bluuebunny/crossref_metadata_2025_split huggingface dataset:
Parvin, a 42 kDa focal adhesion protein, related to the α-actinin superfamily
Authors: Olski Thorsten M. , Noegel Angelika A. , Korenbaum Elena 
Published: April 2021

ABSTRACT
We have identified and cloned a novel 42-kDa protein termed α-parvin, which has a single α-actinin-like actin-binding domain. Unlike other members of the α-actinin superfamily, which are large multidomain proteins, α-parvin lacks a rod domain or any other C-terminal structural modules and therefore represents the smallest known protein of the superfamily. We demonstrate that mouse α-parvin is widely expressed as two mRNA species generated by alternative use of two polyadenylation signals. We analyzed the actin-binding properties of mouse α-parvin and determined the Kd with muscle F-actin to be 8.4±2.1 μM. The GFP-tagged α-parvin co-localizes with actin filaments at membrane ruffles, focal contacts and tensin-rich fibers in the central area of fibroblasts. Domain analysis identifies the second calponin homology domain of parvin as a module sufficient for targeting the focal contacts. In man and mouse, a closely related paralogue β-parvin and a more distant relative γ-parvin have also been identified and cloned. The availability of the genomic sequences of different organisms enabled us to recognize closely related parvin-like proteins in flies and worms, but not in yeast and Dictyostelium. Phylogenetic analysis of α-parvin and its para- and orthologues suggests, that the parvins represent a new family of α-actinin-related proteins that mediate cell-matrix adhesion.
Use of abnormal preprophase bands to decipher division plane determination
Authors: Granger Cheryl L. , Cyr Richard J. 
Published: April 2021

ABSTRACT
Many premitotic plant cells possess a cortical preprophase band of microtubules and actin filaments that encircles the nucleus. In vacuolated cells, the preprophase band is visibly connected to the nucleus by a cytoplasmic raft of actin filaments and microtubules termed the phragmosome. Typically, the location of the preprophase band and phragmosome corresponds to, and thus is thought to influence, the location of the cell division plane. To better understand the function of the preprophase band and phragmosome in orienting division, we used a green fluorescent protein-based microtubule reporter protein to observe mitosis in living tobacco bright yellow 2 cells possessing unusual preprophase bands. Observations of mitosis in these unusual cells support the involvement of the preprophase band/phragmosome in properly positioning the preprophase nucleus, influencing spindle orientation such that the cytokinetic phragmoplast initially grows in an appropriate direction, and delineating a region in the cell cortex that attracts microtubules and directs later stages of phragmoplast growth. Thus, the preprophase band/phragmosome appears to perform several interrelated functions to orient the division plane. However, functional information associated with the preprophase band is not always used or needed and there appears to be an age or distance-dependent character to the information. Cells treated with the anti-actin drug, latrunculin B, are still able to position the preprophase nucleus suggesting that microtubules may play a dominant role in premitotic positioning. Furthermore, in treated cells, spindle location and phragmoplast insertion are frequently abnormal suggesting that actin plays a significant role in nuclear anchoring and phragmoplast guidance. Thus, the microtubule and actin components of the preprophase band/phragmosome execute complementary activities to ensure proper orientation of the division plane.
Microtubules in the fungal pathogen Ustilago maydis are highly dynamic and determine cell polarity
Authors: Steinberg Gero , Wedlich-Söldner Roland , Brill Marianne , Schulz Irene 
Published: April 2021

ABSTRACT
Many fungal pathogens undergo a yeast-hyphal transition during their pathogenic development that requires rearrangement of the cytoskeleton, followed by directed membrane traffic towards the growth region. The role of microtubules and their dynamic behavior during this process is not well understood. Here we set out to elucidate the organization, cellular role and in vivo dynamics of microtubules in the dimorphic phytopathogen Ustilago maydis. Hyphae and unbudded yeast-like cells of U. maydis contain bundles of spindle pole body-independent microtubules. At the onset of bud formation two spherical tubulin structures focus microtubules towards the growth region, suggesting that they support polar growth in G2, while spindle pole body-nucleated astral microtubules participate in nuclear migration in M and early G1. Conditional mutants of an essential α-tubulin gene from U. maydis, tub1, confirmed a role for interphase microtubules in determination of cell polarity and growth. Observation of GFP-Tub1 fusion protein revealed that spindle pole body-independent and astral microtubules are dynamic, with elongation and shrinkage rates comparable to those found in vertebrate systems. In addition, very fast depolymerization was measured within microtubule bundles. Unexpectedly, interphase microtubules underwent bending and rapid translocations within the cell, suggesting that unknown motor activities participate in microtubule organization in U. maydis.
Movies available on-line: http://www.biologists.com/JCS/movies/jcs1792.html
Molecular architecture of the amplified nucleoli of Xenopus oocytes
Authors: Mais Christine , Scheer Ulrich 
Published: April 2021

ABSTRACT
An understanding of the functional organization of nucleoli, the sites of ribosome biosynthesis, is limited by the present uncertainty about the topological arrangement of the transcribing rRNA genes. Since studies with ‘standard’ nucleoli from somatic cells produced conflicting results, we have examined the amplified nucleoli of Xenopus oocytes. These nucleoli are unique in that they contain high copy numbers of rRNA genes, are not attached to chromosomes, lack non-ribosomal DNA and can be examined in light microscopic spread preparations of nuclear contents. By immunostaining and confocal microscopy we show that in growing stage IV oocytes the sites of rDNA are surrounded by the dense fibrillar component. The rDNA is actively transcribed as revealed by BrUTP injection into oocytes and localization of components of the nucleolar transcription machinery (RNA polymerase I and the transcription factor UBF). At the ultrastructural level, the rDNA sites correlate with the fibrillar centers of amplified nucleoli fixed in situ. The results provide clear evidence that the transcriptionally active rRNA genes are confined to the fibrillar centers of the oocyte nucleoli and open the possibility to analyze the protein composition of almost native, transcriptionally highly active nucleolar chromatin by immunofluorescence microscopy.
Diversity in phagocytic signalling
Authors: Greenberg Steven 
Published: April 2021

ABSTRACT
Phagocytosis is a phylogenetically ancient process triggered by particulate ligands, such as bacteria, fungi and environmental particles. The spectrum of phagocytically competent cell types and receptors is diverse, but many of the basic processes of cytoskeletal rearrangement and membrane trafficking that accompany phagocytosis are conserved. Insights from studies of lower eukaryotes, such as Dictyostelium, Caenorhabditis elegans and Drosophila have contributed greatly to our understanding of the phagocytic pathway. Here, I focus on only a few examples of phagocytosis and bacterial invasion in mammalian cells, in which specific receptors have been identified and signal transduction pathways have been mapped. Owing to space constraints, the roles of lipid microdomains in phagocytosis and clearance of apoptotic bodies are not covered. The examples provided below emphasize the ability of the host cell to utilize different surface receptors to engage the phagocytic machinery.
Phosphoinositides in membrane traffic at the synapse
Authors: Cremona Ottavio , Camilli Pietro De 
Published: April 2021

ABSTRACT
Inositol phospholipids represent a minor fraction of membrane phospholipids; yet they play important regulatory functions in signaling pathways and membrane traffic. The phosphorylated inositol ring can act either as a precursor for soluble intracellular messengers or as a binding site for cytosolic or membrane proteins. Hence, phosphorylation-dephosphorylation of phosphoinositides represents a mechanism for regulation of recruitment to the membrane of coat proteins, cytoskeletal scaffolds or signaling complexes and for the regulation of membrane proteins. Recent work suggests that phosphoinositide metabolism has an important role in membrane traffic at the synapse. PtdIns(4,5)P2 generation is implicated in the secretion of at least a subset of neurotransmitters. Furthermore, PtdIns(4,5)P2 plays a role in the nucleation of clathrin coats and of an actin-based cytoskeletal scaffold at endocytic zones of synapses, and PtdIns(4,5)P2 dephosphorylation accompanies the release of newly formed vesicles from these interactions. Thus, the reversible phosphorylation of inositol phospholipids may be one of the mechanisms governing the timing and vectorial progression of synaptic vesicle membranes during their exocytic-endocytic cycle.
Cdk5 mediates changes in morphology and promotes apoptosis of astrocytoma cells in response to heat shock
Authors: Gao Chun , Negash Sewite , Wang Hwai-Shi , Ledee Dolena , Guo Hongtao , Russell Paul , Zelenka Peggy 
Published: April 2021

