摘要:

AbstractFascin is an actin‐bundling protein that was first isolated from cytoplasmic extracts of sea urchin eggs [Kane, 1975:J. Cell Biol.66:305–315] and was the first bundling protein to be charactrized in vitro. Subsequent work has shown that fascin bundles actin filaments in fertilized egg microvilli and filopodia of phagocytic coelomocytes [Otto et al., 1980:Cell Motil.1:31–40; Otto and Bryan, 1981:Cell Motil.1:179–192]. Fifteen years later, the molecular cloning of sea urchin fascin [Bryan et al., 1993:Proc. Natl. Acad. Sci. U.S.A.90:9115–9119] has led to the identification and characterization of homologous proteins inDrosophila[Cant et al., 1994:J. Cell Biol.125:369–380],Xenopus[Holthuis et al., 1994:Biochim. Biophys. Acta.1219:184–188], rodents [Edwards et al., 1995:J. Biol. Chem.270:10764–10770], and humans [Duh et al., 1994:DNA Cell Biol.13:821–827; Mosialos et al., 1994:J. Virol.68:7320–7328] that bundle actin filaments into structures which stabilize cellular processes ranging from mechanosensory bristles to the filopodia of nerve growth cones. Fascin has emerged from relative obscurity as an exotic invertebrate egg protein to being recognized as a widely expressed protein found in a broad spectrum of tissues and organisms. This purpose of this review is to relate the early studies done on sea urchin and HeLa cell fascins to the recent molecular biology that defines a family of bundling proteins, and discuss the current state of knowledge regarding fascin structure and function. ©

ISSN:0886-1544    

DOI:10.1002/cm.970320102

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractCytoskeletal intermediate filaments (IFs) constitute a diverse family of proteins whose members are expressed in tissue‐specific patterns. Although vimentin IFs are normally restricted to mesenchyme, a variety of cell types express vimentin alone or together with cell‐specific IFs during growth, differentiation, and neoplasia. In this study, we have investigated the influence of increased vimentin expression on the simple epithelial cell phenotype. An expression vector encoding a human vimentin cDNA was transfected into the murine HR9 endoderm and F9 embryonal carcinoma cell lines, which serve as models for early extraembryonic epithelial differentiation. Stable clones that expressed varying levels of the human vimentin were characterized by immunofluorescence and biochemical analysis. A relatively high level of vimentin expression in HR9 and differentiated F9 epithelial cells resulted in aberrant vimentin structures with a co‐collapss of keratin K8/K18 filaments and lowered amounts of keratin protein. In F9 epithelial cells, the desmosomal proteins DP I/II did not appear to localize to cell surface desmosomes but rather co‐aggregated with the perturbed IFs. Although overall cell morphology was not dramatically altered, individual nuclei were distorted by excess intracellular vimentin. Furthermore, cell proliferation as well as the cell spreading response time were slowed. There appears to be a threshold effect regarding overall vimentin levels as cells that expressed lower amounts of the human vimentin exhibited no obvious structural nor biological effects. Our results demonstrate that wild‐type vimentin can act as a “mutant” protein when present at high intracellular levels, inducing a variety of phenotypic changes. © 1995 W

ISSN:0886-1544    

DOI:10.1002/cm.970320103

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractWe report the cloning and sequencing of genomic DNA encoding a cytoplasmic dynein heavy chain from the nematodeCaenorhabditis elegans.In a contiguous stretch of 35,103 bp of DNA from the left arm of linkage group I, we have found a gene that is predicted to encode a protein of 4,568 amino acids. This gene is composed of 15 exons and 14 relatively short introns, and it has significant homology of the other dynein heavy chains in the databases. The deduced molecular mass of the derived polypeptide is 512,624 Da. As with other dynein heavy chains that have been sequenced to date, it contains four GXXGXGK(S/T) motifs that form part of the consensus sequence for nucleotide triphosphate‐binding domains. Comparison of axonemal and cytoplasmic dynein heavy chains shows that regions of homology among all dyneins are clustered in the carboxyl terminal two‐thirds of the polypeptide, whereas the amino terminal one‐third of the heavy chains may contain domains that specify functions that differ between axonemal and cytoplasmic forms of the dynein heavy chain. © 1995 Wiley‐L

ISSN:0886-1544    

DOI:10.1002/cm.970320104

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractEntamoeba histolyticatrophozoites are pleiomorphic and highly motile cells. Although scarce fibrous material can be identified in the cytoplasm as elements of an organized cytoskeleton, clearly defined actin‐containing structures are formed at the sites of cell‐matrix contact upon the interaction of trophozoites with fibronectin (FN) and other extracellular matrix substrates. These structures are reminiscent of the adhesion plaques or focal contacts found in higher eukaryotic cells, where actin filament bundles insert into specialized regions of the plasma membrane and function as signal transduction organelles. Thus, the formation of adhesion plates in this parasitic ameba could be related to specific signaling responses involved in its invasive behavior. Here, we report the isolation of amebic adhesion plates and the results of their structural and molecular analyses. Filaments, with the characteristic diameter of F‐actin, radiating from an electrondense matrix, are the main feature. Actin is one of the main protein components of the plate; other proteins identified are a FN‐binding protein—previously reported as a “putative” FN receptor—the actin‐binding proteins myosin II, myosin I, α‐actinin, vinculin, and tropomyosin. The presence in the isolated plates of several proteases and protein kinases, in particular pp125FAK, is also demonstrated. Our results suggest that adhesion plates in amebas are dynamic membrane‐cytoskeletal complexes participating not only in the attachment to FN substrates but also providing the structural basis for their involvement in parasite locomotion and invasiveness.

