bluuebunny/crossref_metadata_2025 huggingface dataset:
Cyclic Expression of Endothelin-converting Enzyme-1 Mediates the Functional Regulation of Seminiferous Tubule Contraction
Authors: Tripiciano Antonella , Peluso Carmelina , Morena Anna Rita , Palombi Fioretta , Stefanini Mario , Ziparo Elio , Yanagisawa Masashi , Filippini Antonio
Published: July 2002

The potent smooth muscle agonist endothelin-1 (ET-1) is involved in the local control of seminiferous tubule contractility, which results in the forward propulsion of tubular fluid and spermatozoa, through its action on peritubular myoid cells. ET-1, known to be produced in the seminiferous epithelium by Sertoli cells, is derived from the inactive intermediate big endothelin-1 (big ET-1) through a specific cleavage operated by the endothelin-converting enzyme (ECE), a membrane-bound metalloprotease with ectoenzymatic activity. The data presented suggest that the timing of seminiferous tubule contractility is controlled locally by the cyclic interplay between different cell types. We have studied the expression of ECE by Sertoli cells and used myoid cell cultures and seminiferous tubule explants to monitor the biological activity of the enzymatic reaction product. Northern blot analysis showed that ECE-1 (and not ECE-2) is specifically expressed in Sertoli cells; competitive enzyme immunoassay of ET production showed that Sertoli cell monolayers are capable of cleaving big ET-1, an activity inhibited by the ECE inhibitor phosphoramidon. Microfluorimetric analysis of intracellular calcium mobilization in single cells showed that myoid cells do not respond to big endothelin, nor to Sertoli cell plain medium, but to the medium conditioned by Sertoli cells in the presence of big ET-1, resulting in cell contraction and desensitization to further ET-1 stimulation; in situ hybridization analysis shows regional differences in ECE expression, suggesting that pulsatile production of endothelin by Sertoli cells (at specific “stages” of the seminiferous epithelium) may regulate the cyclicity of tubular contraction; when viewed in a scanning electron microscope, segments of seminiferous tubules containing the specific stages characterized by high expression of ECE were observed to contract in response to big ET-1, whereas stages with low ECE expression remained virtually unaffected. These data indicate that endothelin-mediated spatiotemporal control of rhythmic tubular contractility might be operated by Sertoli cells through the cyclic expression of ECE-1, which is, in turn, dependent upon the timing of spermatogenesis.
Chs7p, a New Protein Involved in the Control of Protein Export from the Endoplasmic Reticulum that Is Specifically Engaged in the Regulation of Chitin Synthesis in Saccharomyces cerevisiae
Authors: Trilla Jose A. , Durán Angel , Roncero Cesar
Published: July 2002

The Saccharomyces cerevisiae CHS7 gene encodes an integral membrane protein located in the ER which is directly involved in chitin synthesis through the regulation of chitin synthase III (CSIII) activity. In the absence of CHS7 product, Chs3p, but not other secreted proteins, is retained in the ER, leading to a severe defect in CSIII activity and consequently, to a reduced rate of chitin synthesis. In addition, chs7 null mutants show the yeast phenotypes associated with a lack of chitin: reduced mating efficiency and lack of the chitosan ascospore layer, clear indications of Chs7p function throughout the S. cerevisiae biological cycle.

CHS3 overexpression does not lead to increased levels of CSIII because the Chs3p excess is retained in the ER. However, joint overexpression of CHS3 and CHS7 increases the export of Chs3p from the ER and this is accompanied by a concomitant increase in CSIII activity, indicating that the amount of Chs7p is a limiting factor for CSIII activity. Accordingly, CHS7 transcription is increased when elevated amounts of chitin synthesis are detected.

These results show that Chs7p forms part of a new mechanism specifically involved in Chs3p export from the ER and consequently, in the regulation of CSIII activity.
Molecular Characterization of Dendritic Cell-Derived Exosomes
Authors: Théry Clotilde , Regnault Armelle , Garin Jérôme , Wolfers Joseph , Zitvogel Laurence , Ricciardi-Castagnoli Paola , Raposo Graça , Amigorena Sebastian
Published: July 2002

Exosomes are membrane vesicles secreted by hematopoietic cells upon fusion of late multivesicular endosomes with the plasma membrane. Dendritic cell (DC)-derived exosomes induce potent antitumor immune responses in mice, resulting in the regression of established tumors (Zitvogel, L., A. Regnault, A. Lozier, J. Wolfers, C. Flament, D. Tenza, P. Ricciardi-Castagnoli, G. Raposo, and S. Amigorena. 1998. Nat. Med. 4:594–600). To unravel the molecular basis of exosome-induced immune stimulation, we now analyze the regulation of their production during DC maturation and characterize extensively their protein composition by peptide mass mapping. Exosomes contain several cytosolic proteins (including annexin II, heat shock cognate protein hsc73, and heteromeric G protein Gi2α), as well as different integral or peripherally associated membrane proteins (major histocompatiblity complex class II, Mac-1 integrin, CD9, milk fat globule-EGF-factor VIII [MFG-E8]). MFG-E8, the major exosomal component, binds integrins expressed by DCs and macrophages, suggesting that it may be involved in exosome targeting to these professional antigen-presenting cells. Another exosome component is hsc73, a cytosolic heat shock protein (hsp) also present in DC endocytic compartments. hsc73 was shown to induce antitumor immune responses in vivo, and therefore could be involved in the exosome's potent antitumor effects. Finally, exosome production is downregulated upon DC maturation, indicating that in vivo, exosomes are produced by immature DCs in peripheral tissues. Thus, DC-derived exosomes accumulate a defined subset of cellular proteins reflecting their endosomal biogenesis and accounting for their biological function.
The DNA Binding-Independent Function of the Glucocorticoid Receptor Mediates Repression of Ap-1–Dependent Genes in Skin
Authors: Tuckermann Jan P. , Reichardt Holger M. , Arribas Rosa , Richter K. Hartmut , Schütz Günther , Angel Peter
Published: July 2002

The glucocorticoid receptor (GR) mediates the biological effects of glucocorticoids (GCs) through activation or repression of gene expression, either by DNA binding or via interaction with other transcription factors, such as AP-1. Work in tissue culture cells on the regulation of AP-1–dependent genes, such as collagenase (MMP-13) and stromelysin (MMP-3) has suggested that the antitumor and antiinflammatory activity of GCs is mediated, at least in part, by GR-mediated downmodulation of AP-1. Here, we have identified phorbol ester-induced expression of MMP-3 and MMP-13 in mouse skin as the first example of an in vivo system to measure negative interference between AP-1 and GR in the animal. Cell type-specific induction of these genes by tumor promoters is abolished by GCs. Importantly, this is also the case in GRdim mice expressing a DNA binding-defective mutant version of GR. In contrast, the newly identified target genes in skin, plasma glutathione peroxidase and HSP-27, were induced by GC in wild-type, but not in GRdim mice. Thus, these data suggest that the DNA binding-independent function of the GR is dispensable for repression of AP-1 activity in vivo and responsible for the antitumor promoting activity of GCs.
WNT/Frizzled-2 Signaling Induces Aggregation and Adhesion among Cardiac Myocytes by Increased Cadherin–β-Catenin Complex
Authors: Toyofuku Toshihiko , Hong Zhang , Kuzuya Tsunehiko , Tada Michihiko , Hori Masatsugu
Published: July 2002

Wingless is known to be required for induction of cardiac mesoderm in Drosophila, but the function of Wnt family proteins, vertebrate homologues of wingless, in cardiac myocytes remains unknown. When medium conditioned by HEK293 cells overexpressing Wnt-3a or -5a was applied to cultured neonatal cardiac myocytes, Wnt proteins induced myocyte aggregation in the presence of fibroblasts, concomitant with increases in β-catenin and N-cadherin in the myocytes and with E- and M-cadherins in the fibroblasts. The aggregation was inhibited by anti–N-cadherin antibody and induced by constitutively active β-catenin, but was unaffected by dominant negative and dominant positive T cell factor (TCF) mutants. Thus, increased stabilization of complexed cadherin–β-catenin in both cell types appears crucial for the morphological effect of Wnt on cardiac myocytes. Furthermore, myocytes overexpressing a dominant negative frizzled-2, but not a dominant negative frizzled-4, failed to aggregate in response to Wnt, indicating frizzled-2 to be the predominant receptor mediating aggregation. By contrast, analysis of bromodeoxyuridine incorporation and transcription of various cardiogenetic markers showed Wnt to have little or no impact on cell proliferation or differentiation. These findings suggest that a Wnt–frizzled-2 signaling pathway is centrally involved in the morphological arrangement of cardiac myocytes in neonatal heart through stabilization of complexed cadherin– β-catenin.
Adenovirus E4 Open Reading Frame 4–Induced Apoptosis Involves Dysregulation of Src Family Kinases
Authors: Lavoie Josée N. , Champagne Claudia , Gingras Marie-Claude , Robert Amélie
Published: July 2002

The adenoviral early region 4 open reading frame 4 (E4orf4) death factor induces p53-independent apoptosis in many cell types and appears to kill selectively transformed cells. Here we show that expression of E4orf4 in transformed epithelial cells results in early caspase-independent membrane blebbing, associated with changes in the organization of focal adhesions and actin cytoskeleton. Evidence that E4orf4 can associate with and modulate Src family kinase activity, inhibiting Src-dependent phosphorylation of focal adhesion kinase (FAK) and paxillin while increasing phosphorylation of cortactin and some other cellular proteins, is presented. Furthermore, E4orf4 dramatically inhibited the ability of FAK and c-src to cooperate in induction of tyrosine phosphorylation of cellular substrates, suggesting that E4orf4 can interfere with the formation of a signaling complex at focal adhesion sites. Consistent with a functional role for E4orf4–Src interaction, overexpression of activated c-src dramatically potentiated E4orf4-induced membrane blebbing and apoptosis, whereas kinase dead c-src constructs inhibited E4orf4 effects on cell morphology and death. Moreover treatment of E4orf4-expressing cells with PP2, a selective Src kinase inhibitor, led to inhibition of E4orf4-dependent membrane blebbing and later to a marked decrease in E4orf4-induced nuclear condensation. Taken together, these observations indicate that expression of adenovirus 2 E4orf4 can initiate caspase-independent extranuclear manifestations of apoptosis through a modulation of Src family kinases and that these are involved in signaling E4orf4-dependent apoptosis. This study also suggests that Src family kinases are likely to play a role in the cytoplasmic execution of apoptotic programs.
Epimorphin Mediates Mammary Luminal Morphogenesis through Control of C/EBPβ
Authors: Hirai Yohei , Radisky Derek , Boudreau Rosanne , Simian Marina , Stevens Mary E. , Oka Yumiko , Takebe Kyoko , Niwa Shinichiro , Bissell Mina J.
Published: July 2002

