FIP-2, a coiled-coil protein, links Huntingtin to Rab8 and modulates cellular morphogenesis

AbstractHuntington's disease is characterised by the death of cortical and striatal neurons, and is the result of an expanded polyglutamine tract in the Huntingtin protein [1]. Huntingtin is present on both endocytic and secretory membrane organelles but its function is unclear [2,3]. Rab GTPases regulate both of these transport pathways [4]. We have previously shown that Rab8 controls polarised membrane transport by modulating cell morphogenesis [5]. To understand Rab8-mediated processes, we searched for Rab8-interacting proteins by the yeast two-hybrid system. Here, we report that Huntingtin is linked to the Rab8 protein through FIP-2, a tumour necrosis factor-α (TNF-α)-inducible coiled-coil protein related to the NEMO protein [6,7]. The activated form of Rab8 interacted with the amino-terminal region of FIP-2, whereas dominant-negative Rab8 did not. Huntingtin bound to the carboxy-terminal region of FIP-2. Coexpressed FIP-2 and Huntingtin enhanced the recruitment of Huntingtin to Rab8-positive vesicular structures, and FIP-2 promoted cell polarisation in a similar way to Rab8. We propose a model in which Huntingtin, together with FIP-2 and Rab8, are part of a protein network that regulates membrane trafficking and cellular morphogenesis

Clorinated hydrocarbons and mercury in zooplankton near the coast of Finland.

Tiivistelmä: Kloorattujen hiilivetyjen ja elohopean esiintymisestä eläinplanktonissa Suomen rannikkovesissä

Die Modellierung des sportlichen Erfolges : Implikationen für das Sponsoring

Die Auswahl von geeigneten Sponsoring-Engagements mit dem Ziel einer Verbesserung des Unternehmensergebnisses nimmt einen immer wichtigeren Stellenwert im Marketing von Unternehmen ein. Da gerade im Sportsponsoring der finanzielle Erfolg des Sponsors entscheidend vom Erfolg des Gesponserten abhängt, stehen Marketingmanager vor der Herausforderung, den sportlichen Erfolg verlässlich abzuschätzen. Diese Untersuchung stellt daher ein Modell zur Prognose von sowohl kurzfristigem als auch langfristigem sportlichen Erfolg vor. Als primäre Erkenntnisse lassen sich dabei festhalten, dass der kurzfristige sportliche Erfolg durch Leistungs-, situative und Marketingfaktoren relativ gut prognostiziert werden kann und dass die Ergebnisse der kurzfristigen Perspektive ausreichen, um den langfristigen sportlichen Erfolg gut (r = 0,61) vorherzusagen. Die Untersuchung umfasst dabei mehr als 34.000 Spiele des deutschen Profifußballs und berücksichtigt damit mehr als 150 Mannschaften in 35 Jahren. Eine Analyse mittels ordinaler LogitRegression und Markov-Chain-Monte-Carlo-Simulationen zeigt, dass neben der Teamstärke insbesondere die Bedeutung des Spiels für die sportlichen Ziele den größten Einfluss auf den sportlichen Erfolg ausübt. Zudem zeigt sich, dass der sportliche Erfolg positiv durch Fan Commitment und Markenstärke, allerdings negativ durch die Rivalität zwischen den Teams beeinflusst wird. Das kalibrierte Modell, welches auf Basis öffentlich zugänglicher und größtenteils objektiver Informationen ermittelt wird, kann Managern als Tool zur Festlegung des optimalen SponsoringEngagements dienen

Rab8 and Rab8-interacting proteins as players in cell polarization

Rab8 and its interacting proteins as regulators of cell polarization During the development of a multi-cellular organism, progenitor cells have to divide and migrate appropriately as well as organize their differentiation with one another, in order to produce a viable embryo. To divide, differentiate and migrate cells have to undergo polarization, a process where internal and external components such as actin, microtubules and adhesion receptors are reorganized to produce a cell that is asymmetric, with functionally different surfaces. Also in the adult organism there is a continuous need for these processes, as cells need to migrate in response to tissue damage and to fight infection. Improper regulation of cell proliferation and migration can conversely lead to disease such as cancer. GTP-binding proteins function as molecular switches by cycling between a GTP-bound (active) conformation and a GDP-bound (inactive) conformation. The Ras super-family of small GTPases are found in all eukaryotic cells. They can be functionally divided into five subfamilies. The Ras family members mainly regulate gene expression, controlling cell proliferation and differentiation. Ras was in fact the first human oncogene to be characterized, and as much as 30% of all human tumors may be directly or indirectly caused by mutations of Ras molecules The Rho family members mainly regulate cytoskeletal reorganization. Arf proteins are known to regulate vesicle budding and Rab proteins regulate vesicular transport. Ran regulates nuclear transport as well as microtubule organization during mitosis. The focus of the thesis of Katarina Hattula, is on Rab8, a small GTPase of the Rab family. Activated Rab8 has previously been shown to induce the formation of new surface extensions, reorganizing both actin and microtubules, and to have a role in directed membrane transport to cell surfaces. However, the exact membrane route it regulates has remained elusive. In the thesis three novel interactors of Rab8 are presented. Rabin8 is a Rab8-specific GEF that localizes to vesicles where it presumably recruits and activates its target Rab8. Its expression in cells leads to remodelling of actin and the formation of polarized cell surface domains. Optineurin, known to be associated with a leading cause of blindness in humans (open-angle glaucoma), is shown to interact specifically with GTP-bound Rab8. Rab8 binds to an amino-terminal region and interestingly, the Huntingtin protein binds a carboxy-terminal region of optineurin. (Aberrant Huntingtin protein is known to be the cause Huntington s disease in humans.) Co-expression of Huntingtin and optineurin enhanced the recruitment of Huntingtin to Rab8-positive vesicular structures. Furthermore, optineurin promoted cell polarization in a similar way to Rab8. A third novel interactor of Rab8 presented in this thesis is JFC1, a member of the synaptogamin-like protein (Slp) family. JFC1 interacts with Rab8 specifically in its GTP-bound form, co-localizes with endogenous Rab8 on tubular and vesicular structures, and is probably involved in controlling Rab8 membrane dynamics. Rab8 is in this thesis work clearly shown to have a strong effect on cell shape. Blocking Rab8 activity by expression of Rab8 RNAi, or by expressing the dominant negative Rab8 (T22N) mutant leads to loss of cell polarity. Conversely, cells expressing the constitutively active Rab8 (Q67L) mutant exhibit a strongly polarized phenotype. Experiments in live cells show that Rab8 is associated with macropinosomes generated at ruffling areas of the membrane. These macropinosomes fuse with or transform into tubules that move toward the cell centre, from where they are recycled back to the leading edge to participate in protrusion formation. The biogenesis of these tubules is shown to be dependent on both actin and microtubule dynamics. The Rab8-specific membrane route studied contained several markers known to be internalized and recycled (1 integrin, transferrin, transferrin receptor, cholera toxin B subunit (CTxB), and major histocompatibility complex class I protein (MHCI)). Co-expression studies revealed that Rab8 localization overlaps with that of Rab11 and Arf6. Rab8 is furthermore clearly functionally linked to Arf6. The data presented in this thesis strongly suggests a role for Rab8 as a regulator for a recycling compartment, which is involved in providing structural and regulatory components to the leading edge to participate in protrusion formation.Rab8 reglerar cellens form och utseende. En avhandling vid medicinska fakulteten, Helsingfors universitet beskriver hur det GTP bindande proteinet Rab8 reglerar cellens form och utseende (cellpolarisering). I en flercellig organism måste celler specialicera sig och röra på sig för att organismen skall utvecklas korrekt. För att detta skall kunna ske måste de först ändra form i en process kallad polarisering. Cellens komponenter organiseras då assymetriskt för att bilda funktionellt skilda delar. Att en cell kan röra på sig har inte bara en essentiell funktion under utvecklingsskedet, det är också en process som behövs under hela organismens livstid. Det behövs t.ex. rörliga celler för att bekämpa infektioner och för att sår skall kunna läkas. Det är också viktigt att celler som inte skall röra på sig förblir stilla. Strikt kontroll av vilka celler som skall och inte skall röra på sig behövs kontinuerligt. Ett exempel på vad som kan hända om denna kontroll inte fungerar är cancer. Små GTPaser är en stor grupp proteiner som fungerar som omkopplare eller strömbrytare i vidareföring av signaler inuti cellen. Deras funktioner i cellen är diversa och inkluderar bland annat kontroll av celltillväxt, celldelning, och membrantransport. Denna stora grupp (fler än 100 proteiner) kan delas upp i fem familjer: Ras, Rho, Rab, Arf och Ran. Ras var den första humana onkogenen att beskrivas 1981, och i uppskattningsvis en tredjedel av all cancer finns mutationer som leder till ett permanent aktiverat Ras-protein. Rab-familjen av små GTPaser kontrollerar membrantransport i cellen och det är en medlem av denna familj, Rab8, som är fokuset av Katarina Hattulas avhandling. Tidigare forskning har inte klart kunnat visa vilken roll Rab8 har i cellen. I avhandlingen ges bevis på att Rab8 kraftigt påverkar cellens form och utseende. Resultatet av den forskning som presenteras visar en roll för Rab8 i återvinning av material som behövs för att en cell skall kunna bilda och upprätthålla en polariserad fenotyp. Något som bland annat behövs för att cellen skall kunna röra på sig. Avhandlingen presenterar också i detalj hur man sökt efter proteiner som specifikt kan binda till och påverka Rab8s funktion. Tre nya Rab8 bindande protein beskrivs. Ett av dessa är optineurin, som på annat håll har visats vara knutet till glaukom, (en av de främsta anledningarna till blindhet i världen med 33 miljoner drabbade). En koppling mellan optineurin och Huntingtin, det protein vars gen är muterad i personer med Huntingtons sjukdom visas också. Ett annat Rab8 bindande protein som beskrivs är Rabin8. Det aktiverar Rab8 och behövs för att Rab8 ska kunna utföra sin uppgift vid cellpolartisering. Sammantaget ger den forskning som presenteras en ny insikt i hur membrantransport reglerad av Rab8 aktivt bidrar till att ändra cellens form och utseende. Detta kan ge en ny insikt i cellulära processer som cellmigration såväl som en rad sjukdomsförlopp till exempel glaukom, Huntingtons sjukdom och cancer

High-precision mapping of protein–protein interfaces: an integrated genetic strategy combining en masse mutagenesis and DNA-level parallel analysis on a yeast two-hybrid platform

Understanding networks of protein–protein interactions constitutes an essential component on a path towards comprehensive description of cell function. Whereas efficient techniques are readily available for the initial identification of interacting protein partners, practical strategies are lacking for the subsequent high-resolution mapping of regions involved in protein–protein interfaces. We present here a genetic strategy to accurately map interacting protein regions at amino acid precision. The system is based on parallel construction, sampling and analysis of a comprehensive insertion mutant library. The methodology integrates Mu in vitro transposition-based random pentapeptide mutagenesis of proteins, yeast two-hybrid screening and high-resolution genetic footprinting. The strategy is general and applicable to any interacting protein pair. We demonstrate the feasibility of the methodology by mapping the region in human JFC1 that interacts with Rab8A, and we show that the association is mediated by the Slp homology domain 1

Association between the Intrinsically Disordered Protein PEX19 and PEX3

Peer reviewe

A Novel Approach to Detect Malicious User Node by Cognition in Heterogeneous Wireless Networks

Cognitive Networks are characterized by their intelligence and adaptability. Securing layered heterogeneous network architectures has always posed a major challenge to researchers. In this paper, the Observe, Orient, Decide and Act (OODA) loop is adopted to achieve cognition. Intelligence is incorporated by the use of discrete time dynamic neural networks. The use of dynamic neural networks is considered, to monitor the instantaneous changes that occur in heterogeneous network environments when compared to static neural networks. Malicious user node identification is achieved by monitoring the service request rates generated to the cognitive servers. The results and the experimental study presented in this paper prove the improved efficiency in terms of malicious node detection and malicious transaction classification when compared to the existing systems