions.     

An integrated model of epidermal growth factor receptor trafficking and signal transduction

Endocytic trafficking of many types of receptors can have profound effects on subsequent signaling events. Quantitative models of these processes, however, have usually considered trafficking and signaling independently. Here, we present an integrated model of both the trafficking and signaling pathway of the epidermal growth factor receptor (EGFR) using a probability weighted-dynamic Monte Carlo simulation. Our model consists of hundreds of distinct endocytic compartments and approximately 13,000 reactions/events that occur over a broad spatio-temporal range. By using a realistic multicompartment model, we can investigate the distribution of the receptors among cellular compartments as well as their potential signal transduction characteristics. Our new model also allows the incorporation of physiochemical aspects of ligand-receptor interactions, such as pH-dependent binding in different endosomal compartments. To determine the utility of this approach, we simulated the differential activation of the EGFR by two of its ligands, epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha). Our simulations predict that when EGFR is activated with TGF-alpha, receptor activation is biased toward the cell surface whereas EGF produces a signaling bias toward the endosomal compartment. Experiments confirm these predictions from our model and simulations. Our model accurately predicts the kinetics and extent of receptor downregulation induced by either EGF or TGF-alpha. Our results suggest that receptor trafficking controls the compartmental bias of signal transduction, rather than simply modulating signal magnitude. Our model provides a new approach to evaluating the complex effect of receptor trafficking on signal transduction. Importantly, the stochastic and compartmental nature of the simulation allows these models to be directly tested by high-throughput approaches, such as quantitative image analysis.     

A novel glycosphingolipid that may participate in embryo inversion in Volvox carteri

Mouse monoclonal antibody 4-I-244 detects a developmentally regulated antigen in embryos of Volvox carteri and inhibits specifically the morphogenetic process of inversion (the process by which the embryo turns inside out). Antigen 4-I-244 was chemically characterized as a complex phytosphingolipid containing the neutral sugars xylose, galactose, and glucose as well as inositol and phosphate.     

Normal TCR signal transduction in mice that lack catalytically active PTPN3 protein tyrosine phosphatase

PTPN3 (PTPH1) is a cytoskeletal protein tyrosine phosphatase that has been implicated as a negative regulator of early TCR signal transduction and T cell activation. To determine whether PTPN3 functions as a physiological negative regulator of TCR signaling in primary T cells, we generated gene-trapped and gene-targeted mouse strains that lack expression of catalytically active PTPN3. PTPN3 phosphatase-negative mice were born in expected Mendelian ratios and exhibited normal growth and development. Furthermore, numbers and ratios of T cells in primary and secondary lymphoid organs were unaffected by the PTPN3 mutations and there were no signs of spontaneous T cell activation in the mutant mice with increasing age. TCR-induced signal transduction, cytokine production, and proliferation was normal in PTPN3 phosphatase-negative mice. This was observed using both quiescent T cells and recently stimulated T cells where expression of PTPN3 is substantially up-regulated. We conclude, therefore, that the phosphatase activity of PTPN3 is dispensable for negative regulation of TCR signal transduction and T cell activation.     

The caveolin triad: caveolae biogenesis, cholesterol trafficking, and signal transduction

Caveolins are a family of proteins that coat the cytoplasmic face of caveolae, vesicular invaginations of the plasma membrane. These proteins are central to the organization of the proteins and lipids that reside in caveolae. Caveolins transport cholesterol to and from caveolae, and they regulate the activity of signaling proteins that reside in caveolae. Through studying the genes encoding the caveolae coat proteins, we have learned much about how they perform these multiple functions.     

Crystal structure of the VHS and FYVE tandem domains of Hrs, a protein involved in membrane trafficking and signal transduction

We have determined the 2 A X-ray structure of the 219-residue N-terminal VHS and FYVE tandem domain unit of Drosophila Hrs. The unit assumes a pyramidal structure in which the much larger VHS domain (residues 1-153) forms a rectangular base and the FYVE domain occupies the apical end. The VHS domain is comprised of an unusual "superhelix" of eight alpha helices, and the FYVE domain is mainly built of loops, two double-stranded antiparallel sheets, and a helix stabilized by two tetrahedrally coordinated zinc atoms. The two-domain structure forms an exact 2-fold-related homodimer through antiparallel association of mainly FYVE domains. Dimerization creates two identical pockets designed for binding ligands with multiple negative charges such as citrate or phosphatidylinositol 3-phosphate.     

Role of protein phosphorylation in neuronal signal transduction

Protein phosphorylation is involved in the regulation of a wide variety of physiological processes in the nervous system. Studies in which purified protein kinases or kinase inhibitors have been microinjected into defined cells while a specific response is monitored have demonstrated that protein phosphorylation is both necessary and sufficient to mediate responses of excitable cells to extracellular signals. The precise molecular mechanisms involved in neuronal signal transduction processes can be further elucidated by identification and characterization of the substrate proteins for the various protein kinases. The roles of three such substrate proteins in signal transduction are described in this article: 1) synapsin I, whose phosphorylation increases neurotransmitter release and thereby modulates synaptic transmission presynaptically; 2) the nicotinic acetylcholine receptor, whose phosphorylation increases its rate of desensitization and thereby modulates synaptic transmission postsynaptically; and 3) DARPP-32, whose phosphorylation converts it to a protein phosphatase inhibitor and which thereby may mediate interactions between dopamine and other neurotransmitter systems. The characterization of the large number of additional phosphoproteins that have been found in the nervous system should elucidate many additional molecular mechanisms involved in signal transduction in neurons.