We have shown previously that epimorphin (EPM), a protein expressed on the surface of myoepithelial and fibroblast cells of the mammary gland, acts as a multifunctional morphogen of mammary epithelial cells. Here, we present the molecular mechanism by which EPM mediates luminal morphogenesis. Treatment of cells with EPM to induce lumen formation greatly increases the overall expression of transcription factor CCAAT/enhancer binding protein (C/EBP)β and alters the relative expression of its two principal isoforms, LIP and LAP. These alterations were shown to be essential for the morphogenetic activities, since constitutive expression of LIP was sufficient to produce lumen formation, whereas constitutive expression of LAP blocked EPM-mediated luminal morphogenesis. Furthermore, in a transgenic mouse model in which EPM expression was expressed in an apolar fashion on the surface of mammary epithelial cells, we found increased expression of C/EBPβ, increased relative expression of LIP to LAP, and enlarged ductal lumina. Together, our studies demonstrate a role for EPM in luminal morphogenesis through control of C/EBPβ expression.
Condensin and cohesin display different arm conformations with characteristic hinge angles
Authors: Anderson David E. , Losada Ana , Erickson Harold P. , Hirano Tatsuya
Published: July 2002

Structural maintenance of chromosomes (SMC) proteins play central roles in higher-order chromosome dynamics from bacteria to humans. In eukaryotes, two different SMC protein complexes, condensin and cohesin, regulate chromosome condensation and sister chromatid cohesion, respectively. Each of the complexes consists of a heterodimeric pair of SMC subunits and two or three non-SMC subunits. Previous studies have shown that a bacterial SMC homodimer has a symmetrical structure in which two long coiled-coil arms are connected by a flexible hinge. A catalytic domain with DNA- and ATP-binding activities is located at the distal end of each arm. We report here the visualization of vertebrate condensin and cohesin by electron microscopy. Both complexes display the two-armed structure characteristic of SMC proteins, but their conformations are remarkably different. The hinge of condensin is closed and the coiled-coil arms are placed close together. In contrast, the hinge of cohesin is wide open and the coiled-coils are spread apart from each other. The non-SMC subunits of both condensin and cohesin form a globular complex bound to the catalytic domains of the SMC heterodimers. We propose that the “closed” conformation of condensin and the “open” conformation of cohesin are important structural properties that contribute to their specialized biochemical and physiological functions.
An inactive pool of GSK-3 at the leading edge of growth cones is implicated in Semaphorin 3A signaling
Authors: Eickholt Britta J. , Walsh Frank S. , Doherty Patrick
Published: July 2002

Glycogen synthase kinase (GSK)-3 is a serine/threonine kinase that has been implicated in several aspects in embryonic development and several growth factor signaling cascades. We now report that an inactive phosphorylated pool of the enzyme colocalizes with F-actin in both neuronal and nonneuronal cells. Semaphorin 3A (Sema 3A), a molecule that inhibits axonal growth, activates GSK-3 at the leading edge of neuronal growth cones and in Sema 3A–responsive human breast cancer cells, suggesting that GSK-3 activity might play a role in coupling Sema 3A signaling to changes in cell motility. We show that three different GSK-3 antagonists (LiCl, SB-216763, and SB-415286) can inhibit the growth cone collapse response induced by Sema 3A. These studies reveal a novel compartmentalization of inactive GSK-3 in cells and demonstrate for the first time a requirement for GSK-3 activity in the Sema 3A signal transduction pathway.
Role of the 5-Lipoxygenase–activating Protein (FLAP) in Murine Acute Inflammatory Responses
Authors: Byrum Robert S. , Goulet Jennifer L. , Griffiths Richard J. , Koller Beverly H.
Published: July 2002

Leukotrienes are potent inflammatory mediators synthesized from arachidonic acid (AA) predominately by cells of myeloid origin. The synthesis of these lipids is believed to be dependent not only on the expression of the enzyme 5-lipoxygenase (5-LO), which catalyzes the first steps in the synthesis of leukotrienes, but also on expression of a nuclear membrane protein termed the 5-LO–activating protein (FLAP). To study the relationship of these two proteins in mediating the production of leukotrienes in vivo and to determine whether the membrane protein FLAP has additional functions in various inflammatory processes, we have generated a mouse line deficient in this protein. FLAP-deficient mice develop normally and are healthy. However, an array of assays comparing inflammatory reactions in FLAP-deficient mice and in normal controls revealed that FLAP plays a role in a subset of these reactions. Although examination of DTH and IgE-mediated passive anaphylaxis showed no difference between wild-type and FLAP-deficient animals, mice without FLAP possessed a blunted inflammatory response to topical AA and had increased resistance to platelet-activating factor–induced shock compared to controls. Also, edema associated with Zymosan A–induced peritonitis was markedly reduced in animals lacking FLAP. To determine whether these differences relate solely to a deficit in leukotriene production, or whether they reflect an additional role for FLAP in inflammation, we compared the FLAP-deficient mice to 5-LO–deficient animals. Evaluation of mice lacking FLAP and 5-LO indicated that production of leukotrienes during inflammatory responses is dependent upon the availability of FLAP and did not support additional functions for FLAP beyond its role in leukotriene production.
Self-reactive B Cells Are Not Eliminated or Inactivated by Autoantigen Expressed on Thyroid Epithelial Cells
Authors: Akkaraju Srinivas , Canaan Karen , Goodnow Christopher C.
Published: July 2002

