Katherine Bowers

Protein–Protein Interactions of ESCRT Complexes in the Yeast Saccharomyces cerevisiae

Ten class E Vps proteins in yeast are known components of the ESCRT complexes I, II and III, which are required for the sorting of proteins to the lumenal membranes of multivesicular bodies. We used the yeast 2 hybrid system to analyze the protein–protein interactions of all 17 soluble class E Vps proteins, as well as proteins thought to be required for the ubiquitination and deubiquitination of cargo proteins at multivesicular bodies. We identified novel interactions between yeast ESCRT complex components suggesting that ESCRTI binds to both ESCRTII and ESCRTIII. These interactions were confirmed by GST pull‐down experiments. Our data indicate that the link between ESCRTI and ESCRTIII is via Vps28p and Vps37p/Srn2p binding directly to Vps20p, as well as through indirect interactions via ESCRTII. This is in contrast to the situation in mammalian cells where ESCRTI and ESCRTIII interact indirectly via ALIX, the mammalian homologue of yeast proteins Vps31p/Bro1p and Rim20p. Our data also enable us to link all soluble class E Vps proteins to the ESCRT complexes. We propose the formation of a large multimeric complex on the endosome membrane consisting of ESCRTI, ESCRTII, ESCRTIII and other associated proteins.

Degradation of Endocytosed Epidermal Growth Factor and Virally Ubiquitinated Major Histocompatibility Complex Class I Is Independent of Mammalian ESCRTII

Models for protein sorting at multivesicular bodies in the endocytic pathway of mammalian cells have relied largely on data obtained from yeast. These data suggest the essential role of four ESCRT complexes in multivesicular body protein sorting. However, the putative mammalian ESCRTII complex (hVps25p, hVps22p, and hVps36p) has no proven functional role in endosomal transport. We have characterized the human ESCRTII complex and investigated its function in endosomal trafficking. The human ESCRTII proteins interact with one another, with hVps20p (a component of ESCRTIII), and with their yeast homologues. Our interaction data from yeast two-hybrid studies along with experiments with purified proteins suggest an essential role for the N-terminal domain of hVps22p in the formation of a heterotetrameric ESCRTII complex. Although human ESCRTII is found in the cytoplasm and in the nucleus, it can be recruited to endosomes upon overexpression of dominant-negative hVps4Bp. Interestingly, we find that small interference RNA depletion of mammalian ESCRTII does not affect degradation of epidermal growth factor, a known cargo of the multivesicular body protein sorting pathway. We also show that depletion of the deubiquitinating enzymes AMSH (associated molecule with the SH3 domain of STAM (signal transducing adaptor molecule)) and UBPY (ubiquitin isopeptidase Y) have opposite effects on epidermal growth factor degradation, with UBPY depletion causing dramatic swelling of endosomes. Down-regulation of another cargo, the major histocompatibility complex class I in cells expressing the Kaposi sarcoma-associated herpesvirus protein K3, is unaffected in ESCRTII-depleted cells. Our data suggest that mammalian ESCRTII may be redundant, cargo-specific, or not required for protein sorting at the multivesicular body.

A PC12 variant lacking regulated secretory organelles: aberrant protein targeting and evidence for a factor inhibiting neuroendocrine gene expression.

Abstract: A variant of the PC12 pheochromocytoma cell line (termed A35C) has been isolated that lacks regulated secretory organelles and several constituent proteins. Northern and Southern blot analyses suggested a block at the transcriptional level. The proprotein‐converting enzyme carboxypeptidase H was synthesised in the A35C cell line but was secreted by the constitutive pathway. Transient transfection of A35C cells with cDNAs encoding the regulated secretory proteins dopamine β‐hydroxylase and synaptotagmin I resulted in distinct patterns of mistargeting of these proteins. It is surprising that hybrid cells created by fusing normal PC12 cells with A35C cells exhibited the variant phenotype, suggesting that A35C cells express an inhibitory factor that represses neuroendocrine‐specific gene expression.

A non-canonical ESCRT pathway, including histidine domain phosphotyrosine phosphatase (HD-PTP), is used for down-regulation of virally ubiquitinated MHC class I

Using RNAi, a non-canonical pathway of endosomal sorting complexes required for transport was identified that is responsible for sorting virally ubiquitinated MHC class I into multivesicular bodies (MVBs) during down-regulation of this protein from the cell surface.

The trafficking of metal ion transporters of the Zrt- and Irt-like protein family

Metal ion transporters of the Zrt‐ and Irt‐like protein (ZIP, or SLC39A) family transport zinc, iron, manganese and/or cadmium across cellular membranes and into the cytosol. The 14 human ZIP family proteins are expressed in a wide variety of tissues and function in many different cellular processes. Many of these proteins (including ZIP1, 2, 3, 4, 5, 6/10, 8, 9, 11, 12, 14) are situated, at least some of the time, on the plasma membrane, where they mediate metal ion uptake into cells. Their level on the cell surface can be controlled rapidly via protein trafficking in response to the ions they transport. For example, the cell surface level of many ZIPs (including ZIP1, 3, 4, 8 and 12) is mediated by the available concentration of zinc. Zinc depletion causes a decrease in endocytosis and degradation, resulting in more ZIP on the surface to take up the essential ion. ZIP levels on the cell surface are a balance between endocytosis, recycling and degradation. We review the trafficking mechanisms of human ZIP proteins, highlighting possible targeting motifs and suggesting a model of zinc‐mediated endocytic trafficking. We also provide two possible models for ZIP14 trafficking and degradation.

Information and Empire: Mechanisms of Communication in Russia, 1600-1850

From the mid-sixteenth to the mid-nineteenth century Russia was transformed from a moderate-sized, land-locked principality into the largest empire on earth. How did systems of information and communication shape and reflect this extraordinary change? Information and Mechanisms of Communication in Russia, 1600-1850 brings together a range of contributions to shed some light on this complex question. Communication networks such as the postal service and the gathering and circulation of news are examined alongside the growth of a bureaucratic apparatus that informed the government about its country and its people. The inscription of space is considered from the point of view of mapping and the changing public ‘graphosphere’ of signs and monuments. More than a series of institutional histories, this book is concerned with the way Russia discovered itself, envisioned itself and represented itself to its people. Innovative and scholarly, this collection breaks new ground in its approach to communication and information as a field of study in Russia. More broadly, it is an accessible contribution to pre-modern information studies, taking as its basis a country whose history often serves to challenge habitual Western models of development. It is important reading not only for specialists in Russian Studies, but also for students and non-Russianists who are interested in the history of information and communications.Muscovy and the European Information Revolution : Creating the Mechanisms for Obtaining Foreign News

Haunted Ice, Fearful Sounds, and the Arctic Sublime: Exploring Nineteenth-Century Polar Gothic Space

This article considers a unified polar Gothic as a way of examining texts set in Arctic and Antarctic space. Through analysis of Coleridge’s ‘The Rime of the Ancient Mariner’, Shelley’s Frankenstein, and Poe’s The Narrative of Arthur Gordon Pym of Nantucket, the author creates a framework for understanding polar Gothic, which includes liminal space, the supernatural, the Gothic sublime, ghosts and apparitions, and imperial Gothic anxieties about the degradation of ‘civilisation’. Analysing Verne’s scientific-adventure novel The Adventures of Captain Hatteras (1866) with this framework, the author contextualises the continued public interest in the lost Franklin expedition and reflects on nineteenth-century polar Gothic anxieties in the present day. Polar space creates an uncanny potential for seeing one’s own self and examining what lies beneath the surface of one’s own rational mind.

Russian Writers and the Fin de Siècle: Introduction: The fin-de-siècle mood in Russian literature

Often used to describe the period between 1880 and 1900, the expression “fin de siècle,” French for “end of century,” encompasses a meaning considerably broader than its literal definition. In a conversation toward the end of Oscar Wilde’s novel The Picture of Dorian Gray (1890), two characters, Lord Henry and Lady Narborough, relate the fin de siècle to the end of the world, or fin du globe. “I wish it were fin du globe,” weighs in the perpetually bored Dorian Gray, “Life is a great disappointment.” The conflation of fin de siècle with fin du globe, Dorian’s boredom, his despair at reality’s failure to evoke any hope or inspiration, and the tension between life and a longing for the end of the world come together in this scene to illustrate the fin de siècle’s distinctly broader associations. The concept of the fin de siècle runs over its chronology, embodying an overarching mood that affects both individuals and society as a whole. Contemporary scholar Elaine Showalter argues that we tend to experience the end of a century intensely and emotionally, and ascribe to it “metaphors of death and rebirth.” In Degeneration (1892), Max Nordau defines this emotional state in terms of illness, impotence, and exhaustion, but also an unfulfilled yearning for life and vitality: