bluuebunny/crossref_metadata_2025 huggingface dataset:
Myosin IIA deficient cells migrate efficiently despite reduced traction forces at cell periphery
Authors: Jorrisch Melissa H. , Shih Wenting , Yamada Soichiro
Published: February 2013

Cell motility is a cornerstone of embryogenesis, tissue remodeling and repair, and cancer cell invasion. It is generally thought that migrating cells grab and exert traction force onto the extracellular matrix in order to pull the cell body forward. While previous studies have shown that myosin II deficient cells migrate efficiently, whether these cells exert traction forces during cell migration in the absence of the major contractile machinery is currently unknown. Using an array of micron-sized pillars as a force sensor and shRNA specific to each myosin II isoform (A and B), we analyzed how myosin IIA and IIB individually regulate cell migration and traction force generation. Myosin IIA and IIB localized preferentially to the leading edge where traction force was greatest, and the trailing edge, respectively. When individual myosin II isoforms were depleted by shRNA, myosin IIA deficient cells lost actin stress fibers and focal adhesions, whereas myosin IIB deficient cells maintained similar actin organization and focal adhesions as wild-type cells. Interestingly, myosin IIA deficient cells migrated faster than wild-type or myosin IIB deficient cells on both a rigid surface and a pillar array, yet myosin IIA deficient cells exerted significantly less traction force at the leading edge than wild-type or myosin IIB deficient cells. These results suggest that, in the absence of myosin IIA mediated force-generating machinery, cells move with minimal traction forces at the cell periphery, thus demonstrating the remarkable ability of cells to adapt and migrate.
Quantitative assay for TALEN activity at endogenous genomic loci
Authors: Hisano Yu , Ota Satoshi , Arakawa Kazuharu , Muraki Michiko , Kono Nobuaki , Oshita Kazuki , Sakuma Tetsushi , Tomita Masaru , Yamamoto Takashi , Okada Yasushi , Kawahara Atsuo
Published: February 2013

Artificially designed nucleases such as zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) can induce a targeted DNA double-strand break at the specific target genomic locus, leading to the frameshift-mediated gene disruption. However, the assays for their activity on the endogenous genomic loci remain limited. Herein, we describe a versatile modified lacZ assay to detect frameshifts in the nuclease target site. Short fragments of the genome DNA at the target or putative off-target loci were amplified from the genomic DNA of TALEN-treated or control embryos, and were inserted into the lacZα sequence for the conventional blue–white selection. The frequency of the frameshifts in the fragment can be estimated from the numbers of blue and white colonies. Insertions and/or deletions were easily determined by sequencing the plasmid DNAs recovered from the positive colonies. Our technique should offer broad application to the artificial nucleases for genome editing in various types of model organisms.
The KMN protein network – chief conductors of the kinetochore orchestra
Authors: Varma Dileep , Salmon E. D.
Published: February 2013

Successful completion of mitosis requires that sister kinetochores become attached end-on to the plus ends of spindle microtubules (MTs) in prometaphase, thereby forming kinetochore microtubules (kMTs) that tether one sister to one spindle pole and the other sister to the opposite pole. Sites for kMT attachment provide at least four key functions: robust and dynamic kMT anchorage; force generation that can be coupled to kMT plus-end dynamics; correction of errors in kMT attachment; and control of the spindle assembly checkpoint (SAC). The SAC typically delays anaphase until chromosomes achieve metaphase alignment with each sister kinetochore acquiring a full complement of kMTs. Although it has been known for over 30 years that MT motor proteins reside at kinetochores, a highly conserved network of protein complexes, called the KMN network, has emerged in recent years as the primary interface between the kinetochore and kMTs. This Commentary will summarize recent advances in our understanding of the role of the KMN network for the key kinetochore functions, with a focus on human cells.
Epithelial barrier assembly requires coordinated activity of multiple domains of the tight junction protein ZO-1
Authors: Rodgers Laurel S. , Beam M. Tanner , Anderson James M. , Fanning Alan S.
Published: February 2013

Tight junctions (TJ) regulate the paracellular movement of ions, macromolecules and immune cells across epithelia. Zonula Occludens (ZO)-1 is a multi-domain polypeptide required for the assembly of TJs. MDCK II cells lacking ZO-1, and its homolog ZO-2, have three distinct phenotypes: Reduced localization of occludin and some claudins to the TJ, increased epithelial permeability, and expansion of the apical actomyosin contractile array found at the apical junction complex (AJC). However, it is unclear exactly which ZO-1 binding domains are required to coordinate these activities. We addressed this question by examining the ability of ZO-1 domain-deletion transgenes to reverse the effects of ZO-depletion. We found that the SH3 domain and the U5 motif are required to recruit ZO-1 to the AJC and that localization is a prerequisite for normal TJ and cytoskeletal organization. The PDZ2 domain is not required for localization of ZO-1 to the AJC, but is necessary to establish the characteristic continuous circumferential band of ZO-1, occludin and claudin-2. PDZ2 is also required to establish normal permeability, but is not required for normal cytoskeletal organization. Finally, our results demonstrate that PDZ1 is critical for the normal organization of both the TJ and the AJC cytoskeleton. Our results establish that ZO-1 acts as a true scaffolding protein and that the coordinated activity of multiple domains is required for normal TJ structure and function.
4-hydroxytamoxifen leads to PrPSc clearance by conveying both PrPC and PrPSc to lysosomes independently of autophagy
Authors: Marzo Ludovica , Marijanovic Zrinka , Browman Duncan , Chamoun Zeina , Caputo Anna , Zurzolo Chiara
Published: February 2013

Prion diseases are fatal neurodegenerative disorders involving the abnormal folding of a native cellular protein, named PrPC, to a malconformed aggregation-prone state, enriched in beta sheet secondary structure, denoted PrPSc. Recently, autophagy has garnered considerable attention as a cellular process with the potential to counteract neurodegenerative diseases of protein aggregation such as Alzheimer's disease, Huntington's disease, and Parkinson's disease. Stimulation of autophagy by chemical compounds has also been shown to reduce PrPSc in infected neuronal cells and prolong survival times in mice models. Consistent with previous reports we demonstrate that autophagic flux is increased in chronically infected cells. However, in contrast to recent findings we show that autophagy is not causative of a reduction in scrapie burden. We report that in infected neuronal cells different compounds known to stimulate autophagy are ineffective in increasing autophagic flux and in reducing PrPSc. We further demonstrate that tamoxifen and its metabolite 4-hydroxytamoxifen lead to prion degradation in an autophagy-independent manner by diverting the trafficking of both PrP and cholesterol to lysosomes. Because tamoxifen represents a well-characterized, widely available pharmaceutical our data indicate that it may have applications in the therapy of prion diseases.
Dmon1 controls recruitment of Rab7 to maturing endosomes in Drosophila
Authors: Yousefian Jahan , Troost Tobias , Grawe Ferdi , Sasamura Takeshi , Fortini Mark , Klein Thomas
Published: February 2013

The small GTPases Rab5 and Rab7 are important organisers of endosome formation and maturation. In addition they orchestrate the trafficking of cargo through the endosomal pathway. A crucial event during maturation of endosomes is the replacement of the early organiser Rab5 with the late organiser Rab7 in a process called Rab conversion. Rab conversion is a prerequisite for late events, chief among them the fusion of matured endosomes with the lysosome. Recent work identifies members of the Sand1/Mon1 protein family as critical factors during this process. Here, we present the analysis of the function of the Drosophila ortholog of mon1/sand1, Dmon1. We found that loss of function of Dmon1 results in an enlargement of maturing endosomes and loss of their association with Rab7. The enlarged endosomes contain Notch and other trans-membrane proteins as cargo. We report the first EM analysis of Dmon1 cells in a metazoan and extend the analysis of the endosomes in mutant cells. Our results suggest that the phenotype can be explained by the loss of function of Rab7. Moreover, the endosomes of Dmon1 cells matures normally in many aspects despite the loss of association with Rab7. Surprisingly, we do not observe activation of signalling pathways, such as Notch or RTK, in Dmon1 cells, even if the formation of intraluminal vesicles is suppressed in addition.
The E3 ubiquitin ligases RNF126 and Rabring7 regulate endosomal sorting of the Epidermal Growth Factor Receptor
Authors: Smith Christopher J. , Berry Donna M. , McGlade C. Jane
Published: February 2013

Activation of the EGFR results in internalization and ubiquitin dependent endosomal sorting leading to lysosomal degradation. Here we describe the role of the RING finger domain-containing protein RNF126 and the related protein, Rabring7 in EGFR endosomal sorting. We demonstrate that RNF126 specifies K48 linked chains with UbcH5b and also functions with Ubc13/Uev1a to form K63-linked chains in vitro. RNF126 and Rabring7 associate with the epidermal growth factor receptor (EGFR) through a ubiquitin binding zinc finger domain and both E3s promote ubiquitination of EGFR. In the absence of c-Cbl or in cells expressing Cbl-70Z, the binding of RNF126 and Rabring7 to the EGFR is reduced suggesting that RNF126 and Rabring7 function downstream of c-Cbl. In HeLa cells depleted of either RNF126 or Rabring7 the EGFR is retained in a late endocytic compartment and is inefficiently degraded. In addition, depletion of RNF126 or Rabring7 destabilizes ESCRT-II and reduces the number of multivesicular bodies formed after EGF stimulation. In addition, we show that the depletion of Rabring7 also attenuates the degradation of MET and that both RNF126 and Rabring7 regulate the sorting of CXCR4 from an early endocytic compartment. Together these data suggest that RNF126 and Rabring7 play a role in the ubiquitin dependent sorting and down regulation of membrane receptors.
Genetic Interaction between pku300 and fbn2b Controls Endocardial Cell Proliferation and Valve Development in Zebrafish
Authors: Wang Xu , Yu Qingming , Wu Qing , Bu Ye , Chang Nan-Nan , Yan Shouyu , Zhou Xiao-Hai , Zhu Xiaojun , Xiong Jing-Wei
Published: February 2013

Abnormal cardiac valve morphogenesis is a common cause of human congenital heart diseases. The molecular mechanisms regulating endocardial cell proliferation and differentiation into cardiac valves remain largely unknown, although great progress has been made on the endocardial contribution to the atrioventricular cushion and valve formation. Here, we found that scotch tapete382 (scote382) encoded a novel transmembrane protein critical for endocardial cell proliferation and heart valve development. The zebrafish scote382 mutant showed diminished endocardial cell proliferation, lack of heart valve leaflets, abnormal common cardinal and caudal veins. Positional cloning revealed a C946T nonsense mutation of a novel gene pku300 in the scote382 locus, which encoded a 540 amino-acid protein on cell membranes with one putative transmembrane domain and three IgG domains. A known G3935T missense mutation of fbn2b was also found ∼570 kb away from pku300 in scote382 mutants. The genetic mutant scopku300, derived from scote382, only had the C946T mutation of pku300 and showed reduced numbers of atrial endocardial cells and an abnormal common cardinal vein. Morpholino knockdown of fbn2b led to fewer atrial endocardial cells and an abnormal caudal vein. Knockdown of both pku300 and fbn2b phenocopied these phenotypes in scote382 genetic mutants. pku300 transgenic expression in endocardial/endothelial cells, but not myocardial cells, partially rescued the atrial endocardial defects in scote382 mutants. Mechanistically, pku300 and fbn2b were required for endocardial cell proliferation, endocardial Notch signaling, and the proper formation of endocardial cell adhesion and tight junctions, all of which are critical for cardiac valve development. We conclude that pku300 and fbn2b represent the few genes capable of regulating endocardial cell proliferation and signaling in zebrafish cardiac valve development.
Endoglin Regulates the Activation and Quiescence of Endothelium by Participating in Canonical and Non-Canonical TGF-β Signaling Pathways
Authors: Park SunYoung , DiMaio Terri A. , Liu Wei , Wang Shoujian , Sorenson Christine M. , Sheibani Nader
Published: February 2013

Endoglin (Eng) is an auxiliary receptor for TGFβ with important roles in vascular function. TGFβ regulates angiogenesis through balancing the pro-proliferative and pro-differentiation signaling pathways of endothelial cells (EC). However, the contribution of endoglin to these TGFβ activities, and more specifically modulation of EC phenotype remain elusive. Mutations in endoglin cause hereditary hemorrhagic telangiectasia-1 in humans. The Eng +/− mice are viable and exhibit some of the vascular defects seen in humans with endoglin haploinsufficiency. Here we show that haploinsufficiency of endoglin resulted in attenuation of retinal neovascularization during oxygen-induced ischemic retinopathy. Although the importance of endoglin expression in angiogenesis and vascular development has been demonstrated the underlying mechanisms remain obscure. To gain detailed insight into the cell autonomous regulatory mechanisms which impact angiogenic properties of EC, we prepared retinal EC from Eng+/+ and Eng+/− Immorto mice. The Eng +/− EC were more adherent, less migratory, and failed to undergo capillary morphogenesis. Aortic sprouting angiogenesis was similarly attenuated in aortas from Eng +/− mice. In addition, Eng +/− EC expressed increased levels of VEGF but reduced expression of endothelial nitric oxide synthase and NO production. Mechanistically, these changes were consistent with sustained activation of MAPK pathways, and aberrant Smad-dependent signaling pathways in Eng +/− EC. Together our results underscore the importance of endoglin in both canonical and non-canonical TGFβ signaling pathways modulating both the activation and quiescence of the endothelium during angiogenesis.
Coordinate regulation of eIF2α phosphorylation by dPPP1R15 and dGCN2 is required during development
Authors: Malzer Elke , Szajewska-Skuta Małgorzata , Dalton Lucy E. , Thomas Sally E. , Hu Nan , Skaer Helen , Lomas David A. , Crowther Damian C , Marciniak Stefan J.
Published: February 2013

Phosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α) by the kinase GCN2 attenuates protein synthesis during amino acid starvation in yeast, while in mammals a family of related eIF2α kinases regulate translation in response to a variety of stresses. Unlike single-celled eukaryotes, mammals also possess two specific eIF2α phosphatases, PPP1R15a and PPP1R15b, whose combined deletion leads to a poorly understood early embryonic lethality. We report the characterization of the first non-mammalian eIF2α phosphatase and the use of Drosophila to dissect its role during development. The Drosophila protein demonstrates features of both mammalian proteins, including limited sequence homology and association with the endoplasmic reticulum. Of note, although not transcriptionally regulated, its expression is controlled by the presence of upstream open reading frames in its 5′UTR enabling induction in response to eIF2α phosphorylation. Moreover, we show that its expression is necessary for embryonic and larval development and that this is to oppose the inhibitory effects of GCN2 on anabolic growth.
Nrf2 Participates in Regulating Maternal Hepatic Adaptations to Pregnancy
Authors: Zou Yuhong , Hu Min , Bao Qi , Kumar Sudhanshu , Chan Jefferson Y. , Dai Guoli
Published: February 2013

