scholarly journals Mobility of filamentous actin in living cytoplasm.

1987 ◽  
Vol 105 (6) ◽  
pp. 2811-2816 ◽  
Author(s):  
Y L Wang
Keyword(s):  
Cell Division ◽  
Cultured Cells ◽  
Stress Fibers ◽  
Detectable Effect ◽  
Filamentous Actin ◽  
Cellular Functions ◽  
Contractile Ring ◽  
Rhodamine Phalloidin ◽  
Affinity Probe ◽  
Dividing Cells

Filamentous actin in living cultured cells was labeled by microinjecting trace amounts of rhodamine-phalloidin (rh-pha) as a specific, high-affinity probe. The microinjection caused no detectable effect on cell morphology or cell division. The distribution of rh-pha-labeled filaments was then examined in dividing cells using image-intensified fluorescence microscopy, and the exchangeability of labeled filaments along stress fibers was studied during interphase using fluorescence recovery after photobleaching. rh-pha showed a rapid concentration at the contractile ring during cell division. In addition, recovery of fluorescence after photobleaching occurred along stress fibers with a halftime as short as 8 min. These observations suggest that at least some actin filaments undergo continuous movement and reorganization in living cells. This dynamic process may play an important role in various cellular functions.

Enhancement of myosin II/actin turnover at the contractile ring induces slower furrowing in dividing HeLa cells

2011 ◽  
Vol 435 (3) ◽  
pp. 569-576 ◽  
Author(s):  
Tomo Kondo ◽  
Kozue Hamao ◽  
Keiju Kamijo ◽  
Hiroshi Kimura ◽  
Makiko Morita ◽  
...  
Keyword(s):  
Hela Cells ◽  
Fluorescence Recovery After Photobleaching ◽  
Kinase Inhibitor ◽  
Myosin Ii ◽  
Rho Kinase ◽  
Filamentous Actin ◽  
Contractile Ring ◽  
Rho Kinase Inhibitor ◽  
Actin Turnover ◽  
Dividing Cells

Myosin II ATPase activity is enhanced by the phosphorylation of MRLC (myosin II regulatory light chain) in non-muscle cells. It is well known that pMRLC (phosphorylated MRLC) co-localizes with F-actin (filamentous actin) in the CR (contractile ring) of dividing cells. Recently, we reported that HeLa cells expressing non-phosphorylatable MRLC show a delay in the speed of furrow ingression, suggesting that pMRLC plays an important role in the control of furrow ingression. However, it is still unclear how pMRLC regulates myosin II and F-actin at the CR to control furrow ingression during cytokinesis. In the present study, to clarify the roles of pMRLC, we measured the turnover of myosin II and actin at the CR in dividing HeLa cells expressing fluorescent-tagged MRLCs and actin by FRAP (fluorescence recovery after photobleaching). A myosin II inhibitor, blebbistatin, caused an enhancement of the turnover of MRLC and actin at the CR, which induced a delay in furrow ingression. Furthermore, only non-phosphorylatable MRLC and a Rho-kinase inhibitor, Y-27632, accelerated the turnover of MRLC and actin at the CR. Interestingly, the effect of Y-27632 was cancelled in the cell expressing phosphomimic MRLCs. Taken together, these results reveal that pMRLC reduces the turnover of myosin II and also actin at the CR. In conclusion, we show that the enhancement of myosin II and actin turnover at the CR induced slower furrowing in dividing HeLa cells.

scholarly journals Filamentous Actin Disruption and Diminished Inositol Phosphate Response in Gingival Fibroblasts Caused byTreponema denticola

1998 ◽  
Vol 66 (2) ◽  
pp. 696-702 ◽  
Author(s):  
Po Fong Yang ◽  
Meja Song ◽  
David A. Grove ◽  
Richard P. Ellen
Keyword(s):  
Time Course ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Actin Stress Fiber ◽  
Gingival Fibroblasts ◽  
Filamentous Actin ◽  
Rhodamine Phalloidin ◽  
Major Surface

