scholarly journals A novel photoactivatable tool for intermediate filament disruption indicates a role for keratin filaments in early embryogenesis

2018 ◽  
Author(s):  
Rucha Sanghvi-Shah ◽  
Shalaka Paranjpe ◽  
Jiyeon Baek ◽  
Radek Dobrowolski ◽  
Gregory F. Weber
Keyword(s):  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Cell Types ◽  
Model Systems ◽  
Intermediate Filament Proteins ◽  
Morphogenetic Movements ◽  
Cell Functions ◽  
Filament Network ◽  
Keratin Intermediate Filaments

AbstractThe significance of cytoplasmic intermediate filament proteins has previously been examined largely through various genetic approaches, including knockdown, knockout and transgenic overexpression. Few studies to date have attempted to examine the role of specifically the filamentous intermediate filament network in orchestrating various cell functions. To directly assess the role of the filamentous keratin intermediate filament network in regulation of cellular behavior, we created a PhotoActivatable disruptor of keratin Intermediate Filaments (PA-dIF). This genetically encoded construct consists of a peptide derived from the 2B2 region of Keratin 8 fused to the photosensitive LOV2 domain from Avena sativa phototropin-1. Upon 458 nm photoirradiation, PA-dIF disrupts keratin intermediate filaments in multiple species and cell types. Marked remodeling of the keratin intermediate filament network accompanies collective cellular morphogenetic movements that occur during gastrulation and neurulation in the Xenopus laevis frog embryo. Light-based activation of PA-dIF was able to disrupt keratin intermediate filaments in Xenopus cells and lead to tissue-specific disruption of morphogenetic processes. Altogether our data show a fundamental requirement for keratin intermediate filaments in orchestrating morphogenetic movements during early embryonic development that have yet to be revealed in other model systems. Moreover, our data validate the utility of a new genetically encoded photoactivatable tool for the disruption and examination of intermediate filaments.

Role of inflammation in the development of colorectal cancer

2020 ◽  
Vol 20 ◽  
Author(s):  
Sridhar Muthusami ◽  
R. Ileng Kumaran ◽  
Kokelavani Nampalli Babu ◽  
Sneha Krishnamoorthy ◽  
Akash Guruswamy ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Chronic Inflammation ◽  
Interleukin 1 ◽  
Cell Types ◽  
Model Systems ◽  
Cytokine Gene Expression ◽  
Necrosis Factor Alpha ◽  
Tnf Α ◽  
Proinflammatory Molecules

: Chronic inflammation can lead to the development of many diseases including cancer. Inflammatory bowel disease (IBD) that includes both ulcerative colitis (UC) and Crohn's disease (CD) are risk factors for the development of colorectal cancer (CRC). Many cytokines produced primarily by the gut immune cells either during or in response to localized inflammation in the colon and rectum are known to stimulate the complex interactions between the different cell types in the gut environment resulting in acute inflammation. Subsequently, chronic inflammation together with genetic and epigenetic changes has been shown to lead to the development and progression of CRC. Various cell types present in the colon such as enterocytes, Paneth cells, goblet cells and macrophages express receptors for inflammatory cytokines and respond to tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β), IL-6 and other cytokines. Among the several cytokines produced, TNF-α and IL-1β are the key proinflammatory molecules that play critical roles in the development of CRC. The current review is intended to consolidate the published findings to focus on the role of proinflammatory cytokines, namely TNF-α and IL-1β, on inflammation (and the altered immune response) in the gut, to better understand the development of CRC in IBD, using various experimental model systems, preclinical and clinical studies. Moreover, this review also highlights the current therapeutic strategies available (monotherapy and combination therapy), to alleviate the symptoms or treat inflammationassociated CRC by using monoclonal antibodies or aptamers to block proinflammatory molecules, inhibitors of tyrosine kinases in inflammatory signaling cascade, competitive inhibitors of proinflammatory molecules, and the nucleic acid drugs like small activating RNAs (saRNAs) or microRNA (miRNA) mimics to activate tumor suppressor or repress oncogene/proinflammatory cytokine gene expression.

Paranemin and the organization of desmin filament networks

2001 ◽  
Vol 114 (6) ◽  
pp. 1079-1089 ◽  
Author(s):  
S.C. Schweitzer ◽  
M.W. Klymkowsky ◽  
R.M. Bellin ◽  
R.M. Robson ◽  
Y. Capetanaki ◽  
...  
Keyword(s):  
Cell Lines ◽  
Intermediate Filament ◽  
Transgene Expression ◽  
De Novo ◽  
Muscle Cells ◽  
The Other ◽  
Intermediate Filament Proteins ◽  
Mouse Fibroblasts ◽  
Filament Networks ◽  
Filament Network

De novo expression of vimentin, GFAP or peripherin leads to the assembly of an extended intermediate filament network in intermediate filament-free SW13/cl.2 cells. Desmin, in contrast, does not form extended filament networks in either SW13/cl.2 or intermediate filament-free mouse fibroblasts. Rather, desmin formed short thickened filamentous structures and prominent spot-like cytoplasmic aggregates that were composed of densely packed 9–11 nm diameter filaments. Analysis of stably transfected cell lines indicates that the inability of desmin to form extended networks is not due to a difference in the level of transgene expression. Nestin, paranemin and synemin are large intermediate filament proteins that coassemble with desmin in muscle cells. Although each of these large intermediate filament proteins colocalized with desmin when coexpressed in SW-13 cells, expression of paranemin, but not synemin or nestin, led to the formation of an extended desmin network. A similar rescue of desmin network organization was observed when desmin was coexpressed with vimentin, which coassembles with desmin, or with keratins, which formed a distinct filament network. These studies demonstrate that desmin filaments differ in their organizational properties from the other vimentin-like intermediate filament proteins and appear to depend upon coassembly with paranemin, at least when they are expressed in non-muscle cells, in order to form an extended filament network.

Novel features of intermediate filament dynamics revealed by green fluorescent protein chimeras

1998 ◽  
Vol 111 (13) ◽  
pp. 1767-1778 ◽  
Author(s):  
C.L. Ho ◽  
J.L. Martys ◽  
A. Mikhailov ◽  
G.G. Gundersen ◽  
R.K. Liem
Keyword(s):  
Green Fluorescent Protein ◽  
Dynamic Behavior ◽  
Cell Lines ◽  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Fluorescent Protein ◽  
Nih3t3 Cells ◽  
Green Fluorescent ◽  
Filament Network ◽  
Perinuclear Area

In order to study the dynamic behavior of intermediate filament networks in living cells, we have prepared constructs fusing green fluorescent protein to intermediate filament proteins. Vimentin fused to green fluorescent protein labeled the endogenous intermediate filament network. We generated stable SW13 and NIH3T3 cell lines that express an enhanced green fluorescent protein fused to the N-terminus of full-length vimentin. We were able to observe the dynamic behavior of the intermediate filament network in these cells for periods as long as 4 hours (images acquired every 2 minutes). In both cell lines, the vimentin network constantly moves in a wavy manner. In the NIH3T3 cells, we observed extension of individual vimentin filaments at the edge of the cell. This movement is dependent on microtubules, since the addition of nocodazole stopped the extension of the intermediate filaments. Injection of anti-IFA causes the redistribution or ‘collapse’ of intermediate filaments. We injected anti-IFA antibodies into NIH3T3 cells stably expressing green fluorescent protein fused to vimentin and found that individual intermediate filaments move slowly towards the perinuclear area without obvious disassembly. These results demonstrate that individual intermediate filaments are translocated during the collapse, rather than undergoing disassembly-induced redistribution. Injections of tubulin antibodies disrupt the interactions between intermediate filaments and stable microtubules and cause the collapse of the vimentin network showing that these interactions play an important role in keeping the intermediate filament network extended. The nocodazole inhibition of intermediate filament extension and the anti-IFA microinjection experiments are consistent with a model in which intermediate filaments exhibit an extended distribution when tethered to microtubules, but are translocated to the perinuclear area when these connections are severed.

Interactions between peripherin and neurofilaments in cultured cells: disruption of peripherin assembly by the NF-M and NF-H subunits

1999 ◽  
Vol 77 (1) ◽  
pp. 41-45 ◽  
Author(s):  
Jean-Martin Beaulieu ◽  
Janice Robertson ◽  
Jean-Pierre Julien
Keyword(s):  
Nervous System ◽  
Peripheral Nervous System ◽  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Cultured Cells ◽  
Intermediate Filament Protein ◽  
Physiological Conditions ◽  
Filament Protein ◽  
Filament Network ◽  
Filament Type