ABSTRACT
The cyclin-dependent kinase member, Cdk5, is expressed in a variety of cell types, but neuron-specific expression of its activator, p35, is thought to limit its activity to neurons. Here we demonstrate that both Cdk5 and p35 are expressed in the human astrocytoma cell line, U373. Cdk5 and p35 are present in the detergent-insoluble cytoskeletal fraction of this cell line and Cdk5 localizes to filopodia and vinculin-rich regions of cell-matrix contact in lamellopodia. When exposed to a 46°C heat shock, U373 cells change shape, lose cell-matrix contacts and show increased levels of apoptosis. To test whether Cdk5 activation might play a role in these events, U373 cells were stably transfected with histidine-tagged or green fluorescent protein-tagged constructs of Cdk5 or a dominant negative mutation, Cdk5T33. Under normal growth conditions, growth characteristics of the stably transfected lines were indistinguishable from untransfected U373 cells and Cdk5 localization was not changed. However, when subjected to heat shock, cells stably transfected with Cdk5-T33 remained flattened, showed little loss of cell-matrix adhesion, and exhibited significantly lower levels of apoptosis. In contrast, cells that overexpressed wild-type Cdk5 showed morphological changes similar to those seen in untransfected U373 cells in response to heat shock and had significantly higher levels of apoptosis. Heat-shocked cells showed changes in p35 mobility and stability of the Cdk5/p35 complex consistent with endogenous Cdk5 activity. Together these findings suggest that endogenous Cdk5 activity may play a key role in regulating morphology, attachment, and apoptosis in U373 cells, and raise the possibility that Cdk5 may be a general regulator of cytoskeletal organization and cell adhesion in both neuronal and non-neuronal cells.
Rap1 is involved in cell stretching modulation of p38 but not ERK or JNK MAP kinase
Authors: Sawada Yasuhiro , Nakamura Kozo , Doi Koichi , Takeda Kohsuke , Tobiume Kei , Saitoh Masao , Morita Keiichi , Komuro Issei , Vos Kurt De , Sheetz Michael , Ichijo Hidenori 
Published: April 2021

ABSTRACT
Mechanical force or mechanical stress modulates intracellular signal pathways, including the mitogen-activated protein kinase (MAP kinase) cascades. In our system, cell stretching activated and cell contraction inactivated all three MAP kinase pathways (MKK1/2-extracellular signal-regulated kinase (ERK), MKK4 (SEK1)-cJun N-terminal kinase (JNK) and MKK3/6-p38 pathways). However, little is known about the molecular mechanisms that link the mechanical force to the MAP kinase cascades. To test whether Ras and Rap1 are possible components in the stretch-activated MAP kinase pathways, we examined if Ras and Rap1 were activated by cell stretching and if inhibition of their activity decreased the stretch-enhanced MAP kinase activity. Rap1 was activated by cell stretching and inactivated by cell contraction, whereas Ras was inactivated by cell stretching and activated by cell contraction. Rap1GapII and SPA-1, downregulators of Rap1 activity, decreased the stretch-enhanced p38 activity, whereas a dominant-negative mutant of Ras (RasN17) did not inhibit the stretch-initiated activation of MAP kinases. Furthermore, overexpression of Rap1 enhanced p38 activity but not ERK or JNK activity. These results indicate that Rap1 is involved in transducing the stretch-initiated signal to the MKK3/6-p38 pathway, but not to the MEK1/2-ERK or the MKK4 (SEK1)/MKK7-JNK pathway. Thus, Rap1 plays a unique role in force-initiated signal transduction.
VEGF transiently disrupts gap junctional communication in endothelial cells
Authors: Suarez S. , Ballmer-Hofer K. 
Published: April 2021

ABSTRACT
Vascular endothelial growth factor, VEGF, stimulates angiogenesis by directly acting on endothelial cells. The effects of VEGF are mediated by two tyrosine kinase receptors, VEGFR-1 (Flt-1) and VEGFR-2 (Flk-1/KDR) that are highly related to receptors of the platelet derived growth factor (PDGF) receptor family. We are interested in early signalling events downstream from VEGF receptors that affect blood vessel homeostasis.
Endothelial cells form multiple types of cell-cell junctions that are required for cellular organization into complex networks. These junctions also regulate communication among adjacent cells. Stimulation by various growth factors such as epidermal growth factor (EGF) or PDGF has been shown to disrupt cell-cell junctions, consequently affecting cell-to-cell communication. We investigated gap junctional communication (GJC) by monitoring the transfer of a low molecular mass fluorescent tracer molecule between adjacent cells using immunofluorescence microscopy. VEGF maximally blocked GJC 15 minutes after growth factor administration. The cells resumed communication via gap junctions within 1-2 hours after treatment. This early effect of VEGF on communication correlated with changes in the phosphorylation state of one of the proteins involved in gap junction formation, connexin 43 (Cx43). The signalling mechanisms involved in this phenomenon depend on activation of VEGFR-2, impinge on a tyrosine kinase of the Src family and activate the Erk family of MAP kinases. The function of VEGF-mediated disruption of GJC might be to restrict an increase in endothelium permeability to the environment affected by local injury to blood vessels.
Oligomerisation of G-protein-coupled receptors
Authors: Milligan Graeme 
Published: April 2021

ABSTRACT
A range of approaches have recently provided evidence that G-protein-coupled receptors can exist as oligomeric complexes. Both homo-oligomers, comprising multiple copies of the same gene product, and hetero-oligomers containing more than one receptor have been detected. In several, but not all, examples, the extent of oligomerisation is regulated by the presence of agonist ligands, and emerging evidence indicates that receptor hetero-oligomers can display distinct pharmacological characteristics. A chaperonin-like role for receptor oligomerisation in effective delivery of newly synthesised receptors to the cell surface is a developing concept, and recent studies have employed a series of energy-transfer techniques to explore the presence and regulation of receptor oligomerisation in living cells. However, the majority of studies have relied largely on co-immunoprecipitation techniques, and there is still little direct information on the fraction of receptors existing as oligomers in intact cells.
Recruitment of β-catenin to cadherin-mediated intercellular adhesions is involved in myogenic induction
Authors: Goichberg Polina , Shtutman Michael , Ben-Ze’ev Avri , Geiger Benjamin 
Published: April 2021

ABSTRACT
Cadherin-mediated cell adhesion is involved in muscle differentiation from early stages of myogenic induction to late stages of myoblast interaction and fusion. β-Catenin is a major constituent of cadherin-based adherens junctions and also serves as a signal transduction molecule that regulates gene expression during development. In this study, we explored the involvement of β-catenin in myogenic differentiation. We show here that shortly after a switch from growth to differentiation medium, β-catenin translocates to cell-cell junctions and its levels increase. We further show that elevation of β-catenin levels, induced either by inhibition of its breakdown, using LiCl, or by its overexpression, suppresses the formation of adherens junctions, resulting in a sharp decline in myogenin expression and an arrest of myogenic progression. Recruitment of β-catenin to adherens junctions after transfection with N-cadherin restores myogenin expression in the transfected cells. These results suggest that increased cadherin-mediated adhesion and translocation of β-catenin to adherens junctions are involved in activating the early steps of myogenic differentiation.
The GTPase Rac1 selectively regulates Salmonella invasion at the apical plasma membrane of polarized epithelial cells
Authors: Criss Alison K. , Ahlgren Dawn M. , Jou Tzuu-Shuh , McCormick Beth A. , Casanova James E. 
Published: April 2021

ABSTRACT
The bacterial pathogen Salmonella typhimurium colonizes its animal hosts by inducing its internalization into intestinal epithelial cells. This process requires reorganization of the actin cytoskeleton of the apical plasma membrane into elaborate membrane ruffles that engulf the bacteria. Members of the Rho family of small GTPases are critical regulators of actin structure, and in nonpolarized cells, the GTPase Cdc42 has been shown to modulate Salmonella entry. Because the actin architecture of epithelial cells is organized differently from that of nonpolarized cells, we examined the role of two Rho family GTPases, Cdc42 and Rac1, in invasion of polarized monolayers of MDCK cells by S. typhimurium. Surprisingly, we found that endogenous Rac1, but not Cdc42, was activated during bacterial entry at the apical pole, and that this activation required the bacterial effector protein SopE. Furthermore, expression of dominant inhibitory Rac1 but not Cdc42 significantly inhibited apical internalization of Salmonella, indicating that Rac1 activation is integral to the bacterial entry process. In contrast, during basolateral internalization, both Cdc42 and Rac1 were activated; however, neither GTPase was required for entry. These findings, which differ significantly from previous observations in nonpolarized cells, indicate that the host cell signaling pathways activated by bacterial pathogens may vary with cell type, and in epithelial tissues may further differ between plasma membrane domains.
Analysis of R-Ras signalling pathways
Authors: Self Annette J. , Caron Emmanuelle , Paterson Hugh F. , Hall Alan 
Published: April 2021