ISSN:0886-1544    

DOI:10.1002/cm.970320105

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractAxonemal sliding involves both sliding velocity and the extent of sliding, that is how many doublets slide. It is clear that axonemes cannot beat if all doublets were to slide simultaneously, thus sliding extent is important. Using the turbidimetric assay of sliding disintegration ofTetrahymenaaxonemes, we examined the sliding extent and the effect of ADP, ATP, and ATP analogs on the sliding extent. Of course, ATP is necessary to produce sliding disintegration, but ATP alone did not produce extensive sliding disintegration. The addition of ADP allowed greater extent of sliding disintegration. The additions of higher ATP concentration even in the presence of ADP inhibited sliding disintegration. We also observed sliding disintegration using ribose‐modified ATP analogs, anthraniloylATP, and methylanthraniloylATP. The extent of sliding disintegration was proportional to the analog concentration. Thus in contrast to ATP, higher analog concentration was not inhibitory. These results indicate that high ATP concentration acts to inhibit the extent of sliding disintegration and that ADP relieves this inhibition. We propose a model in which the affinity of multiple cooperative active sites are regulated by binding of ATP or ADP to a regulatory site. This model provides a mechanism by which nucleotides regulate the extent of sliding necessary for effective axonemal bending. © 1995 Wiley‐Liss,

ISSN:0886-1544    

DOI:10.1002/cm.970320106

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractPhotoreceptor cells of vertebrate retinae are highly specialized ciliary cells. Their non‐motile ciliated structure is restricted to the so‐called connecting cilium at the joint between the light sensitive outer segment and the metabolically active inner segment. Extensive bidirectional intracellular transport between both segments is forced to occur through this tight connecting cilium. In the present study it is shown that the Ca2+‐binding, phospho‐protein centrin is present in mammalian retinae. Western blot and immunoprecipitation experiments reveal that anti‐centrin antibodies react with purified photoreceptor cell fractions of retinae in bands at a molecular weight of 20 kDa, the molecular weight of centrins found in other cells. Indirect immunofluorescence analysis of cryosections through retinae of different mammalian species show that centrin is present only in centrosomes and basal bodies but also more extensively at the linkage between the inner and the outer segment of the photoreceptor cells. Immunocytological studies on isolated rod cells and immunoelectron microscopy clearly demonstrate a unique presence of centrin in the connecting cilium of photoreceptor cells. High molecular identity between centrins in lower eukaryotes and mammals indicates that centrin may play a role in cellular motility and/or in microtubule severing in the mammalian retina. © 1995 Wiley

ISSN:0886-1544    

DOI:10.1002/cm.970320107

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractA specific peptide inhibitor of the cyclic AMP (cAMP)‐dependent protein kinase (PKI‐peptide) is a very effective inhibitor of the cAMP‐dependent activation of motility ofCionaspermatozoa, when the PKI‐peptide is present at the beginning of incubation of demembranated spermatozoa with cAMP and ATP. Under conditions where approximately 120 sec is required for full activation of motility, the window of sensitivity to the PKI‐peptide lasts for only 25–30 sec. Examination of sperm pellet proteins labeled with32P ATP during activation reveals a major 25 kDa phosphoprotein and 2 minor phosphoproteins whose phosphorylation is highly sensitive to inhibition by the PKI‐peptide and essentially complete during this early phase. These sperm proteins appear to be immediate substrates for cAMP‐dependent protein kinase, and phosphorylation of one or more of these appears to be required, but not sufficient, for activation of motility. The phosphorylation of other proteins is reduced or eliminated when PKI‐peptide is present at the beginning of incubation, but is unaffected by later addition of PKI‐peptide. Some of these substrates appear to be likely candidates for axonemal proteins that must be phosphorylated during the later stages of incubation in order to complete the activation process. This selection is based upon a high degree of inhibition by inclusion of PKI‐peptide or other inhibitors at the start of the incubation process, on near‐completion of their phosphorylation by the end of the 2 min incubation period required for activation of motility, and evidence that these proteins are phosphorylated during in vivo activation of motility. Although these observations suggest the presence of a second kinase activity that is upregulated by the initial activation of the cAMP‐dependent protein kinase, assays using exogenous substrates have not yet been able to identify such a kinase activit

ISSN:0886-1544    

DOI:10.1002/cm.970320108

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

ISSN:0886-1544    

DOI:10.1002/cm.970320101

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY