Yeast oxysterol-binding proteins: sterol transporters or regulators of cell polarization?

Oxysterol-binding protein (OSBP) and OSBP-related proteins (ORPs) are a conserved family of soluble cytoplasmic proteins that can bind sterols, translocate between membrane compartments, and affect sterol trafficking. These properties make ORPs attractive candidates for lipid transfer proteins (LTPs) that directly mediate nonvesicular sterol transfer to the plasma membrane. To test whether yeast ORPs (the Osh proteins) are sterol LTPs, we studied endoplasmic reticulum (ER)-to-plasma membrane (PM) sterol transport in OSH deletion mutants lacking one, several, or all Osh proteins. In conditional OSH mutants, ER-PM ergosterol transport slowed ~20-fold compared with cells expressing a full complement of Osh proteins. Although this initial finding suggested that Osh proteins act as sterol LTPs, the situation is far more complex. Osh proteins have established roles in Rho small GTPase signaling. Osh proteins reinforce cell polarization and they specifically affect the localization of proteins involved in polarized cell growth such as septins, and the GTPases Cdc42p, Rho1p, and Sec4p. In addition, Osh proteins are required for a specific pathway of polarized secretion to sites of membrane growth, suggesting that this is how Osh proteins affect Cdc42p- and Rho1p-dependent polarization. Our findings suggest that Osh proteins integrate sterol trafficking and sterol-dependent cell signaling with the control of cell polarization.     

Are neuronal SNARE proteins Ca2+ sensors?

The neuronal SNARE complex formed by synaptobrevin, syntaxin and SNAP-25 plays a central role in Ca2+-triggered neurotransmitter release. The SNARE complex contains several potential Ca2+-binding sites on the surface, suggesting that the SNAREs may be involved directly in Ca2+-binding during release. Indeed, overexpression of SNAP-25 bearing mutations in two putative Ca2+ ligands (E170A/Q177A) causes a decrease in the Ca2+-cooperativity of exocytosis in chromaffin cells. To test whether the SNARE complex might function in Ca2+-sensing, we analyzed its Ca2+-binding properties using transverse relaxation optimized spectroscopy (TROSY)-based NMR methods. Several Ca2+-binding sites are found on the surface of the SNARE complex, but most of them are not specific for Ca2+ and all have very low affinity. Moreover, we find that the E170A/Q177A SNAP-25 mutation does not alter interactions between the SNAREs and the Ca2+ sensor synaptotagmin 1, but severely impairs SNARE complex assembly. These results suggest that the SNAREs do not act directly as Ca2+ receptors but SNARE complex assembly is coupled tightly to Ca2+-sensing during neurotransmitter release.     

ABC transporters: one,two or four extracytoplasmic substrate-binding sites?

Two families of ATP-binding cassette (ABC) transporters in which one or two extracytoplasmic substrate-binding domains are fused to either the N- or C-terminus of the translocator protein have been detected. This suggests that two, or even four, substrate-binding sites may function in the ABC transporter complex. This domain organization in ABC transporters, widely represented among microorganisms, raises new possibilities for how the substrate-binding protein(s) (SBPs) might interact with the translocator. One appealing hypothesis is that multiple substrate-binding sites in proximity to the entry site of the translocation pore enhance the transport capacity. We also discuss the implications of multiple substrate-binding sites in close proximity to the translocator in terms of broadened substrate specificity and possible cooperative interactions between SBPs and the translocator.     

ABC lipid transporters: extruders, flippases, or flopless activators?

Many mammalian ABC transporters move membrane lipids to acceptor lipid assemblies in the extracellular aqueous milieu. Because the desorption from the membrane costs more energy than provided by two ATPs, the transporter probably only translocates the lipid to a partially hydrophilic site on its extracellular face. From this high-energy site, the lipid may efficiently move to the acceptor, which ideally is bound to the transporter, or, in the absence of an acceptor, fall back into the membrane. If the lipid originated from the cytosolic membrane surface, this represents lipid flop and is probably a side activity of the transporters.     

Microbial cholesterol oxidases: bioconversion enzymes or signal proteins?