Neurofilaments are the principal intermediate filament type expressed by neurons. They are formed by the co-assembly of three subunits: NF-L, NF-M, and NF-H. Peripherin is another intermediate filament protein expressed mostly in neurons of the peripheral nervous system. In contrast to neurofilaments, peripherin can self-assemble to establish an intermediate filament network in cultured cells. The co-expression of neurofilaments and peripherin is found mainly during development and regeneration. We used SW13 cells devoid of endogenous cytoplasmic intermediate filaments to assess the exact assembly characteristics of peripherin with each neurofilament subunit. Our results demonstrate that peripherin can assemble with NF-L. In contrast, the co-expression of peripherin with the large neurofilament subunits interferes with peripherin assembly. These results confirm the existence of interactions between peripherin and neurofilaments in physiological conditions. Moreover, they suggest that perturbations in the stoichiometry of neurofilaments can have an impact on peripherin assembly in vivo.Key words: peripherin, neurofilament, SW13 cells, intermediate filament.

scholarly journals Interaction of Theiler’s Virus with Intermediate Filaments of Infected Cells

1998 ◽  
Vol 72 (12) ◽  
pp. 9553-9560 ◽  
Author(s):  
Patrick Nédellec ◽  
Patrick Vicart ◽  
Christine Laurent-Winter ◽  
Cécile Martinat ◽  
Marie-Christine Prévost ◽  
...  
Keyword(s):  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Close Contact ◽  
Theiler's Virus ◽  
Virus Particles ◽  
Host Cell Proteins ◽  
Encephalomyelitis Virus ◽  
Infected Cells ◽  
Main Components ◽  
Filament Network

ABSTRACT Theiler’s murine encephalomyelitis virus is a neurotropic murine picornavirus which replicates permissively and causes a cytopathic effect in the BHK-21 cell line. We examined the interactions between the GDVII and DA strains of Theiler’s virus and BHK-21 host cell proteins in a virus overlay assay. We observed binding of the virions to two proteins of approximately 60 kDa. These proteins were microsequenced and identified as desmin and vimentin, two main components of the intermediate filament network. The association between desmin or vimentin and virions was demonstrated by immunoprecipitation. Anti-desmin and anti-vimentin monoclonal antibodies precipitated GDVII or DA virions from extracts of infected BHK-21 cells. The intracellular distributions of virions and of the desmin and vimentin intermediate filaments of BHK-21 cells were investigated by two-color immunofluorescence confocal microscopy. Following infection, the intermediate filament network was rearranged into a shell-like structure which surrounded a viral inclusion. Finally, close contact between GDVII virus particles and 10-nm intermediate filaments was observed by electron microscopy.

Peripherin assembles into homopolymers in SW13 cells

1995 ◽  
Vol 108 (10) ◽  
pp. 3279-3284 ◽  
Author(s):  
C. Cui ◽  
P.J. Stambrook ◽  
L.M. Parysek
Keyword(s):  
Intermediate Filament ◽  
Network Formation ◽  
Full Length ◽  
Intermediate Filament Proteins ◽  
Filament Assembly ◽  
Mature Neurons ◽  
Carboxyl Terminal ◽  
Relationship Of ◽  
Filament Network ◽  
Carboxyl Tail

The properties of full-length and mutant peripherins were studied in intermediate filament-less SW13 cells to define regions of peripherin that are essential for initiation of filament assembly. A full-length rat peripherin gene transfected into SW13 cells resulted in filament formation, consistent with the close structural relationship of peripherin to other type III intermediate filament proteins that readily form homopolymers. Translation of full-length rat peripherin is initiated predominantly at the second of two inframe AUGs. Deletions within the amino terminus of wild-type peripherin abolished its ability to form filaments in SW13 cells. In contrast, deletion of the entire carboxyl-terminal tail of peripherin did not affect its ability to form filamentous arrays in transfected SW13 cells. These results indicate that, of the intermediate filament proteins that are expressed in mature neurons, only peripherin and alpha-internexin are capable of making homopolymer intermediate filaments. In addition, mutations of the carboxyl tail of peripherin generally do not interfere with filament network formation.

scholarly journals Assembly of type IV neuronal intermediate filaments in nonneuronal cells in the absence of preexisting cytoplasmic intermediate filaments

1993 ◽  
Vol 122 (6) ◽  
pp. 1323-1335 ◽  
Author(s):  
GY Ching ◽  
RK Liem
Keyword(s):  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Cultured Cells ◽  
Neuronal Cell ◽  
Intermediate Filament Protein ◽  
Amino Acid Residues ◽  
Intermediate Filament Proteins ◽  
Transfected Cells ◽  
Type Iv

We report here on the in vivo assembly of alpha-internexin, a type IV neuronal intermediate filament protein, in transfected cultured cells, comparing its assembly properties with those of the neurofilament triplet proteins (NF-L, NF-M, and NF-H). Like the neurofilament triplet proteins, alpha-internexin coassembles with vimentin into filaments. To study the assembly characteristics of these proteins in the absence of a preexisting filament network, transient transfection experiments were performed with a non-neuronal cell line lacking cytoplasmic intermediate filaments. The results showed that only alpha-internexin was able to self-assemble into extensive filamentous networks. In contrast, the neurofilament triplet proteins were incapable of homopolymeric assembly into filamentous arrays in vivo. NF-L coassembled with either NF-M or NF-H into filamentous structures in the transfected cells, but NF-M could not form filaments with NF-H. alpha-internexin could coassemble with each of the neurofilament triplet proteins in the transfected cells to form filaments. When all but 2 and 10 amino acid residues were removed from the tail domains of NF-L and NF-M, respectively, the resulting NF-L and NF-M deletion mutants retained the ability to coassemble with alpha-internexin into filamentous networks. These mutants were also capable of forming filaments with other wild-type neurofilament triplet protein subunits. These results suggest that the tail domains of NF-L and NF-M are dispensable for normal coassembly of each of these proteins with other type IV intermediate filament proteins to form filaments.

Molecular analysis of intermediate filament cytoskeleton--a putative load-bearing structure

1984 ◽  
Vol 246 (4) ◽  
pp. H566-H572 ◽  
Author(s):  
M. G. Price
Keyword(s):  
Skeletal Muscle ◽  
Molecular Analysis ◽  
Intermediate Filaments ◽  
Intermediate Filament ◽  
Direct Evidence ◽  
Two Dimensional ◽  
Myocardial Cells ◽  
Intermediate Filament Proteins ◽  
Bovine Myocardium ◽  
Bearing Structure

Myocardial cells contain a cytoskeleton of intermediate filaments connecting the myofibrils. The present molecular analysis of the myocardial cytoskeleton was designed to identify the intermediate filament proteins and examine their assembly properties. The intermediate filament proteins desmin and vimentin were isolated from adult bovine myocardium by sequential extraction, urea solubilization, and chromatography on hydroxylapatite and DEAE columns. Desmin was obtained virtually pure in one peak and in a mixture of desmin and vimentin in the trailing fractions. Intermediate filaments of different morphologies polymerized in the desmin and the desmin-vimentin fractions. Isolated myocardial desmin occurs as three isozymes and isolated myocardial vimentin as two isozymes, which co-migrate on two-dimensional gels with corresponding isozymes from bovine skeletal and smooth muscle. Polypeptides of 200,000 and 220,000 daltons that fractionate with myocardial desmin and vimentin are also present in cytoskeletons of smooth and skeletal muscle. The results provide direct evidence that myocardial desmin can assemble to form intermediate filaments, suggesting that desmin is the major component of the cytoskeletal filaments in cardiomyocytes.

scholarly journals Role of MSC in the Tumor Microenvironment

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2107 ◽  
Author(s):  
Ralf Hass
Keyword(s):  
Tumor Microenvironment ◽  
Cancer Cells ◽  
Tumor Development ◽  
Tumor Stroma ◽  
Cell Types ◽  
Cellular Interactions ◽  
Cell Functions ◽  
Metastatic Behavior ◽  
Stem Cell Niches

The tumor microenvironment represents a dynamically composed matrix in which tissue-associated cancer cells are embedded together with a variety of further cell types to form a more or less separate organ-like structure. Constantly mutual interactions between cells of the tumor microenvironment promote continuous restructuring and growth in the tumor. A distinct organization of the tumor stroma also facilitates the formation of transient cancer stem cell niches, thereby contributing to progressive and dynamic tumor development. An important but heterogeneous mixture of cells that communicates among the cancer cells and the different tumor-associated cell types is represented by mesenchymal stroma-/stem-like cells (MSC). Following recruitment to tumor sites, MSC can change their functionalities, adapt to the tumor’s metabolism, undergo differentiation and synergize with cancer cells. Vice versa, cancer cells can alter therapeutic sensitivities and change metastatic behavior depending on the type and intensity of this MSC crosstalk. Thus, close cellular interactions between MSC and cancer cells can eventually promote cell fusion by forming new cancer hybrid cells. Consequently, newly acquired cancer cell functions or new hybrid cancer populations enlarge the plasticity of the tumor and counteract successful interventional strategies. The present review article highlights some important features of MSC within the tumor stroma.

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