Cdc37: a protein kinase chaperone?

The activity of most protein kinases is highly regulated, typically via phosphorylation and/or subunit association. However, the folding of protein kinases into an active state or a form capable of activation is now emerging as another important step through which they can be regulated. The 50-kDa protein Cdc37 and the associated heat-shock protein Hsp90 have been found to bind to, and be required for the activity of, diverse protein kinases, including Cdk4, v-Src, Raf and SEVENLESS. Together, Cdc37 and Hsp90 may act as a general chaperone for protein kinases, in particular those involved in signal-transduction pathways and cell-cycle control.     

Hepatic lipase: a pro- or anti-atherogenic protein?

Hepatic lipase (HL) plays a role in the metabolism of pro- and anti-atherogenic lipoproteins affecting their plasma level and composition. However, there is controversy regarding whether HL accelerates or retards atherosclerosis. Its effects on different lipoprotein classes show that, potentially, HL may promote as well as decrease atherogenesis. Studies in animals with genetically modulated HL expression show that it depends on the model used whether HL acts pro- or anti-atherogenic. In humans, HL activity seems to correlate inversely with atherosclerosis in (familial) hypercholesterolemia, and positively in hypertriglyceridemia. In normolipidemia, HL activity is weakly associated with coronary artery disease (CAD). Genetically low or absent HL activity is usually associated with increased CAD risk, especially if plasma lipid transport is impaired due to other factors. Since HL promotes the uptake of lipoproteins and lipoprotein-associated lipids, HL may affect intracellular lipid content. We hypothesize that the prime role of HL is to maintain, in concert with other factors (e.g., lipoprotein receptors), intracellular lipid homeostasis. This, and the uncertainties about its impact on human atherosclerosis, makes it difficult to predict whether HL is a suitable target for intervention to lower CAD risk. First, the physiological meaning of changes in HL activity under different conditions should be clarified.     

Protease-catalysed protein splicing: a new post-translational modification?

In all organisms, proteases catalyse peptide-bond hydrolysis and mediate protein function for a multitude of cellular processes. Mechanistically, nothing prevents proteases from also catalysing peptide-bond ligation; however, this 'reverse' reaction rarely is observed. In eukaryotes its presence has been viewed as an anomaly. Recent studies from plants and animals now challenge this assumption, indicating that protease-catalysed protein splicing is a bona fide post-translational modification. Increasing evidence indicates that the proximity of protein substrates, imposed either by their structure or by the physical constraints of the local environment, dictates when the splicing reaction will occur. This previously under-recognized splicing mechanism could increase intracellular protein diversity, thereby expanding the size of the proteome and sequence diversity beyond the predictions from genomic studies.     

Is NSF a fusion protein?

N-ethylmaleimide-sensitive fusion protein (NSF) is an ATPase required for vesicular transport throughout the constitutive secretory and endocytic pathways. Recently, NSF has also been implicated in regulated exocytosis in synapses--based on SNAP-mediated binding in vitro to a complex of neurotoxin substrates (termed 'SNAREs'). This work has generated an hypothesis in which the interaction of SNAREs (SNAP receptors) on the vesicle membrane with those on the target membrane forms a docking complex to which SNAPs bind, thus allowing NSF to bind and elicit membrane fusion. However, current evidence supports an earlier, pre-fusion role for NSF. We speculate that this role may be as a molecular chaperone for the membrane docking/fusion machinery.     

AMP-activated protein kinase: a universal regulator of autophagy?

Autophagy is a lysosomal pathway involved in the turnover of cellular macromolecules and organelles. Starvation and various other stresses increase autophagic activity above the low basal levels observed in unstressed cells, where it is kept down by mammalian target of rapamycin complex 1 (mTORC1). In starved cells, LKB1 activates AMP-activated protein kinase (AMPK) that inhibits mTORC1 activity via a pathway involving tuberous sclerosis complex 1 and 2 (TSC1/2) and its substrate Rheb. The present study suggests hat AMPK inhibits mTORC1 and autophagy also in nonstarved cells. Various Ca(2+) mobilizing agents (vitamin D compounds, thapsigargin, ATP and ionomycin) activate MPK via activation of Ca(2+)/calmodulin-dependent kinase kinase-beta (CaMKK-beta), and his pathway is required for Ca(2+)-induced autophagy. Thus, we propose that an increase in free cytosolic Ca(2+) ([Ca(2+)](c)) induces autophagy via the CaMKK/beta-AMPK-TSC1/2-Rheb-mTORC1 signaling pathway and that AMPK is a more general regulator of autophagy than previously expected.     

