scholarly journals Proline Residues in Human Immunodeficiency Virus Type 1 p6Gag Exert a Cell Type-Dependent Effect on Viral Replication and Virion Incorporation of Pol Proteins

1999 ◽  
Vol 73 (6) ◽  
pp. 4696-4704 ◽  
Author(s):  
Markus Dettenhofer ◽  
Xiao-Fang Yu
Keyword(s):  
Microscopic Analysis ◽  
Jurkat Cells ◽  
Cell Type ◽  
Electron Microscopic ◽  
Gag Protein ◽  
Protein Levels ◽  
Rich Domain ◽  
Virion Incorporation ◽  
C Terminus ◽  
A Cell

ABSTRACT The C terminus of the HIV-1 Gag protein contains a proline-rich domain termed p6Gag. This domain has been shown to play a role in efficient virus release and incorporation of Vpr into virions. In a previous study (X. F. Yu, L. Dawson, C. J. Tian, C. Flexner, and M. Dettenhofer, J. Virol. 72:3412–3417, 1998), we observed that the removal of the p6 domain of Gag as well as drastic mutations in the PTAP motif resulted in reduced virion-associated Pol proteins from transfected COS cells. In the present study, amino acid substitutions at residues 5 and 7 of p6Gag resulted in a cell type-dependent replication of the mutant virus in CD4+T cells; the virus was replication competent in Jurkat cells but restricted in H9 cells and primary blood-derived monocytes. Established Jurkat and H9 cell lines expressing p6Gag mutant and parental virus were used to further understand this defect. Mutant virions produced from H9 cells, which displayed no defect in extracellular virion production, showed an ∼16-fold reduction in Pol protein levels, whereas the levels of Pol proteins were only marginally reduced in mutant virions produced from Jurkat cells. The reduction in the virion-associated Pol proteins could not be accounted for by differences in the levels of intracellular p160Gag-Pol or in the interaction between p55Gag and p160Gag-Pol precursors. Electron microscopic analysis of the p6Gag mutant virions showed a predominately immature morphology in the absence of significant defects in Gag proteolytic cleavage. Taken together, these data suggest that the proline-rich motif of p6Gag is involved in the late stages of virus maturation, which include the packaging of cleaved Pol proteins in viral particles, a process which may involve cell-type-specific factors.

Dendritic cells of the human epidermis: immuno-electron microscopic studies of la antigen reactivity

1978 ◽  
Vol 36 (2) ◽  
pp. 644-645
Author(s):  
G. Rowden ◽  
M. G. Lewis ◽  
T. M. Phillips
Keyword(s):  
Dendritic Cells ◽  
Langerhans Cells ◽  
Cell Types ◽  
Cell Type ◽  
Electron Microscopic ◽  
La Antigen ◽  
Microscopic Studies ◽  
A Cell ◽  
C3 Receptors ◽  
Antigen Reactivity

Langerhans cells of mammalian stratified squamous epithelial have proven to be an enigma since their discovery in 1868. These dendritic suprabasal cells have been considered as related to melanocytes either as effete cells, or as post divisional products. Although grafting experiments seemed to demonstrate the independence of the cell types, much confusion still exists. The presence in the epidermis of a cell type with morphological features seemingly shared by melanocytes and Langerhans cells has been especially troublesome. This so called "indeterminate", or " -dendritic cell" lacks both Langerhans cells granules and melanosomes, yet it is clearly not a keratinocyte. Suggestions have been made that it is related to either Langerhans cells or melanocyte. Recent studies have unequivocally demonstrated that Langerhans cells are independent cells with immune function. They display Fc and C3 receptors on their surface as well as la (immune region associated) antigens.

scholarly journals Subcellular distribution and targeting of the intracellular chloride channel p64.

1997 ◽  
Vol 8 (4) ◽  
pp. 691-704 ◽  
Author(s):  
C Redhead ◽  
S K Sullivan ◽  
C Koseki ◽  
K Fujiwara ◽  
J C Edwards
Keyword(s):  
Chloride Channel ◽  
Subcellular Distribution ◽  
Cell Type ◽  
Electron Microscopic ◽  
Intracellular Chloride ◽  
Terminal Domain ◽  
Electron Microscopic Technique ◽  
Membrane Compartment ◽  
A Cell ◽  
Kidney Microsomes

p64 is an intracellular chloride channel originally identified in bovine kidney microsomes. Using a combination of immunofluorescent and electron microscopic technique, we demonstrate that p64 resides in the limiting membranes of perinuclear dense core vesicles which appear to be regulated secretory vesicles. Heterologous expression of p64 in PancI cells, a cell type which does not normally express p64, results in targeting to a similar compartment. Mutagenesis experiments demonstrate that both the N- and C-terminal domains of the protein independently contribute to subcellular distribution of the protein. The C-terminal domain functions to prevent expression of p64 on the plasma membrane and the N-terminal domain is necessary to deliver p64 to the appropriate membrane compartment.

scholarly journals Suppression of Damping-Off Disease in Host Plants by the Rhizoplane Bacterium Lysobacter sp. Strain SB-K88 Is Linked to Plant Colonization and Antibiosis against Soilborne Peronosporomycetes

2005 ◽  
Vol 71 (7) ◽  
pp. 3786-3796 ◽  
Author(s):  
Md. Tofazzal Islam ◽  
Yasuyuki Hashidoko ◽  
Abhinandan Deora ◽  
Toshiaki Ito ◽  
Satoshi Tahara
Keyword(s):  
Sugar Beet ◽  
Microscopic Analysis ◽  
Inhibitory Effect ◽  
Root Surface ◽  
Plant Colonization ◽  
Damping Off ◽  
Electron Microscopic ◽  
Aphanomyces Cochlioides ◽  
Direct Inhibitory Effect ◽  
A Cell

ABSTRACT We previously demonstrated that xanthobaccin A from the rhizoplane bacterium Lysobacter sp. strain SB-K88 suppresses damping-off disease caused by Pythium sp. in sugar beet. In this study we focused on modes of Lysobacter sp. strain SB-K88 root colonization and antibiosis of the bacterium against Aphanomyces cochlioides, a pathogen of damping-off disease. Scanning electron microscopic analysis of 2-week-old sugar beet seedlings from seeds previously inoculated with SB-K88 revealed dense colonization on the root surfaces and a characteristic perpendicular pattern of Lysobacter colonization possibly generated via development of polar, brush-like fimbriae. In colonized regions a semitransparent film apparently enveloping the root and microcolonies were observed on the root surface. This Lysobacter strain also efficiently colonized the roots of several plants, including spinach, tomato, Arabidopsis thaliana, and Amaranthus gangeticus. Plants grown from both sugar beet and spinach seeds that were previously treated with Lysobacter sp. strain SB-K88 displayed significant resistance to the damping-off disease triggered by A. cochlioides. Interestingly, zoospores of A. cochlioides became immotile within 1 min after exposure to a SB-K88 cell suspension, a cell-free supernatant of SB-K88, or pure xanthobaccin A (MIC, 0.01 μg/ml). In all cases, lysis followed within 30 min in the presence of the inhibiting factor(s). Our data indicate that Lysobacter sp. strain SB-K88 has a direct inhibitory effect on A. cochlioides, suppressing damping-off disease. Furthermore, this inhibitory effect of Lysobacter sp. strain SB-K88 is likely due to a combination of antibiosis and characteristic biofilm formation at the rhizoplane of the host plant.

scholarly journals Mapping and Characterization of the N-Terminal I Domain of Human Immunodeficiency Virus Type 1 Pr55Gag

2000 ◽  
Vol 74 (16) ◽  
pp. 7238-7249 ◽  
Author(s):  
Stephanie Sandefur ◽  
Rita M. Smith ◽  
Vasundhara Varthakavi ◽  
Paul Spearman
Keyword(s):  
Human Immunodeficiency Virus ◽  
Plasma Membrane ◽  
Fluorescent Protein ◽  
Microscopic Analysis ◽  
Viral Gene ◽  
Electron Microscopic ◽  
Particle Assembly ◽  
Gag Protein ◽  
Immunodeficiency Virus

ABSTRACT Human immunodeficiency virus (HIV) type 1 particles assemble at the plasma membrane of cells in a manner similar to that of the type C oncoretroviruses. The Pr55Gag molecule directs the assembly process and is sufficient for particle assembly in the absence of all other viral gene products. The I domain is an assembly domain that has been previously localized to the nucleocapsid (NC) region of Gag. In this study we utilized a series of Gag-green fluorescent protein (GFP) fusion proteins to precisely identify sequences that constitute the N-terminal I domain of Pr55Gag. The minimal sequence required for the I domain was localized to the extreme N terminus of NC. Two basic residues (arginine 380 and arginine 384) within the initial seven residues of NC were found to be critical for the function of the N-terminal I domain. The presence of positive charge alone in these two positions, however, was not sufficient to mediate the formation of dense Gag particles. The I domain was required for the formation of detergent-resistant complexes of Gag protein, and confocal microscopy demonstrated that the I domain was also required for the formation of punctate foci of Gag proteins at the plasma membrane. Electron microscopic analysis of cells expressing Gag-GFP fusion constructs with an intact I domain revealed numerous retrovirus-like particles (RVLPs) budding from the plasma membrane, while I domain-deficient constructs failed to generate visible RVLPs. These results provide evidence that Gag-Gag interactions mediated by the I domain play a central role in the assembly of HIV particles.

scholarly journals Sanqi Oral Solution Mitigates Proteinuria in Rat Passive Heymann Nephritis and Blocks Podocyte Apoptosis via Nrf2/HO-1 Pathway

2021 ◽  
Vol 12 ◽  
Author(s):  
Xiaowan Wang ◽  
Jinchu Liu ◽  
Ruimin Tian ◽  
Bidan Zheng ◽  
Chuang Li ◽  
...  
Keyword(s):  
Microscopic Analysis ◽  
Oral Solution ◽  
Complement C3 ◽  
Heme Oxygenase 1 ◽  
Related Factor ◽  
Electron Microscopic ◽  
Protein Levels ◽  
Heymann Nephritis

Idiopathic membranous nephropathy (IMN) is the most common pathological type in adult nephrotic syndrome where podocyte apoptosis was found to mediate the development of proteinuria. Sanqi oral solution (SQ), an effective Chinese herbal preparation clinically used in treatment of IMN for decades, plays an important role in reducing proteinuria, but the underlying mechanisms have not been fully elucidated yet. The current study tested the hypothesis that SQ directly lessens proteinuria in IMN by reducing podocyte apoptosis. To investigate the effects of SQ, we established the experimental passive Heymann nephritis (PHN) rat model induced by anti-Fx1A antiserum in vivo and doxorubicin hydrochloride (ADR)-injured apoptotic podocyte model in vitro. SQ intervention dramatically reduced the level of proteinuria, together with the rat anti-rabbit IgG antibodies, complement C3, and C5b-9 deposition in glomerulus of PHN rats, accompanied by an elevation of serum albumin. Protein expression of synaptopodin, marker of podocyte injury, restored after SQ administration, whereas the electron microscopic analysis indicated that fusion of foot processes, and the pachynsis of glomerular basement membrane was markedly diminished. Further studies showed that SQ treatment could significantly inhibit podocyte apoptosis in PHN rats and ADR-injured podocytes, and protein levels of Cleaved Caspase-3 or the ratio of Bax/Bcl-2 were significantly decreased with SQ treatment in vivo or in vitro. Moreover, we found that the nuclear factor erythroid 2–related factor-2/heme oxygenase 1 (Nrf2/HO-1) pathway mediated the anti-apoptosis effective of SQ in podocyte. Thus, SQ mitigates podocyte apoptosis and proteinuria in PHN rats via the Nrf2/HO-1 pathway.

scholarly journals Human mitochondria and mitochondrial genome function as a single dynamic cellular unit.