Cholesterol oxidases (3beta-hydroxysterol oxidases; EC 1.1.3.6), serve as catalysts for the initial step in the degradation of cholesterol, and probably other natural sterols, that are used as carbon sources for growth of different bacteria. Because of their suitability for attacking cholesterol they have been widely used for the quantification of cholesterol in clinical and food specimens. Cholesterol oxidase has also found application as a probe for membrane structure, as an insecticide, and has been implicated in bacterial pathogenesis. Recently, we have found that a Streptomyces cholesterol oxidase is required for the biosynthesis of the antifungal polyene pimaricin, apparently acting as an antifungal sensor. Here we describe our current understanding of these fascinating enzymes.     

Nucleocytoplasmic trafficking of proteins: With or without Ran?

Proteins and RNAs move between the nucleus and cytoplasm by translocation through nuclear pore complexes in the nuclear envelope. To do this, they require specific targeting signals, energy, and a cellular apparatus that catalyzes their transport. Several of the factors involved in nucleocytoplasmic trafficking of proteins have been identified and characterized in some detail. The emerging picture for nuclear transport proposes a central role for the small GTPase Ran and proteins with which it interacts. In particular, asymmetric distribution of these proteins between nucleus and cytoplasm appears to be responsible for the vectorial nature of nucleocytoplasmic transport. Here, we summarize the role of Ran and Ran-binding proteins in nuclear trafficking of proteins with classical nuclear localisation signals. We also discuss examples of the growing number of alternative pathways that are involved in transport of proteins across the nuclear envelope. BioEssays 21:579–589, 1999. © 1999 John Wiley & Sons, Inc.     

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.     

Plakophilins: Multifunctional proteins or just regulators of desmosomal adhesion?

Plakophilins 1-3 are members of the p120(ctn) family of armadillo-related proteins. The plakophilins have been characterized as desmosomal proteins, whereas p120(ctn) and the closely related delta-catenin, ARVCF and p0071 associate with adherens junctions and play essential roles in stabilizing cadherin mediated adhesion. Recent evidence suggests that plakophilins are essential components of the desmosomal plaque where they interact with desmosomal cadherins as well as the cytoskeletal linker protein desmoplakin. Plakophilins stabilize desmosomal proteins at the plasma membrane and therefore may function in a manner similar to p120(ctn) in the adherens junctions. The three plakophilins reveal distinct expression patterns, and although partially redundant in their function, mediate distinct effects on desmosomal adhesion. Besides a structural role, a function in signaling has been postulated in analogy to other armadillo proteins such as beta-catenin. At least plakophilins 1 and 2 are also localized in the nucleus, and all three proteins occur in a cytoplasmic pool. This review aims to summarize the current knowledge of plakophilin function in the context of cell adhesion, signaling and their putative role in diseases.     

Dipeptidyl peptidase IV-like molecules: homologous proteins or homologous activities?

Membrane-bound proteases are widely distributed among various cell systems. Their expression in a particular cell type is finely regulated, reflecting the specific functional cell implications and engagement in defined physiological pathways. Protein turnover, ontogeny, inflammation, tissue remodeling, cell migration and tumor invasion are among the many physiological and pathological events in which membrane proteases play a crucial role, both as effector as well as regulatory molecules. The presence of proline residues gives unique structural features to peptide chains, substantially influencing the susceptibility of proximal peptide bond to protease cleavage. Among the rare group of proline-specific proteases, dipeptidyl peptidase IV (DPP-IV, EC 3.4.14.5) was originally believed to be the only membrane-bound enzyme specific for proline as the penultimate residue at the amino-terminus of the polypeptide chain. However, other molecules, even structurally non-homologous with the DPP-IV but bearing corresponding enzyme activity, have been identified recently. This review summarizes the present knowledge of "DPP-IV activity- and/or structure-homologues" (DASH) and provides some insight into their multifunctional roles.     

Bcl-2 proteins: regulators of apoptosis or of mitochondrial homeostasis?

Programmed cell death (apoptosis) is used by multicellular organisms during development and to maintain homeostasis within mature tissues. One of the first genes shown to regulate apoptosis was bcl-2. Subsequently, a number of Bcl-2-related proteins have been identified. Despite overwhelming evidence that Bcl-2 proteins are evolutionarily conserved regulators of apoptosis, their precise biochemical function remains controversial. Three biochemical properties of Bcl-2 proteins have been identified: their ability to localize constitutively and/or inducibly to the outer mitochondrial, outer nuclear and endoplasmic reticular membranes, their ability to form heterodimers with proteins bearing an amphipathic helical BH3 domain, and their ability to form ion-conducting channels in synthetic membranes. The discovery that mitochondria can play a key part in the induction of apoptosis has focused attention on the role that Bcl-2 proteins may have in regulating either mitochondrial physiology or mitochondria-dependent caspase activation. Here we attempt to synthesize our current understanding of the part played by mitochondria in apoptosis with a consideration of how Bcl-2 proteins might control cell death through an ability to regulate mitochondrial physiology.     

Heat shock proteins in oncology: Diagnostic biomarkers or therapeutic targets?

Heat shock proteins (HSP) are a family of proteins induced in cells exposed to different insults. This induction of HSPs allows cells to survive stress conditions. Mammalian HSPs have been classified into six families according to their molecular size: HSP100, HSP90, HSP70, HSP60, HSP40 and small HSPs (15 to 30kDa) including HSP27. These proteins act as molecular chaperones either helping in the refolding of misfolded proteins or assisting in their elimination if they become irreversibly damaged. In recent years, proteomic studies have characterized several different HSPs in various tumor types which may be putative clinical biomarkers or molecular targets for cancer therapy. This has led to the development of a series of molecules capable of inhibiting HSPs. Numerous studies speculated that over-expression of HSP is in part responsible for resistance to many anti-tumor agents and chemotherapeutics. Hence, from a pharmacological point of view, the co-administration of HSP inhibitors together with other anti-tumor agents is of major importance in overcoming therapeutic resistance. In this review, we provide an overview of the current status of HSPs in autoimmune, cardiovascular, and neurodegenerative diseases with special emphasis on cancer.     

Uncoupling proteins (UCP) in unicellular eukaryotes: True UCPs or UCP1-like acting proteins?

Uncoupling proteins belong to the superfamily of mitochondrial anion carriers. They are apparently present throughout the Eukarya domain in which only some members have an established physiological function, i.e. UCP1 from brown adipose tissue is involved in non-shivering thermogenesis. However, the proteins responsible for the phenotype observed in unicellular organisms have not been characterized. In this report we analyzed functional evidence concerning unicellular UCPs and found that true UCPs are restricted to some taxonomical groups while proteins conferring a UCP1-like phenotype to fungi and most protists are the result of a promiscuous activity exerted by other mitochondrial anion carriers. We describe a possible evolutionary route followed by these proteins by which they acquire this promiscuous mechanism.     

Cytoplasmic incompatibility in insects: Why sterilize females?

Cytoplasmic incompatibility (CI) in insects is an intrapopulational sterility phenomenon. Although known for nearly 40 years, it has only recently attracted the attention of evolutionary biologists, having been found in an increasing number of species. Apparently, the proximate cause of CI is the presence of rickettsia-like endocellular microorganisms. An ultimate cause can be identified in the fact that males do not transmit cytoplasmic genes, including those of rickettsia.     

Mud-puddling behavior in tropical butterflies: in search of proteins or minerals?

We experimentally investigated the attraction of adult butterflies to moist soil and dirt places (a behavior termed `mud-puddling') in two species-rich tropical communities on the island of Borneo. At a rain forest site, 227 individuals (46 species) were attracted to the baits, compared to 534 individuals (54 species) at a farmland site. With one single exception, all attracted butterflies were males. Of various salt and amino acid solutions, only sodium was accepted, but overall, albumin solutions turned out to be the most attractive puddling resource. Butterfly families differed consistently in their resource preferences. Representatives of the families Papilionidae and Pieridae more often visited NaCl solutions, but still accepted albumin, whereas representatives of the Nymphalidae, Hesperiidae and, in particular, Lycaenidae preferred the protein resource. In experiments using decoys prepared from pinned butterfly specimens, representatives of the Papilionidae and Pieridae were more strongly attracted to baits provided with decoys made from conspicuous, medium-sized yellow Eurema species (Pieridae), whereas dummies made from small, cryptically colored lycaenids (Prosotas and Caleta species) were ineffective. Decoys did not influence the attraction of lycaenid butterflies towards baits. Hence, visual cues play an important role in locating puddling resources for papilionids and pierids, while for lycaenid butterflies searching for nitrogen sources, olfactory cues emitted by decaying organic matter are more likely to be important. The strong attraction of male butterflies to nitrogen-rich resources suggests that, as in the case of sodium, these nutrients may increase reproductive success. Received: 5 October 1998 / Accepted: 7 December 1998