Insulin signal transduction through protein kinase cascades

This review summarizes the evolution of ideas concerning insulin signal transduction, the current information on protein ser/thr kinase cascades as signalling intermediates, and their status as participants in insulin regulation of energy metabolism. Best characterized is the Ras-MAPK pathway, whose input is crucial to cell fate decisions, but relatively dispensable in metabolic regulation. By contrast the effectors downstream of PI-3 kinase, although less well elucidated, include elements indispensable for the insulin regulation of glucose transport, glycogen and cAMP metabolism. Considerable information has accrued on PKB/cAkt, a protein kinase that interacts directly with Ptd Ins 3OH phosphorylated lipids, as well as some of the elements further downstream, such as glycogen synthase kinase-3 and the p70 S6 kinase. Finally, some information implicates other erk pathways (e.g. such as the SAPK/JNK pathway) and Nck/cdc42-regulated PAKs (homologs of the yeast Ste 20) as participants in the cellular response to insulin. Thus insulin recruits a broad array of protein (ser/thr) kinases in its target cells to effectuate its characteristic anabolic and anticatabolic programs.     

The role of tyrosine phosphorylation in regulation of signal transduction pathways in unicellular eukaryotes

The review summarizes for the first time the current concepts of the role of tyrosine phosphorylation in regulation of signal transduction pathways in unicellular eukaryotes. Evolutionary concepts are developed about the origin of protein tyrosine kinases (PTK)-signaling.     

The role of membrane potential shifts at the initial stages of signal transduction in unicellular eukaryotes

The review summariezes current data about the origin of membrane potentials and their contribution to the signal transduction about changes in temperature, ionic composition of the medium, and chemo-, photo-, gravi-taxis signals in unicellular eukaryotes.     

MAP kinase cascades in elicitor signal transduction

 Protein kinases play important roles in elicitor signal transduction. In this article, I describe the current view of the role of mitogen-activated protein kinase (MAPK) cascades in elicitor signal transduction of plant cells based on our own research and recent developments in this field. In the past several years, it has become apparent that MAPK cascades play important roles in elicitor signal transduction in plants. Our early studies demonstrated the identification of p47 MAPK in tobacco as an elicitor-responsive protein kinase and possible involvement of p47 MAPK in elicitor signal transduction to induce defense responses, including defense gene expression and hypersensitive cell death. However, the molecular identity of p47 MAPK is still unclear. Recent important studies suggest that tobacco MAPK cascades that include SIPK, and/or WIPK, and NtMEK2, an upstream kinase for both SIPK and WIPK, have a crucial function in induction of defense responses and hypersensitive cell death. The orthologs of these protein kinases in Arabidopsis and alfalfa are also suggested to have similar functions. Furthermore, the identification of loss-of-function mutation in Arabidopsis reveals a negative regulatory role for putative MAPK cascades in plant defense mechanisms. Received: February 7, 2002 / Accepted: February 25, 2002     

Pathways of transduction of apoptotic signals in unicellular eukaryotes

The review summarizes current data about transduction mechanism of apoptotic signals by caspase-like enzymes and mitochondrial apoptogenic proteins in unicellular eukaryotes. The role of receptor-dependent and receptor-independent caspase cascades is rewieved. The special attention is given to evolution aspects of problem of apoptosis.     

The role of protein kinase C in insulin signal transduction via adenylyl cyclase signaling mechanism

Activation of proteinkinase C with diacylglycerol or phorbol-12-myristate-13-acetate in the rat muscle membrane or Anodonta cygnea mollusc blocks the insulin stimulating signal to adenylyl cyclase via tyrosinekinase type receptor. The same occurs with stimulating effect of biogenic amines to adenylyl cyclase via serpentine type receptor. Transduction of the inhibitory signal induced with isoproterenol to adenylyl cyclase remained unchanged in case of the proteinkinase C activation. The findings suggest that phorbol-sensitive proteinkinase C realizes a negative regulation of insulin-sensitive adenylyl cyclase signalling system. This negative regulation might prove a universal mechanism of the adenylyl cyclase system desensitisation.     

Regulation of protein kinase C by lysophospholipids. Potential role in signal transduction

Certain lysophospholipids, lysophosphatidylcholine (lyso-PC) in particular, stimulated protein kinase C at low concentrations (less than 20 microM) but, conversely, inhibited it at high concentrations (greater than 30 microM). Protein kinase C stimulation by lyso-PC required the presence of phosphatidylserine (PS) and Ca2+ and was associated with a decreased Ka for PS and increased Ka for Ca2+ of the enzyme. Cardiolipin and phosphatidic acid could partially substitute for PS in supporting the stimulatory effect of lyso-PC. Lyso-PC also biphasically regulated protein kinase C activated by diolein. Of several synthetic lyso-PC preparations tested, the oleoyl, myristoyl and palmitoyl derivatives were most active. Data from the Triton X-100 mixed micellar assay indicated that 1.4 and 14.0 mol of lyso-PC/micelle produced a maximal stimulation and a complete abolishment of the stimulation of protein kinase C, respectively. Protein kinase C stimulation by lyso-PC, with a pH optimum of about 7.5, was observed for phosphorylation of histone H1, myelin basic protein, and the 35- and 47-kDa proteins from the rat brain, but not for that of other histone subfractions and protamine. Lyso-PC acted synergistically with diacylglycerol in stimulating protein kinase C, whereas the stimulation by lyso-PC was additive to that by oleic acid. Protein kinase C inhibitors (alkyllysophospholipid, sphingosine, tamoxifen, and polymyxin B) inhibited more potently the protein kinase C activity stimulated by PS/Ca2+/lyso-PC than that stimulated by PS/Ca2+. The stimulatory and inhibitory effects of lyso-PC were not observed for myosin light chain kinase and cAMP-dependent protein kinase, indicating a specificity of its actions. The present findings suggested that lyso-PC, likely derived from membrane PC by the action of phospholipase A2, might play a role in signal transduction via a dual regulation of protein kinase C, and that it could further modulate the enzyme and hence the cellular activity by interplaying with diacylglycerol and unsaturated fatty acid, the two other classes of cellular mediators also shown to be activators of protein kinase C.     

Effects of sequestration on signal transduction cascades

The building blocks of most signal transduction pathways are pairs of enzymes, such as kinases and phosphatases, that control the activity of protein targets by covalent modification. It has previously been shown [Goldbeter A & Koshland DE (1981) Proc Natl Acad Sci USA 78, 6840-6844] that these systems can be highly sensitive to changes in stimuli if their catalysing enzymes are saturated with their target protein substrates. This mechanism, termed zero-order ultrasensitivity, may set thresholds that filter out subthreshold stimuli. Experimental data on protein abundance suggest that the enzymes and their target proteins are present in comparable concentrations. Under these conditions a large fraction of the target protein may be sequestrated by the enzymes. This causes a reduction in ultrasensitivity so that the proposed mechanism is unlikely to account for ultrasensitivity under the conditions present in most in vivo signalling cascades. Furthermore, we show that sequestration changes the dynamics of a covalent modification cycle and may account for signal termination and a sign-sensitive delay. Finally, we analyse the effect of sequestration on the dynamics of a complex signal transduction cascade: the mitogen-activated protein kinase (MAPK) cascade with negative feedback. We show that sequestration limits ultrasensitivity in this cascade and may thereby abolish the potential for oscillations induced by negative feedback.     

Autophagy in unicellular eukaryotes

Cells need a constant supply of precursors to enable the production of macromolecules to sustain growth and survival. Unlike metazoans, unicellular eukaryotes depend exclusively on the extracellular medium for this supply. When environmental nutrients become depleted, existing cytoplasmic components will be catabolized by (macro)autophagy in order to re-use building blocks and to support ATP production. In many cases, autophagy takes care of cellular housekeeping to sustain cellular viability. Autophagy encompasses a multitude of related and often highly specific processes that are implicated in both biogenetic and catabolic processes. Recent data indicate that in some unicellular eukaryotes that undergo profound differentiation during their life cycle (e.g. kinetoplastid parasites and amoebes), autophagy is essential for the developmental change that allows the cell to adapt to a new host or form spores. This review summarizes the knowledge on the molecular mechanisms of autophagy as well as the cytoplasm-to-vacuole-targeting pathway, pexophagy, mitophagy, ER-phagy, ribophagy and piecemeal microautophagy of the nucleus, all highly selective forms of autophagy that have first been uncovered in yeast species. Additionally, a detailed analysis will be presented on the state of knowledge on autophagy in non-yeast unicellular eukaryotes with emphasis on the role of this process in differentiation.     