Graves' Disease results from the production of autoantibodies against receptors for thyroid stimulating hormone (TSH) on thyroid epithelial cells, and represents the prototype for numerous autoimmune diseases caused by autoantibodies that bind to organ-specific cell membrane antigens. To study how humoral tolerance is normally maintained to organ-specific membrane antigens, transgenic mice were generated selectively expressing membrane-bound hen egg lysozyme (mHEL) on the thyroid epithelium. In contrast to the deletion of autoreactive B cells triggered by systemic mHEL (Hartley, S.B., J. Crosbie, R. Brink, A.B. Kantor, A. Basten, and C.C. Goodnow. 1991. Nature. 353:765–769), selective expression of mHEL autoantigen on thyroid cells did not trigger elimination or inactivation of circulating HEL-reactive B cells. These results provide evidence that tolerance is not actively acquired to organ-specific antigens in the preimmune B cell repertoire, underscoring the importance of maintaining tolerance to such antigens by other mechanisms. The role of an intact endothelial barrier in sequestering organ-specific antigens from circulating preimmune B cells is discussed.
A Study of the Interferon Antiviral Mechanism: Apoptosis Activation by the 2–5A System
Authors: Castelli JoAnn C. , Hassel Bret A. , Wood Katherine A. , Li Xiao-Ling , Amemiya Kei , Dalakas Marinos C. , Torrence Paul F. , Youle Richard J.
Published: July 2002

The 2–5A system contributes to the antiviral effect of interferons through the synthesis of 2–5A and its activation of the ribonuclease, RNase L. RNase L degrades viral and cellular RNA after activation by unique, 2′–5′ phosphodiester-linked, oligoadenylates [2–5A, (pp)p5′ A2′(P5′A2′)]n, n ⩾2. Because both the 2–5A system and apoptosis can serve as viral defense mechanisms and RNA degradation occurs during both processes, we investigated the potential role of RNase L in apoptosis. Overexpression of human RNase L by an inducible promoter in NIH3T3 fibroblasts decreased cell viability and triggered apoptosis. Activation of endogenous RNase L, specifically with 2–5A or with dsRNA, induced apoptosis. Inhibition of RNase L with a dominant negative mutant suppressed poly (I)·poly (C)–induced apoptosis in interferon-primed fibroblasts. Moreover, inhibition of RNase L suppressed apoptosis induced by poliovirus. Thus, increased RNase L levels induced apoptosis and inhibition of RNase L activity blocked viral-induced apoptosis. Apoptosis may be one of the antiviral mechanisms regulated by the 2–5A system.
Phosphorylated Peptides Are Naturally Processed and Presented by Major Histocompatibility Complex Class I Molecules in Vivo
Authors: Zarling Angela L. , Ficarro Scott B. , White Forest M. , Shabanowitz Jeffrey , Hunt Donald F. , Engelhard Victor H.
Published: July 2002

Posttranslational modification of peptide antigens has been shown to alter the ability of T cells to recognize major histocompatibility complex (MHC) class I–restricted peptides. However, the existence and origin of naturally processed phosphorylated peptides presented by MHC class I molecules have not been explored. By using mass spectrometry, significant numbers of naturally processed phosphorylated peptides were detected in association with several human MHC class I molecules. In addition, CD8+ T cells could be generated that specifically recognized a phosphorylated epitope. Thus, phosphorylated peptides are part of the repertoire of antigens available for recognition by T cells in vivo.
Involvement of Lat, Gads, and Grb2 in Compartmentation of Slp-76 to the Plasma Membrane
Authors: Ishiai Masamichi , Kurosaki Mari , Inabe Kazunori , Chan Andrew C. , Sugamura Kazuo , Kurosaki Tomohiro
Published: July 2002

B cell linker protein (BLNK) and Src homology 2 domain–containing leukocyte protein of 76 kD (SLP-76) are adaptor proteins required for B cell receptor (BCR) and T cell receptor function, respectively. Here, we show that expression of SLP-76 cannot reconstitute BCR function in Zap-70+BLNK− B cells. This could be attributable to inability of SLP-76 to be recruited into glycolipid-enriched microdomains (GEMs) after antigen receptor cross-linking. Supporting this idea, the BCR function was restored when a membrane-associated SLP-76 chimera was enforcedly localized to GEMs. Moreover, we demonstrate that addition of both linker for activation of T cells (LAT) and Grb2-related adaptor downstream of Shc (Gads) to SLP-76 allow SLP-76 to be recruited into GEMs, whereby the BCR function is reconstituted. The Gads function was able to be replaced by overexpression of Grb2. In contrast to SLP-76, BLNK did not require Grb2 families for its recruitment to GEMs. Hence, these data suggest a functional overlap between BLNK and SLP-76, while emphasizing the difference in requirement for additional adaptor molecules in their targeting to GEMs.
Impaired c-Jun Amino Terminal Kinase Activity and T Cell Differentiation in Death Receptor 6–deficient Mice
Authors: Zhao Haoran , Yan Minhong , Wang Hua , Erickson Sharon , Grewal Iqbal S. , Dixit Vishva M.
Published: July 2002

During an immune response naive T helper (Th) cells differentiate into two functionally distinct subsets, Th1 and Th2, based on their cytokine secretion profile and immunomodulatory function. c-Jun amino terminal kinase (JNK) regulates Th cell differentiation by activating a transcriptional program required for cytokine production. We have recently identified a TNFR superfamily death domain–containing molecule, death receptor (DR)6, which potently activates JNK. T cells from DR6-deficient mice are substantially impaired in JNK activation. When DR6−/− mice were challenged with protein antigen, their T cells hyperproliferate and display a profound polarization toward a Th2 response whereas Th1 differentiation is not equivalently affected. In addition, DR6−/− T cells showed preference toward Th2 differentiation in vitro. The phenotype seen in the DR6−/− mice is not due to the apoptotic pathway. Therefore, DR6, working through JNK, rather than apoptosis, functions to attenuate the Th2 response. This is the first demonstration of a role in the activation and differentiation of Th cells by DR6 in particular and DRs in general.
Requirement for the NF-κB Family Member RelA in the Development of Secondary Lymphoid Organs
Authors: Alcamo Elizabeth , Hacohen Nir , Schulte Leah C. , Rennert Paul D. , Hynes Richard O. , Baltimore David
Published: July 2002

The transcription factor nuclear factor (NF)-κB has been suggested to be a key mediator of the development of lymph nodes and Peyer's patches. However, targeted deletion of NF-κB/ Rel family members has not yet corroborated such a function. Here we report that when mice lacking the RelA subunit of NF-κB are brought to term by breeding onto a tumor necrosis factor receptor (TNFR)1-deficient background, the mice that are born lack lymph nodes, Peyer's patches, and an organized splenic microarchitecture, and have a profound defect in T cell–dependent antigen responses. Analyses of TNFR1/RelA-deficient embryonic tissues and of radiation chimeras suggest that the dependence on RelA is manifest not in hematopoietic cells but rather in radioresistant stromal cells needed for the development of secondary lymphoid organs.
Regulators of G Protein Signaling Attenuate the G Protein–mediated Inhibition of N-Type Ca Channels
Authors: Melliti Karim , Meza Ulises , Fisher Rory , Adams Brett
Published: July 2002

Regulators of G protein signaling (RGS) proteins bind to the α subunits of certain heterotrimeric G proteins and greatly enhance their rate of GTP hydrolysis, thereby determining the time course of interactions among Gα, Gβγ, and their effectors. Voltage-gated N-type Ca channels mediate neurosecretion, and these Ca channels are powerfully inhibited by G proteins. To determine whether RGS proteins could influence Ca channel function, we recorded the activity of N-type Ca channels coexpressed in human embryonic kidney (HEK293) cells with G protein–coupled muscarinic (m2) receptors and various RGS proteins. Coexpression of full-length RGS3T, RGS3, or RGS8 significantly attenuated the magnitude of receptor-mediated Ca channel inhibition. In control cells expressing α1B, α2, and β3 Ca channel subunits and m2 receptors, carbachol (1 μM) inhibited whole-cell currents by ∼80% compared with only ∼55% inhibition in cells also expressing exogenous RGS protein. A similar effect was produced by expression of the conserved core domain of RGS8. The attenuation of Ca current inhibition resulted primarily from a shift in the steady state dose–response relationship to higher agonist concentrations, with the EC50 for carbachol inhibition being ∼18 nM in control cells vs. ∼150 nM in RGS-expressing cells. The kinetics of Ca channel inhibition were also modified by RGS. Thus, in cells expressing RGS3T, the decay of prepulse facilitation was slower, and recovery of Ca channels from inhibition after agonist removal was faster than in control cells. The effects of RGS proteins on Ca channel modulation can be explained by their ability to act as GTPase-accelerating proteins for some Gα subunits. These results suggest that RGS proteins may play important roles in shaping the magnitude and kinetics of physiological events, such as neurosecretion, that involve G protein–modulated Ca channels.
Electrogenic Na+/Ca2+ Exchange
Authors: Danaceau Jonathan P. , Lucero Mary T.
Published: July 2002

Olfactory receptor neurons (ORNs) from the squid, Lolliguncula brevis, respond to the odors l-glutamate or dopamine with increases in internal Ca2+ concentrations ([Ca2+]i). To directly asses the effects of increasing [Ca2+]i in perforated-patched squid ORNs, we applied 10 mM caffeine to release Ca2+ from internal stores. We observed an inward current response to caffeine. Monovalent cation replacement of Na+ from the external bath solution completely and selectively inhibited the caffeine-induced response, and ruled out the possibility of a Ca2+-dependent nonselective cation current. The strict dependence on internal Ca2+ and external Na+ indicated that the inward current was due to an electrogenic Na+/Ca2+ exchanger. Block of the caffeine-induced current by an inhibitor of Na+/Ca2+ exchange (50–100 μM 2′,4′-dichlorobenzamil) and reversibility of the exchanger current, further confirmed its presence. We tested whether Na+/Ca2+ exchange contributed to odor responses by applying the aquatic odor l-glutamate in the presence and absence of 2′,4′-dichlorobenzamil. We found that electrogenic Na+/Ca2+ exchange was responsible for ∼26% of the total current associated with glutamate-induced odor responses. Although Na+/Ca2+ exchangers are known to be present in ORNs from numerous species, this is the first work to demonstrate amplifying contributions of the exchanger current to odor transduction.
Zeros of the Zak transform on locally compact abelian groups
Authors: Kaniuth Eberhard , Kutyniok Gitta
Published: July 2002

Let

G
G

be a locally compact abelian group. The notion of Zak transform on

L
2

(

R

d

)

L^2(\mathbb {R}^d)

extends to

L
2

(
G
)

L^2(G)

. Suppose that

G
G

is compactly generated and its connected component of the identity is non-compact. Generalizing a classical result for

L
2

(

R

)

L^2(\mathbb {R})

, we then prove that if

f
∈

L
2

(
G
)

f \in L^2(G)

is such that its Zak transform

Z
f

Z f

is continuous on

G
×

G
^

G \times \widehat {G}

, then

Z
f

Z f

has a zero.
On a General Form of the Second Main Theorem
Authors: Ru Min
Published: July 2002

We give a proof of a general form of the Second Main Theorem for holomorphic curves with a good error term. Two applications of this general form are also provided.
Symmetric Gibbs measures
Authors: Petersen Karl , Schmidt Klaus
Published: July 2002

We prove that certain Gibbs measures on subshifts of finite type are nonsingular and ergodic for certain countable equivalence relations, including the orbit relation of the adic transformation (the same as equality after a permutation of finitely many coordinates). The relations we consider are defined by cocycles taking values in groups, including some nonabelian ones. This generalizes (half of) the identification of the invariant ergodic probability measures for the Pascal adic transformation as exactly the Bernoulli measures—a version of de Finetti’s theorem. Generalizing the other half, we characterize the measures on subshifts of finite type that are invariant under both the adic and the shift as the Gibbs measures whose potential functions depend on only a single coordinate. There are connections with and implications for exchangeability, ratio limit theorems for transient Markov chains, interval splitting procedures, ‘canonical’ Gibbs states, and the triviality of remote sigma-fields finer than the usual tail field.
Compact Composition Operators on BMOA
Authors: Bourdon P. , Cima J. , Matheson A.
Published: July 2002

We characterize the compact composition operators on BMOA, the space consisting of those holomorphic functions on the open unit disk

U
U

that are Poisson integrals of functions on

∂
U

\partial U

, that have bounded mean oscillation. We then use our characterization to show that compactness of a composition operator on BMOA implies its compactness on the Hardy spaces (a simple example shows the converse does not hold). We also explore how compactness of the composition operator

C
ϕ

:
BMOA
→
BMOA

C_\phi : \operatorname {BMOA}\rightarrow \operatorname {BMOA}

relates to the shape of

ϕ
(
U
)

\phi (U)

near

∂
U

\partial U

, introducing the notion of mean order of contact. Finally, we discuss the relationships among compactness conditions for composition operators on BMOA, VMOA, and the big and little Bloch spaces.
Normal families: New perspectives
Authors: Zalcman Lawrence
Published: July 2002

This paper surveys some surprising applications of a lemma characterizing normal families of meromorphic functions on plane domains. These include short and efficient proofs of generalizations of (i) the Picard Theorems, (ii) Gol’dberg’s Theorem (a meromorphic function on

C

\mathbb {C}

which is the solution of a first-order algebraic differential equation has finite order), and (iii) the Fatou-Julia Theorem (the Julia set of a rational function of degree

d
≥
2

d\ge 2

is the closure of the repelling periodic points). We also discuss Bloch’s Principle and provide simple solutions to some problems of Hayman connected with this principle.
Residue formulae, vector partition functions and lattice points in rational polytopes
Authors: Brion Michel , Vergne Michèle
Published: July 2002

We obtain residue formulae for certain functions of several variables. As an application, we obtain closed formulae for vector partition functions and for their continuous analogs. They imply an Euler-MacLaurin summation formula for vector partition functions, and for rational convex polytopes as well: we express the sum of values of a polynomial function at all lattice points of a rational convex polytope in terms of the variation of the integral of the function over the deformed polytope.
DOLOP—database of bacterial lipoproteins
Authors: Madan Babu M. , Sankaran K.
Published: July 2002

Summary: Bacterial lipoproteins and lipid modification are gaining importance owing to their essential nature, roles in pathogenesis and interesting commercial applications. We have created an exclusive knowledge base for bacterial lipoproteins by processing information from 510 entries to provide a list of 199 distinct lipoproteins with relevant links to molecular details. Features include functional classification, predictive algorithm for query sequences, primary sequence analysis and lists of predicted lipoproteins from 43 completed bacterial genomes along with interactive information exchange facility.

Availability: The website called Database Of bacterial LipOProteins (DOLOP) is available at http://www.mrc-lmb.cam.ac.uk/genomes/dolop along with additional information on the biosynthetic pathway, supplementary material and other related figures.

Contact: ksankaran@annauniv.edu; madanm@mrc-lmb.cam.ac.uk

* To whom correspondence should be addressed.
CONSUMER THEORY, HOME PRODUCTION, AND ENERGY EFFICIENCY
Authors: Thompson Philip B.
Published: July 2002

Arguments about the existence of the so‐called energy efficiency gap are typically based on evaluations of efficiency measures as investments whose end result is to reduce the cost of providing a given level of energy services. This article demonstrates that the investment approach tends to undervalue energy efficiency improvements because it omits the increase in utility that results from the consumer's substitution of energy services for other goods. The substitution occurs because the efficiency measure reduces the marginal cost (implicit price) of energy services. Sample calculations based on a Cobb‐Douglas utility function are used to illustrate the potential size of the undervaluation of efficiency investments.
Ku‐deficient yeast strains exhibit alternative states of silencing competence
Authors: Maillet Laurent , Gaden Florence , Brevet Vanessa , Fourel Geneviève , Martin Sophie G , Dubrana Karine , Gasser Susan M , Gilson Eric
Published: July 2002

In Saccharomyces cerevisiae, efficient silencer function requires telomere proximity, i.e. compartments of the nucleoplasm enriched in silencing factors. Accordingly, silencers located far from telomeres function inefficiently. We show here that cells lacking yKu balance between two mitotically stable states of silencing competence. In one, a partial delocalization of telomeres and silencing factors throughout the nucleoplasm correlates with enhanced silencing at a non‐telomeric locus, while in the other, telomeres retain their focal pattern of distribution and there is no repression at the non‐telomeric locus, as observed in wild‐type cells. The two states also differ in their level of residual telomeric silencing. These findings indicate the existence of a yKu‐independent pathway of telomere clustering and Sir localization. Interestingly, this pathway appears to be under epigenetic control.
The protein Mago provides a link between splicing and mRNA localization
Authors: Hir Hervé Le , Gatfield David , Braun Isabelle C , Forler Daniel , Izaurralde Elisa
Published: July 2002

The proteins Mago and Y14 are evolutionarily conserved binding partners. Y14 is a component of the exon–exon junction complex (EJC), deposited by the spliceosome upstream of messenger RNA (mRNA) exon–exon junctions. The EJC is implicated in post‐splicing events such as mRNA nuclear export and nonsense‐mediated mRNA decay. Drosophila Mago is essential for the localization of oskar mRNA to the posterior pole of the oocyte, but the functional role of Mago in other species is unknown. We show that Mago is a bona fide component of the EJC. Like Y14, Mago escorts spliced mRNAs to the cytoplasm, providing a direct functional link between splicing and the downstream process of mRNA localization. Mago/Y14 heterodimers are essential in cultured Drosophila cells. Taken together, these results suggest that, in addition to its specialized function in mRNA localization, Mago plays an essential role in other steps of mRNA metabolism.
An intragenic methylated region in the imprinted Igf2 gene augments transcription
Authors: Murrell Adele , Heeson Sarah , Bowden Lucy , Constância Miguel , Dean Wendy , Kelsey Gavin , Reik Wolf
Published: July 2002

DNA methylation is usually associated with transcriptional silencing, but in the imprinted mouse Igf2 gene, the paternally expressed copy is methylated in two discrete differentially methylated regions (DMRs). DMR1 is located upstream of the fetal promoters and has been shown to be a methylation sensitive silencer. Here we examine the role of the intragenic DMR2 by gene targeting. In contrast to DMR1, deletion of DMR2 on the maternal allele did not lead to activation of the silent Igf2 gene. Deletion of a 54 bp methylated core region in DMR2 on the paternal allele, however, reduced Igf2 mRNA levels and was associated with fetal growth retardation. Nuclear run‐on assays showed that the core region influenced transcription initiation, and luciferase reporter assays suggested that its methylation increases transcription. These results reveal a novel mechanism of gene expression whereby intragenic methylation can increase levels of transcription.
Safety and tolerability of quinupristin/dalfopristin: administration guidelines
Authors: Rubinstein E. , Prokocimer P. , Talbot G. H.
Published: July 2002

The safety and tolerability of quinupristin/dalfopristin were assessed in both comparative and non-comparative trials (2298 quinupristin/dalfopristin-treated patients). In comparative clinical trials, the most frequent systemic adverse events related to quinupristin/dalfopristin were nausea (4.6%), diarrhoea (2.7%), vomiting (2.7%) and skin rash (2.5%). The comparator group showed similar rates, except that nausea was significantly more common (7.2%; P = 0.01). In non-comparative trials, arthralgia and myalgia were reported most frequently but were reversible upon treatment discontinuation. The renal, inner ear, cardiovascular and central nervous systems were not implicated as significant target organs for toxicity. The most frequent local adverse events related to infusion of quinupristin/dalfopristin were inflammation, pain, oedema, infusion site reaction and thrombophlebitis. Results of laboratory tests while on therapy were comparable for quinupristin/dalfopristin and comparator groups, except that increases in conjugated bilirubin of >5 × the upper limit of normal were reported in 5.5% of quinupristin/dalfopristin recipients; increases in total bilirubin of >5 × the upper limit of normal occurred in 1.5%. Comparator recipients more frequently had increases in alanine aminotransferase and alkaline phosphatase. Quinupristin/dalfopristin inhibits the cytochrome P450 3A4-mediated metabolism of drugs including midazolam, nifedipine, terfenadine and cyclosporin. Therefore, plasma drug monitoring and/or dosage reduction of these agents is prudent. Concomitant administration of drugs that can prolong the electrocardiographic QTc interval should be avoided. Quinupristin/dalfopristin is visually and chemically compatible with commonly used drugs of various classes, but it is not compatible with sodium chloride solution and certain other drugs, including some antimicrobials. Therefore, when prescribing quinupristin/ dalfopristin, clinicians should be aware of the potential for peripheral venous intolerance, arthralgias and myalgias, increases in conjugated bilirubin, interactions with drugs metabolized by the cytochrome P450 3A4 isoenzyme and certain physico-chemical incompatibilities. However, multiple studies have shown that the safety and tolerability of quinupristin/dalfopristin are generally favourable, and that it provides clear benefits to ill patients with severe Gram-positive infections.
Analysis of the subcellular localization of huntingtin with a set of rabbit polyclonal antibodies in cultured mammalian cells of neuronal origin: comparison with the distribution of huntingtin in Huntington'sdisease autopsy brain
Authors: Dorsman J. C. , Smoor M. A. , Schieman M. L. C. Maat , Bout M. , Siesling S. , Duinen S. G. van , Verschuuren J. J. G. M. , Dunnen J. T. den , Roos R. A. C. , Ommen G. J. B. van
Published: July 2002

Huntington'sdisease (HD) is a neurodegenerative disorder with a midlife onset. The disease is caused by expansion of a CAG (glutamine) repeat within the coding region of the HD gene. The molecular mechanism by which the mutated protein causes this disease is still unclear. To study the protein we have generated a set of rabbit polyclonal antibodies raised against different segments of the N–terminal, central and C–terminal parts of the protein. The polyclonal antibodies were affinity purified and characterized in ELISA and Western blotting experiments. All antibodies can react with the mouse and human proteins. The specificity of these antibodies is underscored by their recognition of huntingtin with different repeat sizes in extracts prepared from patient–derived lymphoblasts. The antibodies were used in immunofluorescence experiments to study the subcellular localization of huntingtin in mouse neuroblastoma N1E–115 cells. The results indicate that most huntingtin is present in the cytoplasm, whereas a minor fraction is present in the nucleus. On differentiation of the N1E–115 cells
in vitro
, the subcellular distribution of huntingtin does not change significantly. These results suggest that full–length huntingtin with a normal repeat length can be detected in the nucleus of cycling and non–cycling cultured mammalian cells of neuronal origin. However, in HD autopsy brain the huntingtin–containing neuronal intranuclear inclusions can be detected only with antibodies raised against the N–terminus of huntingtin. Thus several forms of huntingtin display the propensity for nuclear localization, possibly with different functional consequences.
Mechanisms and regulation of the degradation of cyclin B
Authors: Hershko A.
Published: July 2002

The degradation of the cyclin B subunit of protein kinase Cdk1/cyclin B is required for inactivation of the kinase and exit from mitosis. Cyclin B is degraded by the ubiquitin pathway, a system involved in most selective protein degradation in eukaryotic cells. In this pathway, proteins are targeted for degradation by ligation to ubiquitin, a process carried out by the sequential action of three enzymes: the ubiquitin–activating enzyme E1, a ubiquitin–carrier protein E2 and a ubiquitin–protein ligase E3. In the system responsible for cyclin B degradation, the E3–like function is carried out by a large complex called cyclosome or anaphase–promoting complex (APC). In the early embryonic cell cycles, the cyclosome is inactive in the interphase, but becomes active at the end of mitosis. Activation requires phosphorylation of the cyclosome/APC by protein kinase Cdk1/cyclin B. The lag kinetics of cyclosome activation may be explained by Suc1–assisted multiple phosphorylations of partly phosphorylated complex. The presence of a Fizzy/Cdc20–like protein is necessary for maximal activity of the mitotic form of cyclosome/APC in cyclin–ubiquitin ligation.
A. thaliana TRANSPARENT TESTA 1 is involved in seed coat development and defines the WIP subfamily of plant zinc finger proteins
Authors: Sagasser Martin , Lu Gui-Hua , Hahlbrock Klaus , Weisshaar Bernd
Published: July 2002

Seeds of the Arabidopsis thaliana transparent testa 1 mutant (tt1) appear yellow, due to the lack of condensed tannin pigments in the seed coat. The TT1 gene was isolated by reverse genetics using an En-1 transposon mutagenized A. thaliana population. TT1 gene expression was detected in developing ovules and young seeds only, and the gene was shown to encode a nuclear protein. Mutant seeds displayed altered morphology of the seed endothelium in which brown tannin pigments accumulate in wild-type plants, indicating that TT1 is involved in the differentiation of this cell layer. When overexpressed in transgenicA. thaliana plants, TT1 caused aberrant development and organ morphology. The protein contains a novel combination of two TFIIIA-type zinc finger motifs. Closely related motifs were detected in a number of putative proteins deduced from plant genomic and EST sequences. The new protein domain containing this type of zinc finger motifs was designated WIP, according to three strictly conserved amino acid residues. Our data indicate the existence of a small gene family inA. thaliana which is defined by the occurrence of the WIP domain. WIP genes may play important roles in regulating developmental processes, including the control of endothelium differentiation.
Binding of a fibrinogen mimetic stabilizes integrin αIIbβ3's open conformation
Authors: Hantgan Roy R. , Rocco Mattia , Nagaswami Chandrasekaran , Weisel John W.
Published: July 2002

The platelet integrin αIIbβ3 is representative of a class of heterodimeric receptors that upon activation bind extracellular macromolecular ligands and form signaling clusters. This study examined how occupancy of αIIbβ3's fibrinogen binding site affected the receptor's solution structure and stability. Eptifibatide, an integrin antagonist developed to treat cardiovascular disease, served as a high‐affinity, monovalent model ligand with fibrinogen‐like selectivity for αIIbβ3. Eptifibatide binding promptly and reversibly perturbed the conformation of the αIIbβ3 complex. Ligand‐specific decreases in its diffusion and sedimentation coefficient were observed at near‐stoichiometric eptifibatide concentrations, in contrast to the receptor‐perturbing effects of RGD ligands that we previously observed only at a 70‐fold molar excess. Eptifibatide promoted αIIbβ3 dimerization 10‐fold more effectively than less selective RGD ligands, as determined by sedimentation equilibrium. Eptifibatide‐bound integrin receptors displayed an ectodomain separation and enhanced assembly of dimers and larger oligomers linked through their stalk regions, as seen by transmission electron microscopy. Ligation with eptifibatide protected αIIbβ3 from SDS‐induced subunit dissociation, an effect on electrophoretic mobility not seen with RGD ligands. Despite its distinct cleft, the open conformer resisted guanidine unfolding as effectively as the ligand‐free integrin. Thus, we provide the first demonstration that binding a monovalent ligand to αIIbβ3's extracellular fibrinogen‐recognition site stabilizes the receptor's open conformation and enhances self‐association through its distant transmembrane and/or cytoplasmic domains. By showing how eptifibatide and RGD peptides, ligands with distinct binding sites, each affects αIIbβ3's conformation, our findings provide new mechanistic insights into ligand‐linked integrin activation, clustering and signaling.