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PubMed2002

Chemical amplification with encapsulated reagents.

Autocatalysis and chemical amplification are characteristic properties of living systems, and they give rise to behaviors such as increased sensitivity, responsiveness, and self-replication. Here we report a synthetic system in which a unique form of compartmentalization leads to nonlinear, autocatalytic behavior. The compartment is a reversibly formed capsule in which a reagent is sequestered. Reaction products displace the reagent from the capsule into solution and the reaction rate is accelerated. The resulting self-regulation is sensitive to the highly selective molecular recognition properties of the capsule.

Journal Article↗
PubMed1987

Sorting of progeny coronavirus from condensed secretory proteins at the exit from the trans-Golgi network of AtT20 cells.

Murine hepatitis virus (strain A59), (MHV-A59) is a coronavirus that buds into pre-Golgi compartments and then exploits the exocytic pathway of the host cell to reach the exterior. The fibroblastic cells in which replication of this virus is usually studied have only a constitutive exocytic pathway that the virus uses. MHV-A59 also infects, albeit inefficiently, AtT20 cells, murine pituitary tumor cells with a regulated as well as a constitutive exocytic pathway. Here we examine AtT20 cells at early times after the infection, when the Golgi apparatus retains its morphological and biochemical integrity. We observe that progeny coronavirus and secretory protein destined for the secretory granules of the regulated exocytic pathway traverse the same Golgi stacks and accumulate in the trans-Golgi network. Their pathways diverge at this site, the condensed secretory proteins including the ACTH going to the secretory granules and the coronavirus to post-Golgi transport vesicles devoid of ACTH. On very rare occasions there is missorting such that aggregates of condensed secretory proteins and viruses occur together in post-Golgi vesicles. We conclude that the constitutive and regulated exocytic pathways, identified respectively by the progeny virions and the secretory protein ACTH, diverge at the exit from the trans-Golgi network.

Adrenocorticotropic Hormone↗
PubMed2001

Knee bracing for medial compartment osteoarthritis: effects on proprioception and postural control.

OBJECTIVE: To evaluate the effects of a functional knee brace specifically designed for patients with varus gonarthrosis on measures of proprioception and postural control. SUBJECTS: Fourteen men and six women (aged 59+/-9 yr) with measurable varus alignment and osteoarthritis of the knee medial compartment. METHODS: Proprioception was assessed in the sitting position using an isokinetic dynamometer and was quantified as the ability to replicate target knee-joint angles. Postural control was assessed with a force platform using tests of single-limb standing balance performed, while the patient was standing on a stable surface and standing on foam, and was quantified as the total length of the path of the centre of pressure. All tests were performed with and without the patient's own custom-fit valgus brace. RESULTS: Proprioception was significantly improved following application of the brace [mean difference=0.7 degrees, 95% confidence interval (CI)=0.2 to 1.1 degrees ). Postural control was not significantly affected by the use of the brace during the stable surface test (mean difference=2.6 cm, 95% CI=-4.3 to 9.5 cm) or the foam surface test (mean difference=0.9 cm, 95% CI=-7.5 to 9.4 cm). CONCLUSION: Although enhanced proprioception may be partially responsible for reported improvements with the use of a brace, the present findings call into question the functional importance of the small changes observed.

Braces↗
PubMed1993

Hypoexpression of major histocompatibility complex molecules on Legionella pneumophila phagosomes and phagolysosomes.