ABSTRACT
R-Ras has a high degree of sequence homology to Ras and to other members of the Ras subfamily including Rap, TC21 and M-Ras. Activated versions of Ras and TC21 are highly transforming in a variety of cell lines and mutated forms of both proteins have been found in human tumours. R-Ras interacts with many of the same proteins as Ras and TC21, including c-Raf1, and can induce transformed foci, although this activity is weak compared to Ras and appears to be cell-type specific. Here, we have investigated R-Ras signalling pathways in a variety of cell types. We find that microinjection of activated R-Ras into quiescent fibroblasts stimulates cell cycle progression through G1 phase and subsequent DNA synthesis. However, unlike Ras, R-Ras does not activate the ERK MAP kinase pathway nor does it activate the JNK or p38/Mpk2 MAP kinase pathways. Microinjection of R-Ras into PC12 cells does not induce terminal differentiation, but instead causes extensive cell spreading, consistent with R-Ras having a role in integrin activation. Finally, in a macrophage cell line, R-Ras activates the αMβ2 integrin via the small GTPase Rap1, leading to phagocytosis of opsonized red blood cells, whereas Ras does not. These results indicate that R-Ras has an important role in the regulation of cell growth and adhesion, but that this is mediated through downstream signals distinct from those used by Ras.
The Tem1 small GTPase controls actomyosin and septin dynamics during cytokinesis
Authors: Lippincott John , Shannon Katie B. , Shou Wenying , Deshaies Raymond J. , Li Rong 
Published: April 2021

ABSTRACT
Cytokinesis in budding yeast involves an actomyosin-based ring which assembles in a multistepped fashion during the cell cycle and constricts during cytokinesis. In this report, we have investigated the structural and regulatory events that occur at the onset of cytokinesis. The septins, which form an hour-glass like structure during early stages of the cell cycle, undergo dynamic rearrangements prior to cell division: the hourglass structure splits into two separate rings. The contractile ring, localized between the septin double rings, immediately undergoes contraction. Septin ring splitting is independent of actomyosin ring contraction as it still occurs in mutants where contraction fails. We hypothesize that septin ring splitting may remove a structural barrier for actomyosin ring to contract. Because the Tem1 small GTPase (Tem1p) is required for the completion of mitosis, we investigated its role in regulating septin and actomyosin ring dynamics in the background of the net1-1 mutation, which bypasses the anaphase cell cycle arrest in Tem1-deficient cells. We show that Tem1p plays a specific role in cytokinesis in addition to its function in cell cycle progression. Tem1p is not required for the assembly of the actomyosin ring but controls actomyosin and septin dynamics during cytokinesis.
Cell-specific targeting of caveolin-1 to caveolae, secretory vesicles, cytoplasm or mitochondria
Authors: Li Wei-Ping , Liu Pingsheng , Pilcher Brian K. , Anderson Richard G. W. 
Published: April 2021

ABSTRACT
In commonly used tissue culture cells, caveolin-1 is embedded in caveolae membranes. It appears to reach this location after being cotranslationally inserted into ER membranes, processed in the Golgi and shipped to the cell surface. We now report that caveolae are not the preferred location for caveolin-1 in all cell types. Skeletal muscle cells and keratinocytes target caveolin-1 to the cytosol while in exocrine and endocrine cells it accumulates in the secretory pathway. We also found that airway epithelial cells accumulate caveolin-1 in modified mitochondria. The cytosolic and the secreted forms appear to be incorporated into a soluble, lipid complex. We conclude that caveolin-1 can be targeted to a variety of intracellular destinations, which suggests a novel mechanism for the intracellular traffic of this protein.