bluuebunny/crossref_metadata_2025_split huggingface dataset:
Distribution of desmosomal proteins in F9 embryonal carcinoma cells and epithelial cell derivatives
Authors: Trevor Katrina T. , Steben Lara S. 
Published: April 2021

ABSTRACT
In diverse epithelia, cytoskeletal keratin intermediate filaments (IFs) associate with the cytoplasmic face of intercelluar junctional desmosomes. The processes underlying desmosome formation and keratin IF interactions remain unclear. We have examined F9 embryonal carcinoma (EC) cell differentiation as a model for embryonic development of epithelial surface desmosomes. As determined by immunofluorescence microscopy and biochemical protein techniques, F9 EC cells, which lack surface desmosomes and keratin IFs, express the desmosomal proteins desmoplakins I and II (DP I/II), desmoglein I (DG I) and plakoglobin (PK). DP I/II are present at low levels and are relatively soluble in buffer containing Triton X-100. Immunofluorescence localizes DP l/n to the juxtanuclear, centrosomal region. Species of DG I and PK are detected in both the Triton X-100-soluble and-insoluble protein fractions. DG I appears dispersed throughout the cell while PK resides at cell-cell boundaries. In epithelial cell cultures induced by retinoic acid (RA) treatment, each of the desmosomal proteins is organized into punctate desmosome-like structures with the appearance of simple epithelial K8/K18 IFs. The steady-state levels of DP l/ll and PK increase with a partitioning of the majority of the desmosomal components into the insoluble fraction. In epithelial cells which lack distinct surface desmosomes, an intracellular association of keratin bundles with DP I/II is observed, suggesting that keratin filaments may facilitate the translocation of these desmosomal components to the cell surface. Parietal endoderm-like cells, derived by treatment with RA and dibutyryl cAMP, are analogous to F9 EC cells in that the cells express desmosomal components and do not display surface desmosomes. Moreover, K8 and K18 do not form distinct filaments, and the protein and RNA levels of K8 are low relative to epithelial cells induced by RA alone. The F9 system appears to be a relevant model for studies of desmosome assembly and the potential interactions of desmosomal proteins and keratin IFs in embryonic epithelial cell types.
The involvement of discrete genome regions in post-mitotic chromosome decondensation and in G1 timing in Allium cepa L. meristematic cells
Authors: Gonzalez-Fernandez A. , Sans J. , Aller P. , Torre C. De La 
Published: April 2021

ABSTRACT
The role of DNA regions replicated at different times during the S period in the process of chromatin decondensation that takes place in the next G1 period, as well as in the timing of the G1 to S transition, was analyzed in synchronous populations of cells in Allium cepa L. root meristems. For this analysis, DNA bromosubstitution (10 −7 M 5-bromo-2 ′-deoxyuridine feeding) was carried out at similar times corresponding to the first, middle and last thirds of the S period prior to telophase when anoxic 313 nm irradiation was carried out.
Evaluation, after Feulgen staining, of the chromatin pattern corresponding to G0 and G1 phenotypes in these synchronous cells showed that chromatin decondensation after mitosis appeared to be under the control of a double negative/positive regulatory mechanism related, respectively, to early and late replicating sequences.
Thus, such decondensation was favoured when bromosubstitution took place during the first part of the S period, but delayed when bromosubstitution took place during the third part or the whole of the S period. Nevertheless, bromosubstitution at both the early and late stages of the S period delayed the timing of the G1 to S transition. In this way, the initiation of replication appears to be positively controlled by the function of early replicating DNA. Finally, the DNA sequences replicated in the middle third of the S period did not modify the decondensing capacity of the chromatin in G1 or the timing of the G1 to S transition.
Dolichol delays G1-arrest for one cell cycle in human fibroblasts subjected to depletion of serum or mevalonate
Authors: Larsson Olle , Wejde Johan 
Published: April 2021

ABSTRACT
It is well-established that some product(s) or metabolite(s) of mevalonate is (are) critical for growth of mammalian cells. In the search for this (these) compound(s) it seems meaningful to distinguish between compounds needed for cell cycle progression in proliferating cells and compounds needed for growth activation of arrested cells. By using time-lapse video recording we have studied the possible regulatory role of cholesterol, dolichol and mevalonate in the cell cycle of human diploid fibroblasts (HDF). HDF, which are serum-dependent, were rapidly growth-arrested in the first part of G1 upon removal of serum factors. They also responded to mevinolin (an HMG CoA reductase inhibitor) by a similar G1-block, indicating that a mevalonate-derived product is involved in the G1-located cell cycle control of HDF. Interestingly, dolichol counteracted the G1-block caused by both types of treatment. Hence, the early G1-cells could traverse the remainder of the cell cycle and divide despite depletion of serum or mevalonate. We also demonstrated that addition of dolichol resulted in a significant decrease in the rate of protein degradation. This protein stabilizing effect may constitute the mechanism by which dolichol delays the G1-arrest of HDF.
Cell phenotype, binding affinity and promoter structure modulate transactivation by HNF1 and LAP
Authors: Angrand Pierre-Olivier , Rousset Jean-Pierre , Weiss Mary C. 
Published: April 2021

ABSTRACT
To evaluate the importance of the transcription factors known to bind to the albumin promoter as well as the parameters involved in their activity, we have used cotransfections with an albumin promoter-cat plasmid combined with expression vectors driving the expression of cDNAs coding for liver-enriched factors known to interact with this promoter. We describe the characteristics of a set of clones of hepatic origin: well differentiated, partial variants or pleiotropic dedifferentiated variants. These lines have been characterized for the accumulation of RNAs corresponding to each of the albumin promoter-binding factors. Only HNF1, and to a lesser extent C/EBP, show differences depending upon the differentiation state of the cells. Overexpression of exogenous HNF1 in these cells reveals that this factor is able to transactivate the albumin promoter only in variant cells where the endogenous protein is limiting. By contrast, if the HNF1-binding site is of weak affinity, overexpresion of exogenous HNF1 stimulates the albumin promoter even in the HNF1-rich differentiated cells. Overexpression of exogenous LAP strongly transactivates an artificial promoter containing one LAP-binding site, but surprisingly in all the cell lines, it has little effect upon the albumin promoter. These results demonstrate that the transactivation potential of a given transcription factor depends on the degree of differentiation of the recipient cells, on the promoter structure, and on the affinity of the binding site for this factor.
Chondrons from articular cartilage: V.* Immunohistochemical evaluation of type VI collagen organisation in isolated chondrons by light, confocal and electron microscopy
Authors: Poole C. Anthony , Ayad Shirley , Gilbert Raymond T. 
Published: April 2021

ABSTRACT
The pericellular microenvironment around articular cartilage chondrocytes must play a key role in regulating the interaction between the cell and its extracellular matrix. The potential contribution of type VI collagen to this interaction was investigated in this study using isolated canine tibial chondrons embedded in agarose monolayers. The immunohistochemical distribution of an anti-type VI collagen antibody was assessed in these preparations using fluorescence, peroxidase and gold particle probes in combination with light, confocal and transmission electron microscopy. Light and confocal microscopy both showed type VI collagen concentrated in the pericellular capsule and matrix around the chondrocyte with reduced staining in the tail region and the interconnecting segments between adjacent chondrons. Minimal staining was recorded in the territorial and interterritorial matrices. At higher resolution, type VI collagen appeared both as microfibrils and as amorphous deposits that accumulated at the junction of intersecting capsular fibres and microfibrils. Electron microscopy also showed type VI collagen anchored to the chondrocyte membrane at the articular pole of the pericellular capsule and tethered to the radial collagen network through the tail at the basal pole of the capsule. We suggest that type VI collagen plays a dual role in the maintenance of chondron integrity. First, it could bind to the radial collagen network and stabilise the collagens, proteoglycans and glycoproteins of the pericellular microenvironment. Secondly, specific cell surface receptors exist, which could mediate the interaction between the chondrocyte and type VI collagen, providing firm anchorage and signalling potentials between the pericellular matrix and the cell nucleus. In this way type VI collagen could provide a close functional interrelationship between the chondrocyte, its pericellular microenvironment and the load bearing extracellular matrix of adult articular cartilage.
Multiplicity of clathrin light-chain-like polypeptides from developing pea (Pisum sativum L.) cotyledons
Authors: Lin Hong-Bo , Harley Suzanne M. , Butler Juliet M. , Beevers Leonard 
Published: April 2021

ABSTRACT
A comparative study has been made of clathrin-coated vesicles from developing pea (Pisum sativum L.) cotyledons and bovine brains in order to characterize the clathrin light chains from a plant system. Four polypeptides of 31 kDa, 40 kDa, 46 kDa and 50 kDa are considered as candidates for clathrin light chains in the developing pea cotyledons. The 31 kDa, 40 kDa, 46 kDa and 50 kDa polypeptides, together with the 190 kDa heavy chain, are dissociated as triskelions when coated vesicles of developing pea cotyledons are treated with 2 M urea. Partially purified 46 kDa and 50 kDa polypeptides have been demonstrated to bind to purified clathrin heavy chains. The 40 kDa, 46 kDa and 50 kDa polypeptides are sensitive to elastase. They are readily solubilized by neutralization of 10% trichloroacetic acid precipitates of clathrin. The 50 kDa polypeptide of plant clathrin-coated vesicles is heat-stable as are the light chains from bovine brains, while the heat stability of the 31 kDa, 40 kDa and 46 kDa polypeptides of plants is dependent on pH and ionic strength. The 40 kDa, 46 kDa and 50 kDa polypeptides bind calmodulin. The calcium binding properties of these polypeptides are ambiguous. The 40 kDa and 46 kDa polypeptides can be phosphorylated more extensively than the 31 kDa in vitro in the presence of polylysine, as can the smaller light chain of brains. The 50 kDa polypeptide can also be phosphorylated, even without the addition of polylysine. Unlike brain light chains, phosphorylation of the 31 kDa, 40 kDa, 46 kDa and 50 kDa polypeptides from peas is greatly reduced by N-ethylmaleimide (NEM). Our findings contrast with earlier reports of clathrin light chains of 30 and 38 kDa from zucchini and 57 and 60 kDa from carrots, respectively.
Cadherin-mediated adhesion in pancreatic islet cells is modulated by a cell surface N-acetylgalactosaminylphosphotransferase
Authors: Bauer G. Eric , Balsamo Janne , Lilien Jack 
Published: April 2021

ABSTRACT
Rat pancreatic islet cells and RINm5F cells, an islet derived cell line, have at their cell surface an N-acetylgalactosaminylphosphotransferase (GalNAcPTase) similar to that found at the surface of chick neural retina cells and at the rat neuromuscular junction. On islet cells and RINm5F cells the GalNAcPTase is stably associated with cadherin cell-cell adhesion molecules. The effect of antibodies directed against the GalNAcPTase on homophilic, cadherin mediated adhesion was analyzed by measuring their effect on adhesion of islet and RINm5F cells to an immobilized anti-cadherin antibody.
In this experimental paradigm anti-GalNAcPtase antibodies completely inhibit cadherin mediated adhesion. Furthermore, cadherin and GalNAcPTase co-distribute in islet and non-islet tissue. We conclude that pancreatic islet cell-cell adhesion is cadherin mediated and under the control of a tightly associated, cell surface GalNAcPTase.
Expression of naked plasmids by cultured myotubes and entry of plasmids into T tubules and caveolae of mammalian skeletal muscle
Authors: Wolff Jon A. , Dowty Martin E. , Jiao Shoushu , Repetto Gabriella , Berg Randi K. , Ludtke James J. , Williams Phillip , Slautterback David B. 
Published: April 2021

ABSTRACT
Plasmid DNA or artificial mRNA injected intramuscularly into skeletal muscle via a 27 g needle expressed transgenes at relatively efficient levels in skeletal myofibers and cardiac cells. In the present study, several approaches were used to determine the mechanism of cellular uptake. After exposure of naked plasmid DNA, primary rat muscle cells in vitro expressed transgenes to a much greater extent than other types of immortalized or primary cells. In vivo light microscope studies showed that intramuscularly injected plasmid DNA was distributed throughout the muscle and was able to diffuse through the extracellular matrix, cross the external lamina, and enter myofibers. Electron microscope studies showed that colloidal gold conjugated to plasmid DNA traversed the external lamina and entered T tubules and caveolae, while gold complexed with polylysine, polyethylene glycol or polyglutamate primarily remained outside of the myofibers. The results indicate that it is highly unlikely that the plasmid DNA enters the myofiber simply by the needle grossly disrupting the sarcolemma. In addition, transient membrane disruptions do not appear to be responsible for the uptake of DNA. Furthermore, no evidence for endocytosis could be found. The possible uptake of plasmid DNA by some type of cell membrane transporter, in particular via potocytosis, is discussed.
Haemocyte heterogeneity in the cockroach Periplaneta americana analysed using monoclonal antibodies
Authors: Chain Benjamin M. , Ø Rland Karen Leyshon-S , Siva-Jothy Michael T. 
Published: April 2021

ABSTRACT
A panel of monoclonal antibodies (mAbs) raised against whole haemocytes from Periplaneta americana has revealed two broad categories of immunogens.
Three mAbs showed reactivity with membrane-bound antigens on the three major morphological classes of haemocyte as well as the basement membrane of tissues in contact with the haemocoel. Moreover, this reactivity showed heterogeneity within morphotypes: this heterogeneity was quantified using flow cytometry.
In contrast, two other mAbs did not show reactivity with tissues other than haemocytes: both mAbs appeared to label unique intracellular components of the haemocytes.
We have demonstrated that mAbs can be used to define haemocyte subpopulations in P. americana that are not recognisable on morphological criteria alone. We believe this heterogeneity underlies the mechanisms responsible for the identification and neutralisation of non-self in the ‘immune’ response of P. americana.
Occurrence of brown adipocytes in rat white adipose tissue: molecular and morphological characterization
Authors: Cousin B. , Cinti S. , Morroni M. , Raimbault S. , Ricquier D. , Pénicaud L. , Casteilla L. 
Published: April 2021

ABSTRACT
Brown adipocytes are thermogenic cells which play an important role in energy balance. Their thermogenic activity is due to the presence of a mitochondrial uncoupling protein (UCP). Until recently, it was admitted that in rodents brown adipocytes were mainly located in classical brown adipose tissue (BAT). In the present study, we have investigated the presence of UCP protein or mRNA in white adipose tissue (WAT) of rats. Using polymerase chain reaction or Northern blot hybridization, UCP mRNA was detected in mesenteric, epidydimal, retroperitoneal, inguinal and particularly in periovarian adipose depots. The uncoupling protein was detected by Western blotting in mitochondria from periovarian adipose tissue. When rats were submitted to cold or to treatment with a -adrenoceptor agonist, UCP expression was increased in this tissue as in typical brown fat. Moreover, the expression was decreased in obese fa/fa rats compared to lean controls. Morphological studies showed that periovarian adipose tissue of rats kept at 24°C contained cells with numerous typical BAT mitochondria with or without multilocular lipid droplets. Immunocytochemistry confirmed that multilocular cells expressed mitochondrial UCP. Furthermore, the number of brown adipocytes and the density of mitochondrial cristae increased in parallel with exposure to cold. These results demonstrate that adipocytes expressing UCP are present in adipose deposits considered as white fat. They suggest the existence of a continuum in rodents between BAT and WAT, and a great plasticity between adipose tissue phenotypes. The physiological importance of brown adipocytes in WAT and the regulation of UCP expression remain open questions.
Increased microtubule stability and alpha tubulin acetylation in cells transfected with microtubule-associated proteins MAP1B, MAP2 or tau
Authors: Takemura Reiko , Okabe Shigeo , Umeyama Takashige , Kanai Yoshimitsu , Cowan Nicholas J. , Hirokawa Nobutaka 
Published: April 2021

ABSTRACT
We previously transfected MAP2, tau and MAP1B cDNA into fibroblasts and have studied the effect of expression of these microtubule-associated proteins on microtubule organization. In this study, we examined some additional characteristics of microtubule bundles and arrays formed in fibroblasts transfected with these microtubule-associated proteins. It was found that microtubule bundles formed in MAP2cor tau-transfected cells were stabilized against microtubule depolymerizing reagents and were enriched in acetylated alpha tubulin. When mouse MAP1B cDNA was expressed following transfection into COS cells, MAP1B was localized along microtubule arrays, but no extensive reorganization of microtubules such as bundle formation was observed, in agreement with our previous finding using HeLa and 3T3 cells. However, stabilization of microtubules was indicated: (a) microtubules in MAP1B-transfected cells were stabilized against a microtubule depolymerizing reagent, although stabilization was less efficient than that seen in MAP2cor tau-transfected cells, and (b) microtubules in MAP1B-transfected cells were enriched in acetylated alpha tubulin. These results suggest that neuronal microtubule-associated proteins introduced into fibroblasts by cDNA transfection stabilize microtubules and affect the state of post-translational modification of tubulin.
Microtubule and F-actin dynamics at the division site in living Tradescantia stamen hair cells
Authors: Cleary Ann L. , Gunning Brian E. S. , Wasteneys Geoffrey O. , Hepler Peter K. 
Published: April 2021

ABSTRACT
We have visualised F-actin and microtubules in living Tradescantia virginiana stamen hair cells by confocal laser scanning microscopy after microinjecting rhodamine-phalloidin or carboxyfluorescein-labelled brain tubulin. We monitored these components of the cytoskeleton as the cells prepared for division at preprophase and progressed through mitosis to cytokinesis. Reorganisation of the interphase cortical cytoskeleton results in preprophase bands of both F-actin and microtubules that coexist in the cell cortex, centred on the site at which the future cell plate will fuse with the parent cell wall. The preprophase band of microtubules is formed from microtubules that polymerise and incorporate tubulin during prophase. The preprophase band of actin may form either by reorganisation of pre-existing filaments or by de novo polymerisation. Both cytoskeletal components disappear from the future division site approximately five minutes prior to the breakdown of the nuclear envelope. Cortical microtubules are undetectable throughout mitosis and cytokinesis, whereas cortical F-actin remains abundant, although it is notably excluded from the division site. The phragmoplast, containing both F-actin and microtubules, expands towards the cortical actin exclusion-zone through a region that has no detectable microtubules or F-actin. The phragmoplast comes to rest in the predefined region of the cortex that is devoid of F-actin. It is proposed that cortical F-actin may act as a “negative” template which could position the phragmoplast and cell plate correctly. This is the first in vivo documentation of F-actin dynamics at the division site in living plant cells.
Nuclear proteins of the bovine esophageal epithelium: II. The NuMA gene gives rise to multiple mRNAs and gene products reactive with monoclonal antibody W1
Authors: Tang Tang K. , Tang Chieh-ju C. , Chen Yin-Ling , Wu Cheng-Wen 
Published: April 2021

ABSTRACT
Treatment of small cells derived from the basal layer of bovine esophageal epithelium, with Triton X-100, urea and sonication resulted in a nuclear residue that was used as an immunogen for generation of monoclonal antibodies directed against nuclear components. One such antibody, designated W1, was found to label the nuclei of all cells examined. In interphase cells, the target antigen of antibody W1 was diffusely distributed in the nucleus. During metaphase, however, the W1 antigen formed prominent crescents at the poles of the mitotic spindle, diminished gradually in anaphase, and finally redistributed into the regenerating daughter nuclei. Western blotting with antibody W1 yielded a prominent polypeptide of Mr ∼230,000. The amino acid sequence, deduced from the nucleotide sequence of several overlapping cDNA clones that span the entire coding region, revealed that the W1 polypeptide was identical to the Nuclear Mitotic Apparatus (NuMA) protein, with a long a-helical central core flanked by two nonhelical domains. Interestingly, most cDNA sequences were identical to each other, except for six sequence blocks which were either inserted or deleted in individual cDNA clones. Analysis of the cDNA sequences of various clones, coupled with polymerase chain reaction amplification of cellular mRNA and genomic Southern blotting with region-specific probes, all indicated that multiple mRNA species were present in U-251 human glioma cells, derived from alternative splicing of the RNA transcript from a single NuMA/W1 gene. Besides the predominant form of the mRNA giving rise to the polypeptide of Mr ∼230,000, two other forms of mRNA, which arise as a result of alternative splicing and which use different translation termination codons, may yield lower molecular weight polypeptide products. Consistent with this notion, polypeptides of Mr ∼195,000 and ∼194,000 have been observed in this and other studies on the NuMA/W1 protein. These data suggest that multiple isoforms of the NuMA polypeptides generated by alternative mRNA splicing may play some important functions which remain to be characterized.