1994 ◽  
Vol 125 (1) ◽  
pp. 43-50 ◽  
Author(s):  
J Hayashi ◽  
M Takemitsu ◽  
Y Goto ◽  
I Nonaka
Keyword(s):  
Hela Cells ◽  
Single Cells ◽  
Microscopic Analysis ◽  
Large Deletion ◽  
Human Cells ◽  
Homogeneous Distribution ◽  
Electron Microscopic ◽  
Mutant Type ◽  
A Cell ◽  
Dapi Fluorescence

rho 0 HeLa cells entirely lacking mitochondrial DNA (mtDNA) and mitochondrial transfection techniques were used to examine intermitochondrial interactions between mitochondria with and without mtDNA, and also between those with wild-type (wt) and mutant-type mtDNA in living human cells. First, unambiguous evidence was obtained that the DNA-binding dyes ethidium bromide (EtBr) and 4',6-diamidino-2-phenylindole (DAPI) exclusively stained mitochondria containing mtDNA in living human cells. Then, using EtBr or DAPI fluorescence as a probe, mtDNA was shown to spread rapidly to all rho 0 HeLa mitochondria when EtBr- or DAPI-stained HeLa mitochondria were introduced into rho 0 HeLa cells. Moreover, coexisting wt-mtDNA and mutant mtDNA with a large deletion (delta-mtDNA) were shown to mix homogeneously throughout mitochondria, not to remain segregated by use of electron microscopic analysis of cytochrome c oxidase activities of individual mitochondria as a probe to identify mitochondria with predominantly wt- or delta-mtDNA in single cells. This rapid diffusion of mtDNA and the resultant homogeneous distribution of the heteroplasmic wt- and delta-mtDNA molecules throughout mitochondria in a cell suggest that the mitochondria in living human cells have lost their individuality. Thus, the actual number of mitochondria per cell is not of crucial importance, and mitochondria in a cell should be considered as a virtually single dynamic unit.

scholarly journals Breast cancer-associated protein – a novel binding partner of Mason-Pfizer monkey virus protease

2014 ◽  
Vol 95 (6) ◽  
pp. 1383-1389 ◽  
Author(s):  
Michaela Rumlová ◽  
Ivana Křížová ◽  
Romana Hadravová ◽  
Michal Doležal ◽  
Karolína Strohalmová ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Potential Role ◽  
Protein A ◽  
Proteolytic Processing ◽  
Particle Release ◽  
Binding Partner ◽  
Rich Domain ◽  
Virion Incorporation ◽  
C Terminus

We identified breast cancer-associated protein (BCA3) as a novel binding partner of Mason-Pfizer monkey virus (MPMV) protease (PR). The interaction was confirmed by co-immunoprecipitation and immunocolocalization of MPMV PR and BCA3. Full-length but not C-terminally truncated BCA3 was incorporated into MPMV virions. We ruled out the potential role of the G-patch domain, a glycine-rich domain located at the C terminus of MPMV PR, in BCA3 interaction and virion incorporation. Expression of BCA3 did not affect MPMV particle release and proteolytic processing; however, it slightly increased MPMV infectivity.

scholarly journals TNNI1 Mutated in Autosomal Dominant Proximal Arthrogryposis

2021 ◽  
Vol 8 (1) ◽  
pp. e649
Author(s):  
Yukako Nishimori ◽  
Aritoshi Iida ◽  
Masashi Ogasawara ◽  
Mariko Okubo ◽  
Yuki Yonenobu ◽  
...  
Keyword(s):  
Autosomal Dominant ◽  
Genomic Analysis ◽  
Microscopic Analysis ◽  
Electron Microscopic ◽  
Japanese Family ◽  
Genetic Studies ◽  
C Terminus ◽  
The Family