Pregnancy induces widespread adaptive responses in maternal organ systems including the liver. The maternal liver exhibits significant growth by increasing the number and size of hepatocytes, with largely unknown mechanisms. Nrf2 mediates cellular defense against oxidative stress and inflammation and also regulates liver regeneration. To determine whether Nrf2 is involved in the regulation of maternal hepatic adaptations to pregnancy, we assessed the proliferation and size of maternal hepatocytes and the associated molecular events in wild-type and Nrf2-null mice at various stages of gestation. We found that wild-type maternal hepatocytes underwent proliferation and size reduction during the first half, and size increase without overt replication during the second half, of pregnancy. Although pregnancy decreased Nrf2 activity in the maternal liver, Nrf2 deficiency caused a delay in maternal hepatocyte proliferation, concomitant with dysregulation of the activation of Cyclins D1, E1, and, more significantly, A2. Remarkably, due to Nrf2 absence, the maternal hepatocytes were largely prevented from reducing their sizes during the first half of pregnancy, which was associated with an increase in mTOR activation. During the second half of pregnancy, both genotypes of maternal hepatocytes showed continuous volume increase accompanied by persistent activation of mTOR. However, the lack of Nrf2 resulted in dysregulation of the activation of the mTOR upstream regulator AKT1 and the mTOR target p70SK6 and thus disruption of the AKT1/mTOR/p70S6K pathway, which is known to control cell size. The finding suggests an mTOR-dependent and AKT1- and p70S6K-independent compensatory mechanism when Nrf2 is deficient. In summary, our study demonstrates that Nrf2 is required for normal maternal hepatic adjustments to pregnancy by ensuring proper regulation of the number and size of maternal hepatocytes.
cGMP-dependent Protein Kinase Iβ Regulates Breast Cancer Cell Migration and Invasion via a Novel Interaction with the Actin/Myosin-associated Protein Caldesmon
Authors: Schwappacher Raphaela , Rangaswami Hema , Su-Yuo Jacqueline , Hassad Aaron , Spitler Ryan , Casteel Darren E.
Published: February 2013

The two isoforms of type I cGMP-dependent protein kinases (PKGIα and PKGIβ) differ in their first ∼100 amino acids, giving each isoform unique dimerization and autoinhibitory domains. The dimerization domains form coiled-coil structures and serve as platforms for isoform-specific protein-protein interactions. Using the PKGIβ dimerization domain as an affinity probe in a proteomic screen, we identified the actin/myosin-associated protein caldesmon (CaD) as a PKGIβ-specific binding protein. PKGIβ phosphorylated human CaD on serine 12 in vitro and in intact cells. Phosphorylation on serine 12 or mutation of serine 12 to glutamic acid (S12E) reduced the interaction between CaD and myosin IIA. Since CaD inhibits myosin ATPase activity and regulates cell motility, we examined the effects of PKGIβ and CaD on cell migration and invasion. Inhibition of the NO/cGMP/PKG pathway reduced migration and invasion of human breast cancer cells, while PKG activation enhanced their motility and invasion. siRNA-mediated knockdown of endogenous CaD had pro-migratory and pro-invasive effects in human breast cancer cells, and reconstituting cells with wild-type CaD slowed migration/invasion; however, CaD containing a phospho-mimetic S12E mutation failed to reverse the pro-migratory and pro-invasive activity of CaD depletion. Our data suggest that PKGIβ enhances breast cancer cell motility and invasive capacity, at least in part, by phosphorylating CaD. These findings identify a pro-migratory and pro-invasive function for PKGIβ in human breast cancer cells, suggesting that PKGIβ is a potential target for breast cancer treatment.
MicroRNA miR-24 Enhances Tumor Invasion and Metastasis by Targeting PTPN9 and PTPRF to Promote EGF Signaling
Authors: Du William W. , Fang Ling , Li Minhui , Yang Xiangling , Liang Yaoyun , Peng Chun , Qian Wei , O'Malley Yunxia Q. , Askeland Ryan W , Sugg Sonia , Qian Jun , Lin Jiang , Jiang Zide , Yee Albert J. , Sefton Michael , Deng Zhaoqun , Shan Sze Wan , Wang Chia-Hui , Yang Burton B.
Published: February 2013

MicroRNAs are known to play regulatory roles in gene expression associated with cancer development. We analyzed microRNA miR-24 levels in patients with breast carcinoma and found that miR-24 was higher in breast carcinoma samples than in breast benign tissues. We generated constructs expressing miR-24 and studied its functions using both in vitro and in vivo techniques. We found that the ectopic expression of miR-24 promoted breast cancer cell invasion and migration. In vivo experiments indicated that the expression of miR-24 enhanced tumor growth, invasion to local tissues, metastasis to lung tissues, and decreased overall mouse survival. In the miR-24 expressing cells and tumors, EGFR was highly phosphorylated, while expression of the phosphatases PTPN9 and PTPRF were repressed. We confirmed that miR-24 could directly target both PTPN9 and PTPRF. Consistent with this, we found that the levels of pEGFR were higher while the levels of PTPN9 and PTPRF were lower in the patients with metastatic breast carcinoma. Ectopic expression of PTPN9 and PTPRF decreased pEGFR levels, cell invasion, migration, and tumor metastasis. Furthermore, we found that the levels of MMP2, MMP11, pErk, and ADAM15 were up-regulated, while the levels of TIMP2 were down-regulated, all of which supported the roles of miR-24 in tumor invasion and metastasis. Our results suggest that miR-24 plays a key role in breast cancer invasion and metastasis. miR-24 could potentially be a target for cancer intervention.
Tetraspanin18 is a FoxD3-responsive antagonist of cranial neural crest epithelial to mesenchymal transition that maintains Cadherin6B protein
Authors: Fairchild Corinne L. , Gammill Laura S.
Published: February 2013