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.     

The role of phosphoinositides, protein kinase C and protein kinase A in the K+ regulatory signal transduction in human adrenocortical cells

The messenger mechanisms mediating K+ regulatory signals in human adrenocorticocytes were studied. It was shown that potassium ions initiated decay of polyphosphoinositides to inositolphosphates and obviously diacylglycerol. The latter compounds activate protein kinase C as affected by different agonists. Using western blotting method we showed translocation of PKCalpha from cytosol to membranes after adrenal tissue preincubation in the medium with increased K+ content (8.5 mM). Translocation means activation of the enzyme. Activity of PKC increased in the microsomal fraction and did not change in cytosol. Increased concentration of K+ in the incubation medium also activates protein kinase A, although to a lesser extent compared to PKC. Unlike PKC activity of PKA was changed in cytosol as well. The possibility of involvement of several messenger systems in K+ signal transduction in human adrenocortical cells as well as the hypothesis on cross-talk between messenger mechanisms for main physiological agonists controlling aldosterone biosynthesis in the adrenals are discussed.     

The behaviour of unicellular eukaryotes.

The field of ciliated protozoa behaviour is reconsidered. The simple-complexity of their behaviour is seen in its basic nature and adaptive meaning. The Laboratory Conditions (LABCON) are discussed in comparison with those occurring in nature (NATCON). The nature of the arc, the behavioural element commonly performed by a ciliate, is discussed in detail. Several indexes and rates can perfectly describe the tracks of the ciliates. The behaviour can be considered as a kind of adaptive interface between the environment and the organism.     

Sequestration shapes the response of signal transduction cascades

Many signal transduction cascades are composed of covalent modification cycles such as kinase/phosphatase cycles. In the 1980s Goldbeter and Koshland showed that such cycles can exhibit non-linear input-output relations when the enzymes are saturated by their substrates, which may facilitate signal processing. Recent papers show that this mechanism is unlikely to cause non-linearity in mammalian signal transduction cascades as sequestration of the target due to enzyme concentrations present in these cascades will hamper this mechanism. However, sequestration due to high-affinity enzymes can shape the dynamics and steady-state behaviour of signal transduction cascades in different ways, some of which are discussed in this review.     

Functional partitioning of epithelial protein kinase CaMKII in signal transduction

The examination of the physiological role of CaMKII has made substantial progress in non-epithelial systems but little is known about its function in secretory epithelial cells. A prototypic exocrine cell, the acid secreting gastric parietal cell, largely redistributes its cytoplasmic tubulocisternal membranes (TC) to the secretory apical membrane (SA) after stimulation. We here present a dependence of physiological response on partitioned initial CaMKII activities redistributed between TC and SA. Initial acid secretion after cholinergic stimulation has doubled if activities of phosphatases PP1/PP2A and protein kinase PKC-alpha were inhibited by the inhibitors calyculin A and G? 6976. CaMKII appears to be integrated in multienzyme complexes each specific for TC and SA. Therein, PP1/PP2A and PKC-alpha were found to permanently counteract initial CaMKII activities in different modes of transregulation. Remarkably, a dys-transregulated, hyperactive CaMKII at TC did not result in an increased acid secretion to the same extent. A simple mathematical model to estimate the share of TC- and SA-associated CaMKII activities in cholinergically induced acid secretion suggests that TC-associated CaMKII appears to autoregulate its contribution to the physiological response by a negative feedback control. Subcellular distribution and stimulus-dependent redistribution of domain-associated CaMKII signalosomes indicate a fine balanced, adaptive system to guarantee a stable physiological response.