ObjectivesThe main objective of this case report is to identify a gene associated with a Japanese family with autosomal dominant arthrogryposis.MethodsWe performed clinicopathologic diagnosis and genomic analysis using trio-based exome sequencing.ResultsA 14-year-old boy had contractures in the proximal joints, and the serum creatine kinase level was elevated. Muscle biopsy demonstrated a moth-eaten appearance in some type 1 fibers, and electron microscopic analysis revealed that type 1 fibers had Z disk streaming. We identified a heterozygous nonsense variant, c.523A>T (p.K175*), in TNNI1 in the family.DiscussionThe altered amino acid residue is within the tropomyosin-binding site near the C-terminus, in a region homologous to the variational hotspot of Troponin I2 (TNNI2), which is associated with distal arthrogryposis type 1 and 2b. Compared with patients with TNNI2 variants, our patient had a milder phenotype and proximal arthrogryposis. We report here a case of proximal arthrogryposis associated with a TNNI1 nonsense variant, which expands the genetic and clinical spectrum of this disease. Further functional and genetic studies are required to clarify the role of TNNI1 in the disease.

From light to electron microscopic analysis of neurons intracellularly injected in fixed slices with “miniruby”, a fluorescent biocyttn compound

1993 ◽  
Vol 51 ◽  
pp. 290-291
Author(s):  
Wei-lin Liu ◽  
Michael T. Shipley
Keyword(s):  
Piriform Cortex ◽  
Microscopic Analysis ◽  
Microscopic Level ◽  
Electron Microscopic Level ◽  
Light Microscopic Level ◽  
Electron Microscopic ◽  
Fluorescent Compound ◽  
Adult Rat ◽  
Living Slices ◽  
A Cell

Intracellular labeling of neurons in fixed slices is a very useful method for studying morphological structures of neurons both at light and electron microscopic levels. Recently, biocytin has been widely used for intracellular labeling in living slices because this molecule is highly soluble, has high electrophoretic mobility and has high affinity for avidin. However, biocytin cannot be used in fixed slices because in fixed slices membrane potential cannot be used to signify that a cell is impaled. Thus, in fixed slices it is necessary to inject cells with a fluorescent compound so that impalement and filling can be visualized under fluorescent microscope. We have developed a fluorescent biocytin compound, “Miniruby” (MR), dextran-tetramethylrhodamine-biocytin. previously, we showed that mis molecule provides excellent intracellular labels in fixed slices at the light microscopic level. Here, we demonstrate MR can also be visualized at the electron microscopic level.Fixed slices (200-400 ¼m) of adult rat olfactory bulb, piriform cortex and periaqeductal gray were used.

Purification of intact nuclear lamina and identification of novel laminlike proteins in Raji, a cell line devoid of lamins A and C

1997 ◽  
Vol 75 (6) ◽  
pp. 721-732 ◽  
Author(s):  
Sylvain Foisy ◽  
Erik C Joly ◽  
Viviane Bibor-Hardy
Keyword(s):  
Cell Line ◽  
Microscopic Analysis ◽  
Nuclear Lamina ◽  
Intermediate Filament Protein ◽  
Electron Microscopic ◽  
Nuclear Lamins ◽  
A Cell ◽  
Buffer Composition ◽  
Nuclear Structures ◽  
Filament Protein

Research on the structure of the nuclear lamina and the nuclear matrix of cells devoid of lamins A and C has been hampered by the fact that intact residual nuclear structures are difficult to isolate from such cells. In this paper, we show that some extraction parameters, such as buffer composition and the nature of the detergent used to remove nuclear membranes, are critical for achieving isolation of whole nuclear residual structures from the lymphoblastic cell line Raji, used as a model for cells without lamins A and C. Electron microscopic analysis shows that the nuclear lamina of Raji cells is formed by a network of intermediate-sized filaments interrupted with circular discontinuities. Both lamins B1 and B2, and lamin D/E, are present in this structure. In addition, a group of 45-kDa proteins or intermediate filament protein - reacting proteins (IFA-RPs), located uniquely in the lamina, were found to exhibit the same immunological and chemical characteristics as lamins. Although they behave like nuclear lamins, microsequencing analysis of the IFA-RPs has revealed no homology with known lamins. These IFA-RPs may contribute to the formation of the nuclear lamina filament network in the absence of lamins A and C. Key words: nuclear lamina, intermediate filaments, lamin.

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