During epithelial to mesenchymal transition (EMT), tightly associated, polarized epithelial cells become individual mesenchymal cells capable of migrating. Here, we investigate the role of the transmembrane protein tetraspanin18 (Tspan18) in cranial neural crest EMT. Tspan18 mRNA is expressed in premigratory cranial neural crest cells, but is absent from actively migrating neural crest cells. Tspan18 knock down leads to a concomitant loss of Cadherin6B (Cad6B) protein, while Cad6B protein persists when Tspan18 expression is extended. As the temporal profile of Cad6B mRNA downregulation is unaffected in these embryos, this indicates that Tspan18 maintains Cad6B protein levels and reveals that Cad6B is regulated by post-translational mechanisms. Although downregulation of Tspan18 is necessary, it is not sufficient for neural crest migration: the timing of neural crest emigration, basal lamina break down and Cad7 upregulation proceed normally in Tspan18-deficient cells. This emphasizes the need for coordinated transcriptional and post-translational regulation of Cad6B during EMT and illustrates that Tspan18-antagonized remodeling of cell-cell adhesions is only one step in preparing for cranial neural crest migration. Unlike Cad6B, which is transcriptionally repressed by Snail2, Tspan18 expression is downstream of the winged-helix transcription factor FoxD3, providing a new transcriptional input into cranial neural crest EMT. Altogether our data reveal post-translational regulation of Cad6B protein levels by Tspan18 that must be relieved by a FoxD3-dependent mechanism in order for cranial neural crest cells to migrate. These results offer novel insight into the molecular mechanisms of cranial neural crest EMT and expand our understanding of tetraspanin function relevant to metastasis.
Niflumic acid disrupts marine spermatozoan chemotaxis without impairing the spatiotemporal detection of chemoattractant gradients
Authors: Guerrero Adán , Espinal Jesús , Wood Christopher D. , Rendón Juan M. , Carneiro Jorge , Martínez-Mekler Gustavo , Darszon Alberto
Published: February 2013

In many broadcast-spawning marine organisms, oocytes release chemicals that guide conspecific spermatozoa towards their source through chemotaxis. In the sea urchin Lytechinus pictus, the chemoattractant peptide speract triggers a train of fluctuations of intracellular Ca2+ concentration in the sperm flagella. Each transient Ca2+ elevation leads to a momentary increase in flagellar bending asymmetry, known as a chemotactic turn. Furthermore, chemotaxis requires a precise spatiotemporal coordination between the Ca2+-dependent turns and the form of chemoattractant gradient. Spermatozoa that display Ca2+-dependent turns while swimming down the chemoattractant gradient, and conversely suppress turning events while swimming up gradient, successfully approach the center of the gradient. Previous experiments in Strongylocentrotus purpuratus sea urchin spermatozoa showed that niflumic acid (NFA), an inhibitor of several ion channels, drastically altered the speract-induced Ca2+ fluctuations and swimming patterns. In this study, mathematical modeling of the speract-dependent Ca2+ signaling pathway suggests that NFA, by potentially affecting HCN, CaCC and CaKC channels, may alter the temporal organization of Ca2+ fluctuations, and therefore disrupt chemotaxis. Here we investigate our hypothesis using a novel automated method for analyzing sperm behavior. We show that NFA does indeed disrupt chemotactic responses of L. pictus spermatozoa, although the temporal coordination between the Ca2+-dependent turns and the form of chemoattractant gradient is unaltered. Instead, NFA disrupts sperm chemotaxis by altering the arc length traveled during each chemotactic turning event. This alteration in the chemotactic turn trajectory disorientates spermatozoa at the termination of the turning event. We conclude that NFA disrupts chemotaxis without affecting how the spermatozoa decode environmental cues.
AMPK phosphorylates GBF1 for mitotic Golgi disassembly
Authors: Mao Luna , Li Ning , Guo Yajuan , Xu Xiaobin , Gao Luying , Xu Yinfeng , Zhou Linfu , Liu Wei
Published: February 2013

In mammalian cells, the Golgi apparatus undergoes extensive fragmentation during mitosis; this is required not only for the partitioning of the complex but also for the process of mitosis. However, the molecular mechanism underlying the mitotic fragmentation of the Golgi is far from clear. Here, we show that AMP-activated protein kinase (AMPK) is phosphorylated and activated when cells enter mitosis. Activated AMPK phosphorylates GBF1, a guanine nucleotide exchange factor (GEF) for Arf-GTPases at Thr1337, disassociating GBF1 from the Golgi membrane and abolishing the action of GBF1 as an Arf1-GEF. We further demonstrate that the phosphorylation of AMPK and GBF1 is essential for Golgi disassembly and subsequent mitosis entry. These data suggest that AMPK-GBF1-Arf1 signaling is involved in the regulation of Golgi fragmentation during mitosis.
Mature MiR-17-5p and passenger miR-17-3p induce hepatocellular carcinoma by targeting PTEN, GalNT7, and vimentin in different signal pathways
Authors: Shan Sze Wan , Fang Ling , Shatseva Tatiana , Rutnam Zina Jeyapalan , Yang Xiangling , Lu Wei-Yang , Xuan Jim W. , Deng Zhaoqun , Yang Burton B.
Published: February 2013

To study the physiological role of a single miRNA, we generated transgenic mice expressing the miRNA precursor miR-17 and found that the mature miR-17-5p and the passenger strand miR-17-3p were abundantly expressed. Here, we showed that mature MiR-17-5p and passenger strand miR-17-3p could synergistically induce the development of hepatocellular carcinoma. The mature miR-17-5p exerted this function by repressing the expression of PTEN. On the other hand, the passenger strand miR-17-3p repressed expression of vimentin, an intermediate filament with the ability to modulate metabolism, and GalNT7, an enzyme that regulates metabolism of liver toxin galactosamine. Hepatocellular carcinoma cells HepG2 transfected with miR-17 formed larger tumors with more blood vessels and less tumor cell death than mock cells. Expression of miR-17 precursor modulated HepG2 proliferation, migration, survival, morphogenesis, colony formation, and inhibited endothelial tube formation. Silencing of PTEN, vimentin, or GalNT7 with their respective siRNAs enhanced proliferation and migration. Re-expressing these molecules reversed their roles in proliferation, migration and tumorigenesis. Dissecting experiments indicated that these three molecules did not cross talk with each other, but appear to have functioned in different signaling pathways. Our results demonstrated that a mature miRNA can function synergistically with its passenger strand leading to the same phenotype but by regulating different targets located in different signaling pathways. We anticipate that our assay will serve as a helpful model for studying miRNA regulation.
Flagellar regeneration requires cytoplasmic microtubule depolymerization and kinesin-13
Authors: Wang Liang , Piao Tian , Cao Muqing , Qin Tao , Huang Lei , Deng Haiteng , Mao Tonglin , Pan Junmin
Published: February 2013

In ciliated cells, two types of microtubules can be categorized – cytoplasmic and axonemal. It has been shown that axonemal tubulins come from a “cytoplasmic pool” during cilia regeneration. However, the identity and regulation of this “pool” is not understood. Previously, we have shown that Chlamydomonas kinesin-13 (CrKin13) is phosphorylated during flagellar regeneration, and required for proper flagellar assembly. Here, we show that CrKin13 regulates depolymerization of cytoplasmic microtubules to control flagellar regeneration. After flagellar loss and before flagellar regeneration, cytoplasmic microtubules were quickly depolymerized, which were evidenced by appearance of sparse and shorter microtubule arrays and increased free tubulins in the cell body. Knockdown of CrKin13 expression by RNAi inhibited depolymerization of cytoplasmic microtubules and impaired flagellar regeneration. In vitro assay showed that CrKin13 possessed microtubule depolymerization activity. CrKin13 underwent phosphorylation during microtubule depolymerization and phosphorylation induced targeting of CrKin13 to microtubules. The phosphorylation of CrKin13 occurred at residues S100, T469 and S522 as determined by mass spectrometry. Abrogation of CrKin13 phosphorylation at S100 but not at other residues by inducing point mutation prevented CrKin13 targeting to microtubules. We propose that CrKin13 depolymerizes cytoplasmic microtubule to provide tubulin precursors for flagellar regeneration.
Designed Orthogonal Sample Spiking Based Calibrations for Quantitative Liquid Phase Measurements with near Infrared Spectroscopy in an Anaerobic Digestion Process
Authors: Dahlbacka John , Lillhonga Tom , Döring Marco
Published: February 2013

Biogas production through anaerobic digestion (AD) plays an important role in the effort to reduce the amount of greenhouse gas emissions from energy production. At the same time commercial digestion processes are commonly operated well below their maximal capacity. One reason for this is the lack of suitable sensors for monitoring key parameters in the process. Near infrared (NIR) spectroscopy is seen as a potentially interesting technique for liquid phase measurements in AD processes. In this work, the concentrations of the important constituents, ammonium, acetate, propionate and total volatile fatty acids (TVFA), were measured by combining NIR transmittance measurements and partial least-squares (PLS) models. In order to obtain additional calibration data without having to perform additional reference measurements, and in order to enhance the constituent specific correlation in the calibration data, a spiking procedure was implemented according to a central composite design. This methodology reduced the root mean square error of prediction ( RMSEP) for ammonium from 176 mg L−1 to 127 mg L−1, for acetate from 334 mg L−1 to 260 mg L−1 for propionate from 258 mg L−1 to 203 mg L−1 and for TVFA from 858 mg L−1 to 704 mg L−1. The spiking procedure also significantly increased the correlation between model predictions on pure constituent spectra and reference values, compared to that of models based on non-spiked data.
Perihilar branching patterns of renal artery and extrarenal length of arterial branches and tumour-feeding arteries on multidetector CT angiography
Authors: Kang W Y , Sung D J , Park B J , Kim M J , Han N Y , Cho S B , Kang C H , Kang S H
Published: February 2013

Objective:

The purpose of our study was to assess the extrarenal length of renal arterial branches and tumour-feeding arteries on multidetector CT (MDCT) angiography, in addition to the perihilar branching patterns, with relevance to segmental artery clamping.

Methods:

MDCT angiograms of 64 patients with renal masses <4 cm were retrospectively reviewed by 2 radiologists. The perihilar branching patterns of the single main renal artery were assessed according to the number of pre-segmental and segmental arteries. The extrarenal lengths of segmental plus pre-segmental arteries and the tumour-feeding arteries, measured on volume-rendered images, were compared according to the vascular segmentation and the tumour location, respectively.

Results:

In the 116 kidneys, 1 pre-segmental plus 5 segmental arteries (n=48) was the most common branching pattern. The mean extrarenal length of the inferior segmental plus pre-segmental arteries (33.05 mm) and the posterior segmental plus pre-segmental arteries (32.30 mm) was longer than any of the other segmental plus pre-segmental arteries (apical, 23.87 mm; superior, 26.80 mm; middle, 29.23 mm) (p<0.05). The mean extrarenal length of the lower pole tumour-feeding arteries (35.94 mm) was longer than those of the upper and mid-pole tumour-feeding arteries (24.95 mm, 29.62 mm), with significant difference between the lower and the upper pole tumour-feeding arteries (p<0.05).

Conclusion:

Tumours in the lower pole, supplied by the inferior or posterior segmental artery, may be more amenable to segmental artery clamping.

Advances in knowledge:

MDCT angiography with volume rendering can demonstrate the extrarenal length of tumour-feeding arteries and may help in determining the accessibility for segmental artery clamping.
Tertiary structure-based analysis of microRNA–target interactions
Authors: Gan Hin Hark , Gunsalus Kristin C.
Published: February 2013

Current computational analysis of microRNA interactions is based largely on primary and secondary structure analysis. Computationally efficient tertiary structure-based methods are needed to enable more realistic modeling of the molecular interactions underlying miRNA-mediated translational repression. We incorporate algorithms for predicting duplex RNA structures, ionic strength effects, duplex entropy and free energy, and docking of duplex–Argonaute protein complexes into a pipeline to model and predict miRNA–target duplex binding energies. To ensure modeling accuracy and computational efficiency, we use an all-atom description of RNA and a continuum description of ionic interactions using the Poisson–Boltzmann equation. Our method predicts the conformations of two constructs of Caenorhabditis elegans let-7 miRNA–target duplexes to an accuracy of ∼3.8 Å root mean square distance of their NMR structures. We also show that the computed duplex formation enthalpies, entropies, and free energies for eight miRNA–target duplexes agree with titration calorimetry data. Analysis of duplex–Argonaute docking shows that structural distortions arising from single-base-pair mismatches in the seed region influence the activity of the complex by destabilizing both duplex hybridization and its association with Argonaute. Collectively, these results demonstrate that tertiary structure-based modeling of miRNA interactions can reveal structural mechanisms not accessible with current secondary structure-based methods.
The Stock Selection and Performance of Buy-Side Analysts
Authors: Groysberg Boris , Healy Paul , Serafeim George , Shanthikumar Devin
Published: February 2013

Prior research on equity analysts focuses almost exclusively on those employed by sell-side investment banks and brokerage houses. Yet investment firms undertake their own buy-side research, and their analysts face different stock selection and recommendation incentives than their sell-side peers. We examine the selection and performance of stocks recommended by analysts at a large investment firm relative to those of sell-side analysts from mid-1997 to 2004. We find that the buy-side firm's analysts issue less optimistic recommendations for stocks with larger market capitalizations and lower return volatility than their sell-side peers, consistent with their facing fewer conflicts of interest and having a preference for liquid stocks. Tests with no controls for these effects indicate that annualized buy-side strong buy/buy recommendations underperform those for sell-side peers by 5.9% using market-adjusted returns and by 3.8% using four-factor model abnormal returns. However, these findings are driven by differences in the stocks recommended and their market capitalization. After controlling for these selection effects, we find no difference in the performance of the buy- and sell-side analysts' strong buy/buy recommendations.

This paper was accepted by Mary Barth, accounting.
Guilt by Association: Strategic Failure Prevention and Recovery Capacity Investments
Authors: Kim Sang-Hyun , Tomlin Brian
Published: February 2013

We examine technological systems that display the following characteristics: (i) unplanned outages incur significant costs, (ii) the system experiences an outage if one or more of its subsystems fails, (iii) subsystem failures may occur simultaneously, and (iv) subsystem recovery requires specific resources and capabilities that are provided by different firms. Firms may invest in two measures to enhance system availability—namely, failure prevention and recovery capacity. If recovery capacity investment is the only option, we find that the firms in a decentralized setting overinvest in capacity, resulting in higher system availability but at a higher cost. If both investments can be made, we find that the firms underinvest in failure prevention and overinvest in recovery capacity. That is, firms in a decentralized setting shift their focus from preventing failures to responding to failures. The net effect is lower system availability, reversing the conclusion above. We also find that, unexpectedly, firms are more willing to let their subsystems fail if a joint failure is more likely to occur.

This paper was accepted by Martin Lariviere, operations management.
Designing Buyback Contracts for Irrational But Predictable Newsvendors
Authors: Becker-Peth Michael , Katok Elena , Thonemann Ulrich W.
Published: February 2013

One of the main assumptions in research on designing supply contracts is that decision makers act in a way that maximizes their expected profit. A number of laboratory experiments demonstrate that this assumption does not hold. Specifically, faced with uncertain demand, decision makers place orders that systematically deviate from the expected profit maximizing levels. We have added to this body of knowledge by demonstrating that ordering decisions also systematically depend on individual contract parameters and by developing a behavioral model that captures this systematic behavior. We proceed to test our behavioral model using laboratory experiments and use the data to derive empirical model parameters. We then test our approach in out-of-sample validation experiments that confirm that, indeed, contracts designed using the behavioral model perform better than contracts designed using the standard model.