Legionella pneumophila is a facultative intracellular pathogen that parasitizes host mononuclear phagocytes. Cell-mediated immunity is pivotal to host defense against L. pneumophila, and the infected host cell may play a central role in processing and presenting parasite antigens to lymphocytes mediating cell-mediated immune response. However, in the case of L. pneumophila and intracellular parasites in general, little is known about the intracellular trafficking of parasite antigens, the influence of parasite infection on major histocompatibility complex (MHC) expression, or the relationship of MHC molecules to sites of parasite replication. To learn more about this, we have used flow cytometry to study the expression of HLA-DR by monocytes infected with L. pneumophila and cryosection immunogold electron microscopy to study the distribution of MHC class I and II molecules on L. pneumophila phagosomes. Flow cytometry analysis demonstrated that L. pneumophila infection has little effect on the overall expression of HLA-DR by monocytes. Cryosection immunogold studies revealed abundant staining for MHC class I and II molecules on the plasma membrane of infected monocytes but little or no staining on the membranes of mature L. pneumophila phagosomes. Cryosection immunogold studies of an avirulent mutant of L. pneumophila that, unlike the wild type, does not inhibit phagosome-lysosome fusion and subsequently survives but does not multiply in a phagolysosome yielded similar results. We have previously found that MHC class I and II molecules are excluded from nascent phagosomes during coiling and conventional phagocytosis. The present work demonstrates that MHC molecules do not accumulate appreciably in the L. pneumophila phagosome as it matures and at a point in the life cycle of the organism in which it is replicating and producing immunoprotective T-cell antigens. This suggests that L. pneumophila does not reside in a typical endosomal compartment in the host cell and that L. pneumophila antigens may encounter MHC molecules at extraphagosomal sites within the host cell.

Animals↗
PubMed1994

Fate of Listeria monocytogenes in murine macrophages: evidence for simultaneous killing and survival of intracellular bacteria.

The intracellular survival of the ubiquitous pathogen Listeria monocytogenes was studied in primary cultures of bone marrow-derived mouse macrophages. Bacteria were able to grow rapidly in these cells, with an apparent multiplication rate of about 40 min. Electron microscopy demonstrated that intracellular bacterial replication was the consequence of simultaneous intracellular killing and replication of bacteria in the same cells. Within the first hour following phagocytosis, most bacteria were destroyed in the phagosomal compartment to which they were confined. This was due to early transfer of hydrolytic enzymes to phagosomes, undoubtedly via phagosome-lysosome (P-L) fusion, as demonstrated by a quantitative analysis after staining for a lysosomal marker, acid phosphatase. One hour after infection, about 14% of the bacteria were free in the cytoplasm, in which they multiplied and induced actin polymerization and spreading to adjacent macrophages, as in epithelial cells. By using the 3-(2,4-dinitroanilino)-3'-amino-N-methyldipropylamine staining procedure, direct evidence is presented that all phagosomes were acidified immediately after phagocytosis, thus indicating that intraphagosomal bacteria were exposed to an acidic environment that might favor vacuolar lysis by listeriolysin O. Intracellular growth in macrophages, therefore, appears to be the result of a competition between the expression of the hydrolytic activity of these cells following P-L fusion and the capacity of L. monocytogenes to escape from the acidified phagosomal compartment before P-L fusion has occurred. The finding that concomitant intracellular killing and survival of L. monocytogenes occurs in the same macrophages might explain the high immunogenicity observed in vivo with live bacteria, as opposed to killed bacteria.

Acid Phosphatase↗
PubMed1998

Macrophage NRAMP1 and its role in resistance to microbial infections.

The identification and characterization of genetic factors influencing natural susceptibility to infectious diseases in humans and in model organisms, such as the laboratory mouse, can provide new insight into the basic mechanisms of host defense against infections. In the mouse, resistance or susceptibility to infection with intracellular pathogens such as Salmonella, Mycobacterium and Leishmnania is controlled by the Natural resistance associated macrophage protein (Nramp1) gene on chromosome 1, which influences the rate of intracellular replication of these parasites in macrophages. Nramp1 codes for an integral membrane protein, which is expressed exclusively in macrophage/monocytes and polymorphonuclear leukocytes. The protein is localized to the endosomal/lysosomal compartment of the macrophage and is rapidly recruited to the membrane of the particle-containing phagosome upon phagocytosis. Nramp defines a novel family of functionally related membrane proteins including Nramp2, which was recently shown to be the major transferrin-independent uptake system of the intestine in mammals. This observation supports the hypothesis that the phagocyte-specific Nramp1 protein may regulate the intraphagosomal replication of antigenically unrelated bacteria by controlling divalent cation concentrations at that site. Recent genetic studies have found that allelic variants at the human NRAMP1 locus are associated with susceptibility to leprosy (Mycobacterium leprae) and tuberculosis (Mycobacterium tuberculosis) and possibly with the onset of rheumatoid arthritis.