Autophagy: a broad role in unconventional protein secretion?

Autophagy, a cellular 'self-eating' process in eukaryotic cells, exists in both a basal and in an activated state that is induced in response to starvation. Basal and induced autophagy are associated with the packaging of cellular components, including damaged and/or redundant organelles, into double-membrane vesicles called autophagosomes, followed by autophagosome fusion with lysosomes, in which their contents are degraded and recycled. Recent results highlight a novel role for autophagy that does not involve lysosomal degradation of autophagosomal contents, but instead involves their redirection towards the extracellular delivery of an unconventionally secreted protein. Here, we discuss these findings, evaluate the strength of evidence, consider their implications for the field of protein trafficking, and suggest the next steps required to probe this interesting pathway.     

Galactose dialdehyde: the forgotten candidate for a protein cross-linker?

Chitosan derivatives with quaternary ammonium salt, such as N,N,N-trimethyl chitosan, N-N-propyl-N,N-dimethyl chitosan and N-furfuryl-N,N-dimethyl chitosan were prepared using different 96% deacetylated chitosan of M(v) 2.14x10(5), 1.9x10(4), 7.8x10(3). Amino groups on chitosan react with aldehydes to from a Schiff base intermediate. Quaternized chitosan were obtained by reaction of a Schiff base with methyl iodide. The yields, degree of quaternization and water-solubility of quaternized chitosan were influenced by the molecular weight of the chitosan sample. The antibacterial activities of quaternized chitosan against Escherichia coli were explored by calculation of the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) in water, 0.25 and 0.50% acetic acid medium. Results show the antibacterial activities of quaternized chitosan against E. coli is related to its molecular weight. Antibacterial activities of quaternized chitosan in acetic acid medium is stronger than that in water. Their antibacterial activities is increased as the concentration of acetic acid is increased. It was also found that the antibacterial activity of quaternized chitosan against E. coli is stronger than that of chitosan.     

Agouti-related protein: more than a melanocortin-4 receptor antagonist?

It is well established that agouti-related protein (AGRP) can act as a competitive antagonist to proopiomelanocortin (POMC)-derived peptides at the melanocortin-4 receptor (MC4R), and that this homeostatic mechanism is important as a means of coordinating appetite with perceived metabolic requirement. However, there are clearly additional facets to the physiological role of AGRP, given that it is active in MC4R knockout mice and it has strikingly long-lasting effects on food intake, compared with MC4R agonists. In this review we focus on: (i) evidence that AGRP is more sensitive to perturbations in energy balance than POMC and is therefore the primary basis of melanocortinergic regulation. (ii) Evidence that the bioactive peptide AGRP83-132, acts by alternate mechanism(s) to elicit its long-term effects on food intake. (iii) Evidence that AGRP is post-translationally cleaved to generate AGRP83-132 and one or more N terminal peptides, which may have an important physiological role(s) that are independent of the melanocortin system. A clear understanding of how proAGRP processing is regulated, and the role of resultant peptides, may define additional therapeutic targets in the treatment of obesity.     

The mitochondrial trifunctional protein: centre of a beta-oxidation metabolon?

The trifunctional enzyme comprises three consecutive steps in the mitochondrial beta-oxidation of long-chain acyl-CoA esters: 2-enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase and 3-ketoacyl-CoA thiolase. Deficiencies in either 3-hydroxyacyl-CoA dehydrogenase activity, or all three activities, are important causes of human disease. The dehydrogenase and thiolase have a requirement for NAD+ and CoA respectively, whose levels are conserved within the mitochondrion and thus provide possible means for control and regulation of beta-oxidation. Using analysis of the intact CoA ester intermediates produced by the complex, we have examined the sensitivity of the complex to NAD+/NADH and acetyl-CoA. We consider the evidence for channelling within the trifunctional protein and propose a model for a beta-oxidation 'metabolon'.     

The human Cas1 protein: a sialic acid-specific O-acetyltransferase?

Sialic acids are important sugars at the reducing end of glycoproteins and glycolipids. They are among many other functions involved in cell-cell interactions, host-pathogen recognition and the regulation of serum half-life of glycoproteins. An important modification of sialic acids is O-acetylation, which can alter or mask the biological properties of the parent sialic acid molecule. The nature of mammalian sialate-O-acetyltransferases (EC 2.3.1.45) involved in their biosynthesis is still unknown. We have identified the human CasD1 (capsule structure1 domain containing 1) gene as a candidate to encode the elusive enzyme. The human CasD1 gene encodes a protein with a serine-glycine-asparagine-histidine hydrolase domain and a hydrophobic transmembrane domain. Expression of the Cas1 protein tagged with enhanced green fluorescent protein in mammalian and insect cells directed the protein to the medial and trans-cisternae of the Golgi. Overexpression of the Cas1 protein in combination with α-N-acetyl-neuraminide α-2,8-sialyltransferase 1 (GD3 synthase) resulted in an up to 40% increased biosynthesis of 7-O-acetylated ganglioside GD3. By quantitative real-time polymerase chain reaction, we found up to 5-fold increase in CasD1 mRNA in tumor cells overexpressing O-Ac-GD3. CasD1-specific small interfering RNA reduced O-acetylation in tumor cells. These results suggest that the human Cas1 protein is directly involved in O-acetylation of α2-8-linked sialic acids.     

Polyglutamylation: a fine‐regulator of protein function?

Polyglutamylation is a post-translational modification in which glutamate side chains of variable lengths are formed on the modified protein. It is evolutionarily conserved from protists to mammals and its most prominent substrate is tubulin, the microtubule (MT) building block. Various polyglutamylation states of MTs can be distinguished within a single cell and they are also characteristic of specific cell types or organelles. Polyglutamylation has been proposed to be involved in the functional adaptation of MTs, as it occurs within the carboxy-terminal tubulin tails that participate directly in the binding of many structural and motor MT-associated proteins. The discovery of a new family of enzymes that catalyse this modification has brought new insight into the mechanism of polyglutamylation and now allows for direct functional studies of the role of tubulin polyglutamylation. Moreover, the recent identification of new substrates of polyglutamylation indicates that this post-translational modification could be a potential regulator of diverse cellular processes.     

Ageing and the regulation of protein synthesis: a balancing act?

Ageing in diverse species ranging from yeast to humans is associated with extensive changes in both general and specific protein synthesis. Accumulating evidence now indicates that these alterations are not simply a corollary of the ageing process but, rather, they have a causative role in senescent decline. Indeed, interfering with mRNA translation significantly influences longevity. Interestingly, the mechanisms that control mRNA translation interface with intricate, conserved signalling pathways and specific conditions that regulate ageing, such as the insulin-insulin growth factor 1 signalling pathway and caloric restriction. This emerging relationship reveals that protein synthesis is a novel determinant of ageing in diverse organisms such as yeast, worms, flies and mice and can thus be considered as a universal component of the ageing process.     

Nonvesicular sterol transport: two protein families and a sterol sensor?

Sterols, essential components of eukaryotic membranes, are actively transported between cellular membranes. Although it is known that both vesicular and non-vesicular means are used to move sterols, the molecules and molecular mechanisms involved have yet to be identified. Recent studies point to a key role for oxysterol binding protein (OSBP) and its related proteins (ORPs) in nonvesicular sterol transport. Here, evidence that OSBP and ORPs are bona fide sterol carriers is discussed. In addition, I hypothesize that ATPases associated with various cellular activities regulate the recycling of soluble lipid carriers and that the Niemann Pick C1 protein facilitates the delivery of sterols from endosomal membranes to ORPs and/or the ensuing membrane dissociation of ORPs.