This paper was accepted by Christian Terwiesch, operations management.
Mixed Bundling of Two Independently Valued Goods
Authors: Bhargava Hemant K.
Published: February 2013

This paper develops analytical results and insights for the mixed bundling problem of pricing a product line consisting of two component goods (with valuations distributed uniformly and independently in [0, a1] and [0, a2], respectively, with a1≤ a2) and a bundle of the two goods. This setting has previously been considered analytically intractable. By deriving component good prices as exact algebraic functions of the optimal bundle price, I reduce the multiproduct pricing problem to a univariate nonlinear optimization problem in the bundle price. When the two component goods are sufficiently asymmetric in valuations, the optimal solution is partial mixed bundling (one component good is not sold separately), for which I derive exact conditions and optimal prices. An exact analytical solution is also given for mixed bundling of two information goods (component goods with zero marginal costs). For the general case, I derive a closed-form approximation of the optimal bundle price: pB= 0.5724w1+ 0.5695w2+ 0.3516a1+ 0.4889a2+ 0.0054(a2/a1)w1−0.0201(a2/a1)w2. The approximation is highly accurate (the resulting profit is within a percent of the exact optimal profit), efficient (instant solutions for specific problem instances), and useful in other applications where mixed bundling is a subproblem. I demonstrate this by applying the solution to the mixed bundle problem in a vertical channel where a retailer must bundle and price component goods from multiple independent manufacturers.

This paper was accepted by Lorin Hitt, information systems.
Demand Uncertainty and Excess Supply in Commodity Contracting
Authors: Popescu Dana G. , Seshadri Sridhar
Published: February 2013

We examine how different characteristics of product demand and market impact the relative sales volume in the forward and spot markets for a commodity whose aggregate demand is uncertain. In a setting where either the forward contracts are binding quantity commitments between buyers and suppliers or the forward production takes place before the uncertainty in demand is resolved, we find that a combination of factors that include market concentration, demand risk, and price elasticity of demand will determine whether a commodity will be sold mainly through forward contracts or in the spot market. Previous findings in the literature show that when participants are risk neutral, the ratio of forward sales to spot sales is a function of market concentration alone; also, the lower the concentration, the higher this ratio. These findings hold under the assumption that demand is either deterministic or, if demand is uncertain, all production takes place after uncertainty is fully resolved and production plans can be altered instantaneously and costlessly. In our setting, however, we find that even a low level of demand risk can reverse the nature of supply in a highly competitive (low concentration) market, by shifting it from predominantly forward-driven to predominantly spot-driven supply. In markets with high concentration, the price elasticity of demand will determine whether the supply will be predominantly spot-driven or forward-driven. Our analysis suggests various new hypotheses on the structure of supply in commodity markets.

This paper was accepted by Martin Lariviere, operations management.
Customer-Driven Misconduct: How Competition Corrupts Business Practices
Authors: Bennett Victor Manuel , Pierce Lamar , Snyder Jason A. , Toffel Michael W.
Published: February 2013

Competition among firms yields many benefits but can also encourage firms to engage in corrupt or unethical activities. We argue that competition can lead organizations to provide services that customers demand but that violate government regulations, especially when price competition is restricted. Using 28 million vehicle emissions tests from more than 11,000 facilities, we show that increased competition is associated with greater inspection leniency, a service quality attribute that customers value but is illegal and socially costly. Firms with more competitors pass customer vehicles at higher rates and are more likely to lose customers whom they fail, suggesting that competition intensifies pressure on facilities to provide illegal leniency. We also show that, at least in markets in which pricing is restricted, firms use corrupt and unethical practices as an entry strategy.

This paper was accepted by Bruno Cassiman, business strategy.
Shouting to Be Heard in Advertising
Authors: Anderson Simon P. , de Palma André
Published: February 2013

Advertising competes for scarce consumer attention, so more profitable advertisers send more messages to break through the others' clutter. Multiple equilibria can arise: more messages in aggregate induce more “shouting to be heard,” dissipating profit. Equilibria can involve a small range of loud shouters or large range of quiet whisperers. All advertisers prefer there to be less shouting. There is the largest diversity in message levels for a middling width of advertiser types: both very wide and very narrow widths have only one message per advertiser. The number of advertisers at each message level decreases with the level if the profit distribution is log-convex. Increasing the cost of sending messages can make all advertisers better off. A new technique is given for describing multiple equilibria, by determining how much examination is consistent with a given marginal advertiser.

This paper was accepted by J. Miguel Villas-Boas, marketing.
Is Pay for Performance Detrimental to Innovation?
Authors: Ederer Florian , Manso Gustavo
Published: February 2013

Previous research in economics shows that compensation based on the pay-for-performance principle is effective in inducing higher levels of effort and productivity. On the other hand, research in psychology argues that performance-based financial incentives inhibit creativity and innovation. How should managerial compensation be structured if the goal is to induce managers to pursue more innovative business strategies? In a controlled laboratory setting, we provide evidence that the combination of tolerance for early failure and reward for long-term success is effective in motivating innovation. Subjects under such an incentive scheme explore more and are more likely to discover a novel business strategy than subjects under fixed-wage and standard pay-for-performance incentive schemes. We also find evidence that the threat of termination can undermine incentives for innovation, whereas golden parachutes can alleviate these innovation-reducing effects.

This paper was accepted by David Hsu, entrepreneurship and innovation.
Ethnic Innovation and U.S. Multinational Firm Activity
Authors: Foley C. Fritz , Kerr William R.
Published: February 2013

This paper studies the impact that ethnic innovators have on the global activities of U.S. firms by analyzing detailed data on patent applications and on the operations of the foreign affiliates of U.S. multinational firms. The results indicate that increases in the share of a firm's innovation performed by inventors of a particular ethnicity are associated with increases in the share of that firm's affiliate activity in countries related to that ethnicity. Ethnic innovators also appear to facilitate the disintegration of innovative activity across borders and to allow U.S. multinationals to form new affiliates abroad without the support of local joint venture partners.

This paper was accepted by Bruno Cassiman, business strategy.