bluuebunny/crossref_metadata_2025 huggingface dataset:
Complex interactions between the laminin α4 subunit and integrins regulate endothelial cell behavior
in vitro
and angiogenesis
in vivo
Authors: Gonzalez Annette M. , Gonzales Meredith , Herron G. Scott , Nagavarapu Usha , Hopkinson Susan B. , Tsuruta Daisuke , Jones Jonathan C. R.
Published: December 2002

The α4 laminin subunit is a component of the basement membrane of blood vessels where it codistributes with the integrins αvβ3, α3β1, and α6β1. An antibody against the G domain (residues 919-1207; G
919–1207
) of the α4 laminin subunit inhibits angiogenesis in a mouse–human chimeric model, indicating the functional importance of this domain. Additional support for the latter derives from the ability of recombinant G
919–1207
to support endothelial cell adhesion. In particular, endothelial cell adhesion to G
919–1207
is half-maximal at 1.4 nM, whereas residues 919-1018 and 1016–1207 of the G domain are poor cellular ligands. Function blocking antibodies against integrins αvβ3 and β1 and a combination of antibodies against α3 and α6 integrin subunits inhibit endothelial cell attachment to G
919–1207
. Moreover, both αvβ3 and α3β1 integrin bind with high affinity to G
919–1207
. Together, our studies demonstrate that the G domain of laminin α4 chain is a specific, high affinity ligand for the αvβ3 and α3β1 integrin heterodimers and that these integrins, together with α6β1, function cooperatively to mediate endothelial cell–α4 laminin interaction and hence blood vessel development. We propose a model based on these data that reconcile apparent discrepancies in the recent literature with regard to the role of the αvβ3 integrin in angiogenesis.
Changes in the middle region of Sup35 profoundly alter the nature of epigenetic inheritance for the yeast prion [
PSI
+
]
Authors: Liu Jia-Jia , Sondheimer Neal , Lindquist Susan L.
Published: December 2002

The yeast prion [
PSI
+
] provides an epigenetic mechanism for the inheritance of new phenotypes through self-perpetuating changes in protein conformation. [
PSI
+
] is a nonfunctional, ordered aggregate of the translation termination factor Sup35p that influences new Sup35 proteins to adopt the same state. The N-terminal region of Sup35p plays a central role in prion induction and propagation. The C-terminal region provides translation termination activity. The function of the highly charged, conformationally flexible middle region (M) is unknown. An M deletion mutant was capable of existing in either the prion or the nonprion state, but in either case it was mostly insoluble. Substituting a charged synthetic polypeptide for M restored solubility, but the prions formed by this variant were mitotically very unstable. Substituting charged flexible regions from two other proteins for M created variants that acquired prion states (defined as self-perpetuating changes in function transferred to them from wild-type [
PSI
+
] elements), but had profoundly different properties. One was soluble in both the prion and the nonprion form, mitotically stable but meiotically unstable, and cured by guanidine HCl but not by alterations in heat shock protein 104 (Hsp104p). The other could only maintain the prion state in the presence of wild-type protein, producing Mendelian segregation patterns. The unique character of these M variants, all carrying the same N-terminal prion-determining region, demonstrate the importance of M for [
PSI
+
] and suggest that a much wider range of epigenetic phenomena might be based on self-perpetuating, prion-like changes in protein conformation than suggested by our current methods for defining prion states.
Nonlymphoid Fas ligand in peptide-induced peripheral lymphocyte deletion
Authors: Pinkoski Michael J. , Droin Nathalie M. , Lin Tesu , Genestier Laurent , Ferguson Thomas A. , Green Douglas R.
Published: December 2002

Peripheral lymphocyte deletion is required for reduction of lymphocyte numbers after expansion in response to antigen. Peripheral deletion is mediated in part by the activation of apoptosis by engagement of the death receptor, Fas (CD95), by its ligand, Fas ligand (FasL; CD95L), among other mechanisms. Here we used T cell receptor (TCR) transgenic animals to examine the role of inducible expression of nonlymphoid FasL in response to peptide antigen. Antigenic challenge of TCR transgenic mice resulted in increased expression of FasL in a number of nonlymphoid tissues including the epithelium of the small intestine. Similar results were obtained in an adoptive transfer system in which TCR transgenic T cells were transferred into recipient animals. The functional relevance of nonlymphoid FasL in peripheral deletion is supported by the observation that FasL-deficientgldanimals showed a significantly reduced rate of clearance of transferred antigen-specific lymphocytes, although the lymphocytes themselves were wild type for FasL. These observations were supported further by studies in a transgenic mouse model wherelacZwas expressed under the control of the proximal promoter of theFasLgene. Using these transgenic mice, we observed induced activity of theFasLpromoter in intestinal epithelial cells throughout the crypts and villi, where we also observed infiltration of activated T cells. These data demonstrate that nonlymphoid FasL is expressed in response to peripheral T cell activation and participates in the regulation of T cells that infiltrate peripheral tissues.
The intranuclear mobility of messenger RNA binding proteins is ATP dependent and temperature sensitive
Authors: Calapez Alexandre , Pereira Henrique M. , Calado Angelo , Braga José , Rino José , Carvalho Célia , Tavanez João Paulo , Wahle Elmar , Rosa Agostinho C. , Carmo-Fonseca Maria
Published: December 2002