ABSTRACT Previous reports have shown that Treponema denticolacauses rearrangement of filamentous actin (F-actin) in human gingival fibroblasts (HGF). The purpose of this investigation was to determine the effect of T. denticola on the generation of inositol phosphates (IPs) in relation to a time course for F-actin disruption in HGF. Cultured HGF were exposed to washed cells of T. denticola ATCC 35405 for 140 min. Changes in the fluorescence intensity of rhodamine-phalloidin-labeled F-actin in serial optical sections of single HGF were quantified by confocal microscopy image analysis. The percentage of cells with stress fiber disruption was also determined by fluorescence microscopy. Challenge with T. denticola caused a significant reduction in F-actin within the first hour, especially at the expense of F-actin in the ventral third of the cells, and a significant increase in the percentage of HGF with altered stress fiber patterns. Significant concentration-dependent disruption of stress fibers was also caused by HGF exposure to a Triton X-100 extract of T. denticola outer membrane (OM). IPs were measured by a radiotracer assay based on the incorporation ofmyo-[3H]inositol into IPs in HGF incubated with LiCl to inhibit endogenous phosphatases. HGF challenge with several strains of T. denticola and the OM extract ofT. denticola ATCC 35405 resulted in a diminished accumulation of radiolabeled IPs relative to both 15 and 1% fetal bovine serum, which served as strongly positive and background control agonists, respectively. The significantly diminished IP response toT. denticola ATCC 35405 occurred within 60 min, concomitant with significant reduction of total F-actin and disruption of stress fibers. Pretreatment with the proteinase inhibitor phenylmethylsulfonyl fluoride, which had previously been found to block T. denticola’s degradation of endogenous fibronectin and detachment of HGF from the extracellular matrix, had little effect on F-actin stress fiber disruption and the IP response. Therefore, in addition to its major surface chymotrypsin-like properties, T. denticola expresses cytopathogenic activities that diminish the generation of IPs during the time course associated with significant cytoskeletal disruption in fibroblasts.

scholarly journals Diphosphorylated MRLC is Required for Organization of Stress Fibers in Interphase Cells and the Contractile Ring in Dividing Cells

2001 ◽  
Vol 26 (6) ◽  
pp. 677-683 ◽  
Author(s):  
Takahiro Iwasaki ◽  
Maki Murata-Hori ◽  
Shu Ishitobi ◽  
Hiroshi Hosoya
Keyword(s):  
Stress Fibers ◽  
Contractile Ring ◽  
Interphase Cells ◽  
Dividing Cells

Polyamine deficiency causes reorganization of F-actin and tropomyosin in IEC-6 cells

1994 ◽  
Vol 267 (3) ◽  
pp. C715-C722 ◽  
Author(s):  
S. A. McCormack ◽  
J. Y. Wang ◽  
L. R. Johnson
Keyword(s):  
Actin Filaments ◽  
Cytoskeletal Proteins ◽  
Stress Fibers ◽  
Filamentous Actin ◽  
Rhodamine Phalloidin ◽  
Tropomyosin Isoforms ◽  
Actin Cortex ◽  
Beta Actin ◽  
Actin Mrna ◽  
As Stress

In earlier work we have shown that polyamine-deficient IEC-6 cells lose most of their ability to migrate. In this report we describe the effect of polyamine deficiency on the cytoskeleton of migrating IEC-6 cells. Cells were grown on cover slips for 4 days. One-third of the monolayer was removed, and the remainder was incubated for 6 h. The monolayers were fixed and stained with rhodamine phalloidin for actin filaments and by immunocytochemistry for tropomyosin. In control cells, actin filaments were found as stress fibers traversing the cell, in a thin actin cortex often visible on only one edge of the cell, and in fine fibers extending into the lamellipodia. Tropomyosin was found in the same distribution. A Western blot showed that tropomyosin was present as 35- and 37-kDa isoforms. In polyamine-deficient cells, actin stress fibers were less dense, whereas the actin cortex was greatly increased in density and lamellipodia were less extensive. Tropomyosin distribution was similar and included a 30-kDa isoform not seen previously. In spite of the obvious changes in the distribution of these cytoskeletal proteins, the concentrations of filamentous actin, beta-actin mRNA, and the higher molecular weight tropomyosin isoforms did not change. In all cases the addition of putrescine to polyamine-deficient cells prevented the changes described. We conclude that polyamines are essential for migration in this system because of their effects on the organization of cytoskeletal actin, tropomyosin, and perhaps other proteins as well.

scholarly journals Comparison of purified anti-actin and fluorescent-heavy meromyosin staining patterns in dividing cells.

1979 ◽  
Vol 80 (3) ◽  
pp. 509-520 ◽  
Author(s):  
I M Herman ◽  
T D Pollard
Keyword(s):  
Cultured Cells ◽  
Direct Method ◽  
Fluorescent Antibody ◽  
Stress Fibers ◽  
Fluorescent Antibody Technique ◽  
Heavy Meromyosin ◽  
Antibody Technique ◽  
Nonmuscle Cells ◽  
Dividing Cells ◽  
Fluorescent Heavy Meromyosin

We purified actin antibodies by affinity chromatography from the serum of rabbits immunized with glutaraldehyde-fixed chicken gizzard actin filaments and used this anti-actin to localize actin in myofibrils and fixed cultured cells at each stage of the cell cycle. By double immunodiffusion the anti-actin reacted with both smooth and skeletal muscle actin. Several blocking and absorption experiments demonstrated that the antibodies also bound specifically to actin in nonmuscle cells. The same structures stained using either the direct or the indirect fluorescent antibody technique; and, while the indirect method was more sensitive, the direct method was superior because there was no detectable nonspecific staining. As expected, anti-actin stained the I-band of myofibrils. It also stained stress fibers and membrane ruffles in HeLa cells. Some PtK-2 cells have straight stress fibers which stained with anti-actin, but in confluent cultures all PtK-2 cells have, instead, sinuous phase-dense fibers which stained with antibody. At prophase the whole cytoplasm stained uniformly with anti-actin. During metaphase and anaphase, anti-actin staining was concentrated diffusely in the mitotic spindle. In contrast, fluorescent heavy meromyosin stained discrete fine spindle fibers in these fixed cells. During cytokinesis, anti-actin stained the whole cytoplasm uniformly and was not concentrated in the cleavage furrow.

scholarly journals On the embryonic cell division beyond the contractile ring mechanism: experimental and computational investigation of effects of vitelline confinement, temperature and egg size

PeerJ ◽  
2015 ◽  
Vol 3 ◽  
pp. e1490 ◽  
Author(s):  
Evgeny Gladilin ◽  
Roland Eils ◽  
Leonid Peshkin
Keyword(s):  
Cell Division ◽  
Egg Size ◽  
Early Embryo ◽  
Embryonic Cell ◽  
Vitelline Membrane ◽  
Imaging Data ◽  
Contractile Ring ◽  
Dividing Cells ◽  
Vitelline Envelope ◽  
Effects Of Temperature

Embryonic cell division is a mechanical process which is predominantly driven by contraction of the cleavage furrow and response of the remaining cellular matter. While most previous studies focused on contractile ring mechanisms of cytokinesis, effects of environmental factors such as pericellular vitelline membrane and temperature on the mechanics of dividing cells were rarely studied. Here, we apply a model-based analysis to the time-lapse imaging data of two species (Saccoglossus kowalevskiiandXenopus laevis) with relatively large eggs, with the goal of revealing the effects of temperature and vitelline envelope on the mechanics of the first embryonic cell division. We constructed a numerical model of cytokinesis to estimate the effects of vitelline confinement on cellular deformation and to predict deformation of cellular contours. We used the deviations of our computational predictions from experimentally observed cell elongation to adjust variable parameters of the contractile ring model and to quantify the contribution of other factors (constitutive cell properties, spindle polarization) that may influence the mechanics and shape of dividing cells. We find that temperature affects the size and rate of dilatation of the vitelline membrane surrounding fertilized eggs and show that in native (not artificially devitellinized) egg cells the effects of temperature and vitelline envelope on mechanics of cell division are tightly interlinked. In particular, our results support the view that vitelline membrane fulfills an important role of micromechanical environment around the early embryo the absence or improper function of which under moderately elevated temperature impairs normal development. Furthermore, our findings suggest the existence of scale-dependent mechanisms that contribute to cytokinesis in species with different egg size, and challenge the view of mechanics of embryonic cell division as a scale-independent phenomenon.

scholarly journals THz irradiation inhibits cell division by affecting actin dynamics

2021 ◽  
Author(s):  
shota yamazaki ◽  
Yuya Ueno ◽  
Ryosuke Hosoki ◽  
Takanori Saito ◽  
Toshitaka Idehara ◽  
...  
Keyword(s):  
Cell Division ◽  
Actin Polymerization ◽  
Time Lapse ◽  
Actin Dynamics ◽  
Filamentous Actin ◽  
Contractile Ring ◽  
Biological Phenomena ◽  
Underlying Mechanisms

Biological phenomena induced by terahertz (THz) irradiation are described in recent reports, but underlying mechanisms, structural and dynamical change of specific molecules are still unclear. In this paper, we performed time-lapse morphological analysis of human cells and found that THz irradiation halts cell division at cytokinesis. At the end of cytokinesis, the contractile ring, which consists of filamentous actin (F-actin), needs to disappear; however, it remained for 1 hour under THz irradiation. Induction of the functional structures of F-actin was also observed in interphase cells. Similar phenomena were also observed under chemical treatment (jasplakinolide), indicating that THz irradiation assists actin polymerization. We previously reported that THz irradiation enhances the polymerization of purified actin in vitro; our current work shows that it increases cytoplasmic F-actin in vivo. Thus, we identified one of the key biomechanisms affected by THz waves.

scholarly journals THz irradiation inhibits cell division by affecting actin dynamics

PLoS ONE ◽  
2021 ◽  
Vol 16 (8) ◽  
pp. e0248381
Author(s):  
Shota Yamazaki ◽  
Yuya Ueno ◽  
Ryosuke Hosoki ◽  
Takanori Saito ◽  
Toshitaka Idehara ◽  
...  
Keyword(s):  
Cell Division ◽  
Actin Polymerization ◽  
Time Lapse ◽  
Actin Dynamics ◽  
Filamentous Actin ◽  
Contractile Ring ◽  
Biological Phenomena ◽  
Underlying Mechanisms

Biological phenomena induced by terahertz (THz) irradiation are described in recent reports, but underlying mechanisms, structural and dynamical change of specific molecules are still unclear. In this paper, we performed time-lapse morphological analysis of human cells and found that THz irradiation halts cell division at cytokinesis. At the end of cytokinesis, the contractile ring, which consists of filamentous actin (F-actin), needs to disappear; however, it remained for 1 hour under THz irradiation. Induction of the functional structures of F-actin was also observed in interphase cells. Similar phenomena were also observed under chemical treatment (jasplakinolide), indicating that THz irradiation assists actin polymerization. We previously reported that THz irradiation enhances the polymerization of purified actin in vitro; our current work shows that it increases cytoplasmic F-actin in vivo. Thus, we identified one of the key biomechanisms affected by THz waves.

scholarly journals Cortical PAR polarity proteins promote robust cytokinesis during asymmetric cell division

2016 ◽  
Vol 212 (1) ◽  
pp. 39-49 ◽  
Author(s):  
Shawn N. Jordan ◽  
Tim Davies ◽  
Yelena Zhuravlev ◽  
Julien Dumont ◽  
Mimi Shirasu-Hiza ◽  
...  
Keyword(s):  
Cell Division ◽  
Cell Fate ◽  
Asymmetric Cell Division ◽  
Animal Cell ◽  
Cell Formation ◽  
Active Role ◽  
Filamentous Actin ◽  
Contractile Ring ◽  
Par Proteins ◽  
Cell Divisions

Cytokinesis, the physical division of one cell into two, is thought to be fundamentally similar in most animal cell divisions and driven by the constriction of a contractile ring positioned and controlled solely by the mitotic spindle. During asymmetric cell divisions, the core polarity machinery (partitioning defective [PAR] proteins) controls the unequal inheritance of key cell fate determinants. Here, we show that in asymmetrically dividing Caenorhabditis elegans embryos, the cortical PAR proteins (including the small guanosine triphosphatase CDC-42) have an active role in regulating recruitment of a critical component of the contractile ring, filamentous actin (F-actin). We found that the cortical PAR proteins are required for the retention of anillin and septin in the anterior pole, which are cytokinesis proteins that our genetic data suggest act as inhibitors of F-actin at the contractile ring. Collectively, our results suggest that the cortical PAR proteins coordinate the establishment of cell polarity with the physical process of cytokinesis during asymmetric cell division to ensure the fidelity of daughter cell formation.

Confocal microscopy of the cytoskeleton in living plant cells following microinjection of fluorescent probes

1993 ◽  
Vol 51 ◽  
pp. 130-131
Author(s):  
Ann Cleary
Keyword(s):  
Cell Division ◽  
Fluorescent Probes ◽  
Actin Filaments ◽  
Intracellular Transport ◽  
Cell Structure ◽  
Plant Cells ◽  
Cell Plate ◽  
Cell Cortex ◽  
Living Plant ◽  
Rhodamine Phalloidin

Microinjection of fluorescent probes into living plant cells reveals new aspects of cell structure and function. Microtubules and actin filaments are dynamic components of the cytoskeleton and are involved in cell growth, division and intracellular transport. To date, cytoskeletal probes used in microinjection studies have included rhodamine-phalloidin for labelling actin filaments and fluorescently labelled animal tubulin for incorporation into microtubules. From a recent study of Tradescantia stamen hair cells it appears that actin may have a role in defining the plane of cell division. Unlike microtubules, actin is present in the cell cortex and delimits the division site throughout mitosis. Herein, I shall describe actin, its arrangement and putative role in cell plate placement, in another material, living cells of Tradescantia leaf epidermis.The epidermis is peeled from the abaxial surface of young leaves usually without disruption to cytoplasmic streaming or cell division. The peel is stuck to the base of a well slide using 0.1% polyethylenimine and bathed in a solution of 1% mannitol +/− 1 mM probenecid.

Sign in / Sign up

Export Citation Format

Share Document