fAter being released from transcription sites, messenger ribonucleoprotein particles (mRNPs) must reach the nuclear pore complexes in order to be translocated to the cytoplasm. Whether the intranuclear movement of mRNPs results largely from Brownian motion or involves molecular motors remains unknown. Here we have used quantitative photobleaching techniques to monitor the intranuclear mobility of protein components of mRNPs tagged with GFP. The results show that the diffusion coefficients of the poly(A)-binding protein II (PABP2) and the export factor TAP are significantly reduced when these proteins are bound to mRNP complexes, as compared with nonbound proteins. The data further show that the mobility of wild-type PABP2 and TAP, but not of a point mutant variant of PABP2 that fails to bind to RNA, is significantly reduced when cells are ATP depleted or incubated at 22°C. Energy depletion has only minor effects on the intranuclear mobility of a 2,000-kD dextran (which corresponds approximately in size to 40S mRNP particles), suggesting that the reduced mobility of PABP2 and TAP is not caused by a general alteration of the nuclear environment. Taken together, the data suggest that the mobility of mRNPs in the living cell nucleus involves a combination of passive diffusion and ATP-dependent processes.
The yeast nuclear pore complex functionally interacts with components of the spindle assembly checkpoint
Authors: Iouk Tatiana , Kerscher Oliver , Scott Robert J. , Basrai Munira A. , Wozniak Richard W.
Published: December 2002

Aphysical and functional link between the nuclear pore complex (NPC) and the spindle checkpoint machinery has been established in the yeast Saccharomyces cerevisiae. We show that two proteins required for the execution of the spindle checkpoint, Mad1p and Mad2p, reside predominantly at the NPC throughout the cell cycle. There they are associated with a subcomplex of nucleoporins containing Nup53p, Nup170p, and Nup157p. The association of the Mad1p–Mad2p complex with the NPC requires Mad1p and is mediated in part by Nup53p. On activation of the spindle checkpoint, we detect changes in the interactions between these proteins, including the release of Mad2p (but not Mad1p) from the NPC and the accumulation of Mad2p at kinetochores. Accompanying these events is the Nup53p-dependent hyperphosphorylation of Mad1p. On the basis of these results and genetic analysis of double mutants, we propose a model in which Mad1p bound to a Nup53p-containing complex sequesters Mad2p at the NPC until its release by activation of the spindle checkpoint. Furthermore, we show that the association of Mad1p with the NPC is not passive and that it plays a role in nuclear transport.
Recruitment of the Arp2/3 complex to vinculin
Authors: DeMali Kris A. , Barlow Christy A. , Burridge Keith
Published: December 2002

Cell migration involves many steps, including membrane protrusion and the development of new adhesions. Here we have investigated whether there is a link between actin polymerization and integrin engagement. In response to signals that trigger membrane protrusion, the actin-related protein (Arp)2/3 complex transiently binds to vinculin, an integrin-associated protein. The interaction is regulated, requiring phosphatidylinositol-4,5-bisphosphate and Rac1 activation, and is sufficient to recruit the Arp2/3 complex to new sites of integrin aggregation. Binding of the Arp2/3 complex to vinculin is direct and does not depend on the ability of vinculin to associate with actin. We have mapped the binding site for the Arp2/3 complex to the hinge region of vinculin, and a point mutation in this region selectively blocks binding to the Arp2/3 complex. Compared with WT vinculin, expression of this mutant in vinculin-null cells results in diminished lamellipodial protrusion and spreading on fibronectin. The recruitment of the Arp2/3 complex to vinculin may be one mechanism through which actin polymerization and membrane protrusion are coupled to integrin-mediated adhesion.
CENP-B box is required for de novo centromere chromatin assembly on human alphoid DNA
Authors: Ohzeki Jun-ichirou , Nakano Megumi , Okada Teruaki , Masumoto Hiroshi
Published: December 2002

Centromere protein (CENP) B boxes, recognition sequences of CENP-B, appear at regular intervals in human centromeric α-satellite DNA (alphoid DNA). In this study, to determine whether information carried by the primary sequence of alphoid DNA is involved in assembly of functional human centromeres, we created four kinds of synthetic repetitive sequences: modified alphoid DNA with point mutations in all CENP-B boxes, resulting in loss of all CENP-B binding activity; unmodified alphoid DNA containing functional CENP-B boxes; and nonalphoid repetitive DNA sequences with or without functional CENP-B boxes. These four synthetic repetitive DNAs were introduced into cultured human cells (HT1080), and de novo centromere assembly was assessed using the mammalian artificial chromosome (MAC) formation assay. We found that both the CENP-B box and the alphoid DNA sequence are required for de novo MAC formation and assembly of functional centromere components such as CENP-A, CENP-C, and CENP-E. Using the chromatin immunoprecipitation assay, we found that direct assembly of CENP-A and CENP-B in cells with synthetic alphoid DNA required functional CENP-B boxes. To the best of our knowledge, this is the first reported evidence of a functional molecular link between a centromere-specific DNA sequence and centromeric chromatin assembly in humans.
VCIP135, a novel essential factor for p97/p47-mediated membrane fusion, is required for Golgi and ER assembly in vivo
Authors: Uchiyama Keiji , Jokitalo Eija , Kano Fumi , Murata Masayuki , Zhang Xiaodong , Canas Benito , Newman Richard , Rabouille Catherine , Pappin Darryl , Freemont Paul , Kondo Hisao
Published: December 2002

NSF and p97 are ATPases required for the heterotypic fusion of transport vesicles with their target membranes and the homotypic fusion of organelles. NSF uses ATP hydrolysis to dissociate NSF/SNAPs/SNAREs complexes, separating the v- and t-SNAREs, which are then primed for subsequent rounds of fusion. In contrast, p97 does not dissociate the p97/p47/SNARE complex even in the presence of ATP. Now we have identified a novel essential factor for p97/p47-mediated membrane fusion, named VCIP135 (valosin-containing protein [VCP][p97]/p47 complex-interacting protein, p135), and show that it binds to the p97/p47/syntaxin5 complex and dissociates it via p97 catalyzed ATP hydrolysis. In living cells, VCIP135 and p47 are shown to function in Golgi and ER assembly.
A novel Apaf-1–independent putative caspase-2 activation complex
Authors: Read Stuart H. , Baliga Belinda C. , Ekert Paul G. , Vaux David L. , Kumar Sharad
Published: December 2002

CVaspase activation is a key event in apoptosis execution. In stress-induced apoptosis, the mitochondrial pathway of caspase activation is believed to be of central importance. In this pathway, cytochrome c released from mitochondria facilitates the formation of an Apaf-1 apoptosome that recruits and activates caspase-9. Recent data indicate that in some cells caspase-9 may not be the initiator caspase in stress-mediated apoptosis because caspase-2 is required upstream of mitochondria for the release of cytochrome c and other apoptogenic factors. To determine how caspase-2 is activated, we have studied the formation of a complex that mediates caspase-2 activation. Using gel filtration analysis of cell lysates, we show that caspase-2 is spontaneously recruited to a large protein complex independent of cytochrome c and Apaf-1 and that recruitment of caspase-2 to this complex is sufficient to mediate its activation. Using substrate-binding assays, we also provide the first evidence that caspase-2 activation may occur without processing of the precursor molecule. Our data are consistent with a model where caspase-2 activation occurs by oligomerization, independent of the Apaf-1 apoptosome.
Lineage-specific requirements of β-catenin in neural crest development
Authors: Hari Lisette , Brault Véronique , Kléber Maurice , Lee Hye-Youn , Ille Fabian , Leimeroth Rainer , Paratore Christian , Suter Ueli , Kemler Rolf , Sommer Lukas
Published: December 2002

β-Catenin plays a pivotal role in cadherin-mediated cell adhesion. Moreover, it is a downstream signaling component of Wnt that controls multiple developmental processes such as cell proliferation, apoptosis, and fate decisions. To study the role of β-catenin in neural crest development, we used the Cre/loxP system to ablate β-catenin specifically in neural crest stem cells. Although several neural crest–derived structures develop normally, mutant animals lack melanocytes and dorsal root ganglia (DRG). In vivo and in vitro analyses revealed that mutant neural crest cells emigrate but fail to generate an early wave of sensory neurogenesis that is normally marked by the transcription factor neurogenin (ngn) 2. This indicates a role of β-catenin in premigratory or early migratory neural crest and points to heterogeneity of neural crest cells at the earliest stages of crest development. In addition, migratory neural crest cells lateral to the neural tube do not aggregate to form DRG and are unable to produce a later wave of sensory neurogenesis usually marked by the transcription factor ngn1. We propose that the requirement of β-catenin for the specification of melanocytes and sensory neuronal lineages reflects roles of β-catenin both in Wnt signaling and in mediating cell–cell interactions.
Electronic Structures of the Filled Tetrahedral Semiconductor
LiMgN with a Zincblende-Type Structure
Authors: Li Hui-Ping  , Hou Zhu-Feng  , Huang Mei-Chun  , Zhu Zi-Zhong
Published: December 2002

An ab initio method with mixed-basis norm-conserving non-local
pseudo-potentials has been employed to investigate the electronic
structures of LiMgN. The band structure, electronic density of states
and charge density contour plot of LiMgN are also presented. By the
calculation, we have found that LiMgN with a zincblende-type structure
was an indirect gap semiconductor, and the value of indirect
(Γ-X) energy band gap under the local density approximation was
2.97 eV. In addition, the strong covalent character for Li-N and Mg-N
has also been found in LiMgN.
Regulation of IκBβ Expression in Testis
Authors: Budde Lucy M. , Wu Chun , Tilman Christopher , Douglas Iris , Ghosh Sankar
Published: December 2002

IκBα and IκBβ are regulators of the nuclear factor-κB (NF-κB) transcription factor family. Both IκBs bind to the same NF-κB dimers and are widely expressed in different cells and tissues. To better understand how these two IκB isoforms differ biologically, we have characterized the expression of IκBβ in testis, a tissue in which IκBα is only minimally expressed. We have found that IκBβ expression is localized within the haploid spermatid stages of spermatogenesis and follows the expression of nuclear NF-κB. IκBβ expression in haploid spermatids is likely regulated by Sox family proteins, members of which are also expressed within spermatids. We have shown that both SRY and Sox-5 can bind to multiple Sox binding sites found within the IκBβ promoter and can enhance transcription of a reporter gene in transient transfection assays. We also demonstrate that IκBβ mRNA is strongly expressed in developing male gonads. These results therefore suggest that IκBβ may be a novel target for transcription factors of the HMG-box SRY/Sox family and imply a potential role for NF-κB/IκBβ in spermatogenesis.
Kar9p-independent Microtubule Capture at Bud6p Cortical Sites Primes Spindle Polarity before Bud Emergence inSaccharomyces cerevisiae
Authors: Segal Marisa , Bloom Kerry , Reed Steven I.
Published: December 2002

Spindle orientation is critical for accurate chromosomal segregation in eukaryotic cells. In the yeast Saccharomyces cerevisiae, orientation of the mitotic spindle is achieved by a program of microtubule–cortex interactions coupled to spindle morphogenesis. We previously implicated Bud6p in directing microtubule capture throughout this program. Herein, we have analyzed cells coexpressing GFP:Bud6 and GFP:Tub1 fusions, providing a kinetic view of Bud6p–microtubule interactions in live cells. Surprisingly, even during the G1 phase, microtubule capture at the recent division site and the incipient bud is dictated by Bud6p. These contacts are eliminated in bud6Δ cells but are proficient inkar9Δ cells. Thus, Bud6p cues microtubule capture, as soon as a new cell polarity axis is established independent of Kar9p. Bud6p increases the duration of interactions and promotes distinct modes of cortical association within the bud and neck regions. In particular, microtubule shrinkage and growth at the cortex rarely occur away from Bud6p sites. These are the interactions selectively impaired at the bud cortex in bud6Δ cells. Finally, interactions away from Bud6p sites within the bud differ from those occurring at the mother cell cortex, pointing to the existence of an independent factor controlling cortical contacts in mother cells after bud emergence.
Mutant Actins Demonstrate a Role for Unpolymerized Actin in Control of Transcription by Serum Response Factor
Authors: Posern Guido , Sotiropoulos Athanassia , Treisman Richard
Published: December 2002

Signal-induced activation of the transcription factor serum response factor (SRF) requires alterations in actin dynamics. SRF activity can be inhibited by ectopic expression of β-actin, either because actin itself participates in SRF regulation or as a consequence of cytoskeletal perturbations. To distinguish between these possibilities, we studied actin mutants. Three mutant actins, G13R, R62D, and a C-terminal VP16 fusion protein, were shown not to polymerize in vivo, as judged by two-hybrid, immunofluorescence, and cell fractionation studies. These actins effectively inhibited SRF activation, as did wild-type actin, which increased the G-actin level without altering the F:G-actin ratio. Physical interaction between SRF and actin was not detectable by mammalian or yeast two-hybrid assays, suggesting that SRF regulation involves an unidentified cofactor. SRF activity was not blocked upon inhibition of CRM1-mediated nuclear export by leptomycin B. Two actin mutants were identified, V159N and S14C, whose expression favored F-actin formation and which strongly activated SRF in the absence of external signals. These mutants seemed unable to inhibit SRF activity, because their expression did not reduce the absolute level of G-actin as assessed by DNase I binding. Taken together, these results provide strong evidence that G-actin, or a subpopulation of it, plays a direct role in signal transduction to SRF.
Selective Protein Exit from Yeast Endoplasmic Reticulum in Absence of Functional COPII Coat Component Sec13p
Authors: Fatal Netta , Suntio Taina , Makarow Marja
Published: December 2002

Sec13p has been thought to be an essential component of the COPII coat, required for exit of proteins from the yeast endoplasmic reticulum (ER). We show herein that normal function of Sec13p was not required for ER exit of the Hsp150 glycoprotein. Hsp150 was secreted to the medium under restrictive conditions in a sec13-1mutant. The COPII components Sec23p and Sec31p and the GTP/GDP exchange factor Sec12p were required in functional form for secretion of Hsp150. Hsp150 leaves the ER in the absence of retrograde COPI traffic, and the responsible determinant is a peptide repeated 11 times in the middle of the Hsp150 sequence. Herein, we localized the sorting determinant for Sec13p-independent ER exit to the C-terminal domain. Sec13p-dependent invertase left the ER in the absence of normal Sec13p function, when fused to the C-terminal domain of Hsp150, demonstrating that this domain contained an active mediator of Sec13p-independent secretion. Thus, Hsp150 harbors two different signatures that regulate its ER exit. Our data show that transport vesicles lacking functional Sec13p can carry out ER-to-Golgi transport, but select only specific cargo protein(s) for ER exit.
Characterization of Human Alpha-Dystrobrevin Isoforms in HL-60 Human Promyelocytic Leukemia Cells Undergoing Granulocytic Differentiation
Authors: Kulyte Agné , Navakauskiene Ruta , Treigyte Grazina , Gineitis Arunas , Bergman Tomas , Magnusson Karl-Eric
Published: December 2002

The biochemical properties and spatial localization of the protein alpha-dystrobrevin and other isoforms were investigated in cells of the human promyelocytic leukemia line HL-60 granulocytic differentiation as induced by retinoic acid (RA). Alpha-dystrobrevin was detected both in the cytosol and the nuclei of these cells, and a short isoform (gamma-dystrobrevin) was modified by tyrosine phosphorylation soon after the onset of the RA-triggered differentiation. Varying patterns of distribution of alpha-dystrobrevin and its isoforms could be discerned in HL-60 promyelocytes, RA-differentiated mature granulocytes, and human neutrophils. Moreover, the gamma-dystrobrevin isoform was found in association with actin and myosin light chain. The results provide new information about potential involvement of alpha-dystrobrevin and its splice isoforms in signal transduction in myeloid cells during induction of granulocytic differentiation and/or at the commitment stage of differentiation or phagocytic cells.
Alternative Splicing Regulates the Subcellular Localization of Divalent Metal Transporter 1 Isoforms
Authors: Tabuchi Mitsuaki , Tanaka Naotaka , Nishida-Kitayama Junko , Ohno Hiroshi , Kishi Fumio
Published: December 2002

Divalent metal transporter 1 (DMT1) is responsible for dietary-iron absorption from apical plasma membrane in the duodenum and iron acquisition from the transferrin cycle endosomes in peripheral tissues. Two isoforms of the DMT1 transcript generated by alternative splicing of the 3′ exons have been identified in mouse, rat, and human. These isoforms can be distinguished by the different C-terminal amino acid sequences and by the presence (DMT1A) or absence (DMT1B) of an iron response element located in the 3′ untranslated region of the mRNA. However, it has been still unknown whether the structural differences between the two DMT1 isoforms is functionally important. Here, we report that each DMT1 isoform exhibits a differential cell type–specific expression patterns and distinct subcellular localizations. DMT1A is predominantly expressed by epithelial cell lines, whereas DMT1B is expressed by the blood cell lines. In HEp-2 cells, GFP-tagged DMT1A is localized in late endosomes and lysosomes, whereas GFP-tagged DMT1B is localized in early endosomes. Using site-directed mutagenesis, a Y555XLXX sequence in the cytoplasmic tail of DMT1B has been identified as an important signal sequence for the early endosomal-targeting of DMT1B. In polarized MDCK cells, GFP-tagged DMT1A and DMT1B are localized in the apical plasma membrane and their respective specific endosomes. Disruption of the N-glycosylation sites in each of the DMT1 isoforms affects their polarized distribution into the apical plasma membrane but not their correct endosomal localization. Our data indicate that the cell type–specific expression patterns and the distinct subcellular localizations of two DMT1 isoforms may be involved in the different iron acquisition steps from the subcellular membranes in various cell types.
Adaptor Protein Crk Is Required for Ephrin-B1-induced Membrane Ruffling and Focal Complex Assembly of Human Aortic Endothelial Cells
Authors: Nagashima Ken-Ichiro , Endo Akira , Ogita Hisakazu , Kawana Akiko , Yamagishi Akiko , Kitabatake Akira , Matsuda Michiyuki , Mochizuki Naoki
Published: December 2002

Endothelial cell migration is an essential step in vasculogenesis and angiogenesis, in which receptor tyrosine kinases play a pivotal role. We investigated the mechanism by which ephrin-B1 promotes membrane ruffling in human aortic endothelial cells, because membrane ruffling heralds cell body migration. We especially focused on the role of Crk adaptor protein in EphB-mediated signaling. Using DsRed-tagged Crk and a fluorescent time-lapse microscope, we showed that Crk was recruited to the nascent focal complex after ephrin-B1 stimulation. Furthermore, we found that p130Cas, but not paxillin, recruited Crk to the nascent focal complex. The necessity of Crk in ephrin-B1–induced membrane ruffling was shown both by the overexpression of dominant negative Crk mutants and by the depletion of Crk by using RNA interference. Then, we examined the role of two major downstream molecules of Crk, Rac1 and Rap1. The dominant negative mutant of Rac1 completely inhibited ephrin-B1–induced membrane ruffling and focal complex assembly. In contrast, rap1GAPII, a negative regulator of Rap1, did not inhibit ephrin-B1–induced membrane ruffling. However, in rap1GAPII-expressing cells, ephrin-B1 did not induce membrane spreading, probably due to instability of the focal complex. These results indicated that Crk plays a critical role in Rac1-induced membrane ruffling and Rap1-mediated nascent focal complex stabilization contributing to ephrin-B1–induced human aortic endothelial cells migration.
Lipid-dependent Subcellular Relocalization of the Acyl Chain Desaturase in Yeast
Authors: Tatzer Verena , Zellnig Günther , Kohlwein Sepp D. , Schneiter Roger
Published: December 2002

The degree of acyl chain desaturation of membrane lipids is a critical determinant of membrane fluidity. Temperature-sensitive mutants of the single essential acyl chain desaturase, Ole1p, of yeast have previously been isolated in screens for mitochondrial inheritance mutants ( Stewart, L.C., and Yaffe, M.P. (1991). J. Cell Biol.115, 1249–1257 ). We now report that the mutant desaturase relocalizes from its uniform ER distribution to a more punctuate localization at the cell periphery upon inactivation of the enzyme. This relocalization takes place within minutes at nonpermissive conditions, a time scale at which mitochondrial morphology and inheritance is not yet affected. Relocalization of the desaturase is fully reversible and does not affect the steady state localization of other ER resident proteins or the kinetic and fidelity of the secretory pathway, indicating a high degree of selectivity for the desaturase. Relocalization of the desaturase is energy independent but is lipid dependent because it is rescued by supplementation with unsaturated fatty acids. Relocalization of the desaturase is also observed in cells treated with inhibitors of the enzyme, indicating that it is independent of temperature-induced alterations of the enzyme. In the absence of desaturase function, lipid synthesis continues, resulting in the generation of lipids with saturated acyl chains. A model is discussed in which the accumulation of saturated lipids in a microdomain around the desaturase could induce the observed segregation and relocalization of the enzyme.
Different Transmembrane Domains Associate with Distinct Endoplasmic Reticulum Components during Membrane Integration of a Polytopic Protein
Authors: Meacock Suzanna L. , Lecomte Fabienne J.L. , Crawshaw Samuel G. , High Stephen
Published: December 2002

We have been studying the insertion of the seven transmembrane domain (TM) protein opsin to gain insights into how the multiple TMs of polytopic proteins are integrated at the endoplasmic reticulum (ER). We find that the ER components associated with the first and second TMs of the nascent opsin polypeptide chain are clearly distinct. The first TM (TM1) is adjacent to the α and β subunits of the Sec61 complex, and a novel component, a protein associated with the ER translocon of 10 kDa (PAT-10). The most striking characteristic of PAT-10 is that it remains adjacent to TM1 throughout the biogenesis and membrane integration of the full-length opsin polypeptide. TM2 is also found to be adjacent to Sec61α and Sec61β during its membrane integration. However, TM2 does not form any adducts with PAT-10; rather, a transient association with the TRAM protein is observed. We show that the association of PAT-10 with opsin TM1 does not require theN-glycosylation of the nascent chain and occurs irrespective of the amino acid sequence and transmembrane topology of TM1. We conclude that the precise makeup of the ER membrane insertion site can be distinct for the different transmembrane domains of a polytopic protein. We find that the environment of a particular TM can be influenced by both the “stage” of nascent chain biosynthesis reached, and the TM's relative location within the polypeptide.
Novel Myosin Heavy Chain Kinase Involved in Disassembly of Myosin II Filaments and Efficient Cleavage in MitoticDictyosteliumCells
Authors: Nagasaki Akira , Itoh Go , Yumura Shigehiko , Uyeda Taro Q.P.
Published: December 2002

We have cloned a full-length cDNA encoding a novel myosin II heavy chain kinase (mhckC) from Dictyostelium. Like other members of the myosin heavy chain kinase family, themhckC gene product, MHCK C, has a kinase domain in its N-terminal half and six WD repeats in the C-terminal half. GFP-MHCK C fusion protein localized to the cortex of interphase cells, to the cleavage furrow of mitotic cells, and to the posterior of migrating cells. These distributions of GFP-MHCK C always corresponded with that of myosin II filaments and were not observed in myosin II-null cells, where GFP-MHCK C was diffusely distributed in the cytoplasm. Thus, localization of MHCK C seems to be myosin II-dependent. Cells lacking the mhckC gene exhibited excessive aggregation of myosin II filaments in the cleavage furrows and in the posteriors of the daughter cells once cleavage was complete. The cleavage process of these cells took longer than that of wild-type cells. Taken together, these findings suggest MHCK C drives the disassembly of myosin II filaments for efficient cytokinesis and recycling of myosin II that occurs during cytokinesis.
Microtubule Asymmetry during Neutrophil Polarization and Migration
Authors: Eddy Robert J. , Pierini Lynda M. , Maxfield Frederick R.
Published: December 2002

The development of cell polarity in response to chemoattractant stimulation in human polymorphonuclear neutrophils (PMNs) is characterized by the rapid conversion from round to polarized morphology with a leading lamellipod at the front and a uropod at the rear. During PMN polarization, the microtubule (MT) array undergoes a dramatic reorientation toward the uropod that is maintained during motility and does not require large-scale MT disassembly or cell adhesion to the substratum. MTs are excluded from the leading lamella during polarization and motility, but treatment with a myosin light chain kinase inhibitor (ML-7) or the actin-disrupting drug cytochalasin D causes an expansion of the MT array and penetration of MTs into the lamellipod. Depolymerization of the MT array before stimulation caused 10% of the cells to lose their polarity by extending two opposing lateral lamellipodia. These multipolar cells showed altered localization of a leading lamella-specific marker, talin, and a uropod-specific marker, CD44. In summary, these results indicate that F-actin– and myosin II-dependent forces lead to the development and maintenance of MT asymmetry that may act to reinforce cell polarity during PMN migration.
A Novel Transforming Growth Factor-β Receptor-interacting Protein That Is Also a Light Chain of the Motor Protein Dynein
Authors: Tang Qian , Staub Cory M. , Gao Guofeng , Jin Qunyan , Wang Zhengke , Ding Wei , Aurigemma Rosemarie E. , Mulder Kathleen M.
Published: December 2002

The phosphorylated, activated cytoplasmic domains of the transforming growth factor-β (TGFβ) receptors were used as probes to screen an expression library that was prepared from a highly TGFβ-responsive intestinal epithelial cell line. One of the TGFβ receptor-interacting proteins isolated was identified to be the mammalian homologue of the LC7 family (mLC7) of dynein light chains (DLCs). This 11-kDa cytoplasmic protein interacts with the TGFβ receptor complex intracellularly and is phosphorylated on serine residues after ligand-receptor engagement. Forced expression of mLC7-1 induces specific TGFβ responses, including an activation of Jun N-terminal kinase (JNK), a phosphorylation of c-Jun, and an inhibition of cell growth. Furthermore, TGFβ induces the recruitment of mLC7-1 to the intermediate chain of dynein. A kinase-deficient form of TGFβ RII prevents both mLC7-1 phosphorylation and interaction with the dynein intermediate chain (DIC). This is the first demonstration of a link between cytoplasmic dynein and a natural growth inhibitory cytokine. Furthermore, our results suggest that TGFβ pathway components may use a motor protein light chain as a receptor for the recruitment and transport of specific cargo along microtublules.
Interorganellar Regulation of Lysosome Positioning by the Golgi Apparatus through Rab34 Interaction with Rab-interacting Lysosomal Protein
Authors: Wang Tuanlao , Hong Wanjin
Published: December 2002

We present evidence to suggest the existence of a regulatory pathway for the Golgi apparatus to modulate the spatial positioning of otherwise distantly located lysosomes. Rab34, a new member of the Rab GTPase family, is associated primarily with the Golgi apparatus. Expression of wild-type or GTP-restricted but not GDP-restricted versions of Rab34 causes spatial redistribution of lysosomes from the periphery to the peri-Golgi region. The regulation of lysosomal positioning by Rab34 depends on its association with the membrane mediated by prenylation and its direct interaction with Rab-interacting lysosomal protein (RILP). This biological activity, mediated by Rab34-RILP interaction, is dependent on Lys82 in the switch I region. Our results have uncovered a novel mechanism for the Golgi apparatus to regulate the spatial distribution of another organelle.
Ran GTPase Cycle and Importins α and β Are Essential for Spindle Formation and Nuclear Envelope Assembly in LivingCaenorhabditis elegansEmbryos
Authors: Askjaer Peter , Galy Vincent , Hannak Eva , Mattaj Iain W.
Published: December 2002

The small GTPase Ran has been found to play pivotal roles in several aspects of cell function. We have investigated the role of the Ran GTPase cycle in spindle formation and nuclear envelope assembly in dividing Caenorhabditis elegans embryos in real time. We found that Ran and its cofactors RanBP2, RanGAP, and RCC1 are all essential for reformation of the nuclear envelope after cell division. Reducing the expression of any of these components of the Ran GTPase cycle by RNAi leads to strong extranuclear clustering of integral nuclear envelope proteins and nucleoporins. Ran, RanBP2, and RanGAP are also required for building a mitotic spindle, whereas astral microtubules are normal in the absence of these proteins. RCC1(RNAi) embryos have similar abnormalities in the initial phase of spindle formation but eventually recover to form a bipolar spindle. Irregular chromatin structures and chromatin bridges due to spindle failure were frequently observed in embryos where the Ran cycle was perturbed. In addition, connection between the centrosomes and the male pronucleus, and thus centrosome positioning, depends upon the Ran cycle components. Finally, we have demonstrated that both IMA-2 and IMB-1, the homologues of vertebrate importin α and β, are essential for both spindle assembly and nuclear formation in early embryos.
Requirement of the N-Terminal Extension for Vacuolar Trafficking and Transport Activity of Yeast Ycf1p, an ATP-binding Cassette Transporter
Authors: Mason Deborah L. , Michaelis Susan
Published: December 2002

Ycf1p is the prototypical member of the yeast multidrug resistance-associated protein (MRP) subfamily of ATP-binding cassette (ABC) transporters. Ycf1p resides in the vacuolar membrane and mediates glutathione-dependent transport processes that result in resistance to cadmium and other xenobiotics. A feature common to many MRP proteins that distinguishes them from other ABC transporters is the presence of a hydrophobic N-terminal extension (NTE), whose function is not clearly established. The NTE contains a membrane spanning domain (MSD0) with five transmembrane spans and a cytosolic linker region (L0). The goal of this study was to determine the functional significance of the NTE of Ycf1p by examining the localization and functional properties of Ycf1p partial molecules, expressed either singly or together. We show that MSD0 plays a critical role in the vacuolar membrane trafficking of Ycf1p, whereas L0 is dispensable for localization. On the other hand, L0 is required for transport function, as determined by monitoring cadmium resistance. We also examine an unusual aspect of Ycf1p biology, namely, the posttranslational proteolytic processing that occurs within a lumenal loop of Ycf1p. Processing is shown to be Pep4p dependent and thus serves as a convenient marker for proper vacuolar localization. The processed fragments associate with each other, suggesting that these natural cleavage products contribute together to Ycf1p function.
Mammalian Septins Nomenclature
Authors: Macara Ian G. , Baldarelli Richard , Field Christine M. , Glotzer Michael , Hayashi Yasuhide , Hsu Shu-Chan , Kennedy Mary B. , Kinoshita Makoto , Longtine Mark , Low Claudia , Maltais Lois J. , McKenzie Louise , Mitchison Timothy J. , Nishikawa Toru , Noda Makoto , Petty Elizabeth M. , Peifer Mark , Pringle John R. , Robinson Phillip J. , Roth Dagmar , Russell S.E. Hilary , Stuhlmann Heidi , Tanaka Manami , Tanaka Tomoo , Trimble William S. , Ware Jerry , Zeleznik-Le Nancy J. , Zieger Barbara
Published: December 2002

There are 10 known mammalian septin genes, some of which produce multiple splice variants. The current nomenclature for the genes and gene products is very confusing, with several different names having been given to the same gene product and distinct names given to splice variants of the same gene. Moreover, some names are based on those of yeast or Drosophilaseptins that are not the closest homologues. Therefore, we suggest that the mammalian septin field adopt a common nomenclature system, based on that adopted by the Mouse Genomic Nomenclature Committee and accepted by the Human Genome Organization Gene Nomenclature Committee. The human and mouse septin genes will be namedSEPT1–SEPT10 and Sept1–Sept10, respectively. Splice variants will be designated by an underscore followed by a lowercase “v” and a number, e.g., SEPT4_v1.
FORECASTING THE DEMAND FOR HEALTH CARE IN OECD NATIONS AND CHINA
Authors: Fogel Robert W.
Published: December 2002

This article argues that the nations of the Organisation for Economic Co‐operation and Development (OECD) are facing a potential disaster: intergenerational conflicts between the large number of workers approaching retirement at an earlier age than ever before in history and the smaller number entering the labor market. These nations are confronted with the choice of whether to default on commitments to pensions and health care, to reduce benefits, or to increase the taxes borne by younger workers. The author argues that this new intergenerational conflict is the result of technophysio evolution, a synergism between technological and physiological improvements that has produced a form of human evolution that is biological but not genetic, rapid, culturally transmitted, and not necessarily stable. The author goes on to argue that an important aspect of technophysio evolution has been a change in the structure of consumption and in the division of discretionary time between work and leisure. The ongoing debate over whether or not the rapid advances in biotechnology will spare OECD nations' health systems from a financial crisis is addressed. Finally, the implications this argument has for forecasting China's future health care costs are discussed.
THE COST OF THE U.S. SUGAR PROGRAM REVISITED
Authors: Beghin John C. , Osta Barbara El , Cherlow Jay R. , Mohanty Samarendu
Published: December 2002

The article analyzes the welfare cost of the U.S. sugar program using a multimarket model of U.S. sweetener markets. The latter includes raw crops, sugar extraction and refining, and sweetener users (food‐processing industries and final consumers). The authors address the industrial organization of food industries using sweeteners and treat the United States as a large importer. With the removal of the program, this article estimates (all figures in 1999 dollars) that in 1998 cane growers, sugar beet growers, and processors would have lost $307, $650, and $89 million, respectively; sweetener users would have gained $1.9 billion. World prices would have increased by 13.2%. The deadweight loss of the program is estimated at $532 million.
A REVISIONIST VIEW OF CHINESE AGRICULTURAL PRODUCTIVITY?
Authors: Mead Robert W.
Published: December 2002

Although productivity increased during the initial Chinese agricultural reforms, there is less clarity about subsequent reform efforts. Several studies find that productivity growth either slowed or stopped in the mid‐1980s, but these studies use official labor statistics that indicate an increasing farm labor force during the late 1980s and early 1990s. Using an alternative calculation of China's farm labor force showing a decrease in agricultural labor, this article reexamines the data on Chinese agricultural productivity and notes that the changes in productivity appear to correlate to changes in economic policies.
TO TAX OR NOT TO TAX? THE CASE OF ELECTRONIC COMMERCE
Authors: Bruce Donald , Fox William , Murray Matthew
Published: December 2002

Despite the intensifying debate over the taxation of Internet commerce, the relevant issues have not been given a systematic treatment in the context of the literature on optimal taxation. This article presents such an analysis and investigates separately the taxation of business purchases of intermediate goods, the taxation of consumer purchases of final goods and services, and the various issues of administration and compliance costs as they apply to the development of E‐commerce. The authors conclude that generally the optimal tax literature cannot be used in support of a blanket tax exemption for Internet purchases. Certain conditions could lead to the optimality of an exemption, but those conditions are not likely to be met in practice.
THE RELATIONSHIP BETWEEN LARGE FISCAL ADJUSTMENTS AND SHORT‐TERM OUTPUT GROWTH UNDER ALTERNATIVE FISCAL POLICY REGIMES
Authors: Miller Stephen M. , Russek Frank S.
Published: December 2002

A small but growing body of literature searches for evidence of non‐Keynesian effects of fiscal contractions. That is, some evidence exists that large fiscal contractions stimulate short‐run economic activity. This article continues that research effort by systematically examining the effects (if any) of unusual fiscal events—either non‐Keynesian results within a Keynesian model or Keynesian results within a neoclassical model—on short‐run economic activity. The authors examine this issue within three separate models—a St. Louis equation, a Hall‐type consumption equation, and a growth accounting equation. The empirical findings do not provide strong systematic support for the view that unusually large fiscal contractions/expansions reverse the effects of normal fiscal events. Moreover, the authors find only limited evidence that trigger points are empirically important.
PACIFIC SALMON RESTORATION: TRADE‐OFFS BETWEEN ECONOMIC EFFICIENCY AND POLITICAL ACCEPTANCE
Authors: Wu Junjie , Skelton‐Groth Katharine , Boggess William G. , Adams Richard M.
Published: December 2002

There is no simple solution to the problems of salmon restoration given substantial political and scientific uncertainties. There are, however, some local findings in Oregon that can provide guidance to resource managers charged with allocating funds for conservation purposes. This article shows that in most salmon habitat investments, there are likely to be strong nonlinearities (cumulative and threshold effects) that mitigate against politically palatable allocation criteria. In fact, this research indicates that decisions based on political equity concerns may actually lead to the lowest possible benefits to society. These scientific nonlinearities may make the political resolution of salmon recovery more difficult.
CORRELATION FUNCTIONS IN BERKOVITS' PURE SPINOR FORMULATION
Authors: TRIVEDI GAUTAM
Published: December 2002

We use Berkovits' pure spinor quantization to compute various three-point tree correlation functions in position-space for the Type IIB superstring. We solve the constraint equations for the vertex operators and obtain explicit expressions for the graviton and axion components of the vertex operators. Using these operators we compute tree level correlation functions in flat space and discuss their extension to the AdS5 × S5 background.
DECOMPOSING THE SU(N) CONNECTION AND THE WU–YANG POTENTIAl
Authors: DUAN YI-SHI , ZHANG PENG-MING
Published: December 2002

Based on the decomposition of SU(2) gauge field, we derive a generalization of the decomposition theory for the SU (N) gauge field. We thus obtain the invariant electromagnetic tensors of SU (N) groups and the extended Wu–Yang potentials. The sourceless solutions are also discussed.
THE HOSOTANI MECHANISM IN BULK GAUGE THEORIES WITH AN ORBIFOLD EXTRA SPACE S1/Z2
Authors: KUBO MASAHIRO , LIM C. S. , YAMASHITA HIROYUKI
Published: December 2002

We pursue the possibility of the scenario in which the Higgs field is identified with the extra-space component of a bulk gauge field. The space–time we take is M4 ⊗ S1/Z2. We show that a nontrivial Z2-parity assignment allows some of the extra-space component to have radiatively induced vev, which strongly modifies the mass spectrum and gauge symmetry of the theory, realized by orbifolding. In particular we investigate the dynamical mass generation of zero-mode fermion and spontaneous gauge symmetry breaking due to the vev. The gauge theories we adopt are a prototype model SU(2) and SU(3) model, of special interest as the realistic minimal scheme to incorporate the standard model SU(2) × U(1).
A NOTE ON SCHWARZSCHILD BLACK HOLE THERMODYNAMICS IN A MAGNETIC UNIVERSE
Authors: RADU EUGEN
Published: December 2002

We prove that the thermodynamic properties of a Schwarzschild black hole are unaffected by an external magnetic field passing through it. Apart from the background subtraction prescription, this result is also obtained by using a counterterm method.