Animals↗
PubMed1998

Cytokine response in multiple lymphoid tissues during the primary phase of feline immunodeficiency virus infection.

Type 1 and 2 cytokine mRNA responses were measured at various time periods and in various lymphoid compartments during the acute stage (first 4 months) of feline immunodeficiency virus (FIV) infection in laboratory cats. Cytokine responses were correlated with virus replication. Virus was detected in plasma and tissue from day 14 postinfection (p.i.) onward, peaked at 56 to 70 days, and declined greatly by 70 days. Virus replication was highest in the thymus, followed by spleen, mesenteric lymph nodes, and cervical lymph nodes. Baseline cytokine levels were highest in the mesenteric lymph nodes and lowest in the cervical lymph nodes. Cytokine upregulation after FIV infection was most dramatic in the cervical lymph nodes, with the greatest increase in interleukin-10 (IL-10) and gamma interferon (IFN-gamma). Cytokine transcription in the mesenteric lymph node increased above baseline by day 14 p.i. for IFN-gamma, IL-12p40, IL-4, and IL-10, while elevations in the spleen were mainly for IFN-gamma, IL-12p40 and IL-10. An increase in IFN-gamma, IL-10, and IL-12p40 occurred in the thymus at day 56 p.i., concomitant with the onset of thymitis. In general, type 2 cytokines (IL-4 and IL-10) were increased greater than 1 log over baseline, while the elevations in type 1 cytokines were less than 1 log. In the tissues tested, CD4(+) cells were the primary source of IL-2, IL-4, and IL-10. Both CD4(+) and CD8(+) cells produced IFN-gamma, while no cytokine mRNA was detected in B cells. These results demonstrate the presence of a heterogeneous cytokine response in lymphoid tissues during the primary stage of FIV infection. The nature and intensity of the response differed from one compartment to the other and, in the case of the thymus, also with inflammatory changes. Although limited in scope, the present study confirms the usefulness of the FIV infection model in studying early cytokine events that lead to the secondary subclinical carrier state typical of most lentivirus infections.

Animals↗
PubMedDate not supplied

[Site-specific expression of single-stranded antibody fragments in Nicotiana tabacum].

Antibody expression and immunomodulation are modern molecular techniques to produce pharmaceuticals and to interfere with cellular metabolism or pathogen infectivity in plants. Nonetheless, there is still no generally applicable strategy to express correctly folded active antibodies or antibody fragments in different cell compartments. To facilitate expression, single-chain antibody fragments (scFvs) were made of mouse monoclonal antibodies, J2 and P6 that specifically recognize double-stranded RNA (dsRNA). Stabilizing double-stranded replication intermediates could modulate the biological activity of dsRNAs in plants, especially to influence virus replication. Along with cytoplasmic expression, scFvs were anchored to the plasma membrane; targeted to the apoplast for secretion and made ER-resident. Expression levels were analysed and transgenic plants were evaluated for resistance or tolerance to potato virus Y infection. We have established strategies for expression of correctly assembled antibodies or antibody fragments in different plant cell compartments.

Animals↗
PubMed1988

Coupled induction of exocrine proteins and intracellular compartments involved in the secretory pathway in AR4-2J cells by glucocorticoids.

Treatment of AR4-2J cells with dexamethasone at 10 nM for 96 h inhibited cell replication by 75% and increased cell size (30%), protein content (1.6-fold) and protein synthesis (2-fold). The increase in protein synthesis was largely due to a 5 to 10-fold increase in the synthesis of secretory proteins. Amylase activity increased 20 to 30-fold in cellular homogenates and 10 to 20-fold in culture medium. Both in the presence and absence of dexamethasone AR4-2J cells release their secretory proteins by constitutive secretion. The proportion of newly synthesized amylase retained by the cells over the 14 h labeling period increased from 15 to 30% with hormone treatment. As judged by comigration on polyacrylamide gels and Western blots analyzed by immunospecific sera, AR4-2J cells synthesize and secrete the majority of known pancreatic secretory proteins. Dexamethasone increased the synthesis of trypsinogen 12 to 16-fold, chymotrypsinogen 4.5 to 6-fold, the group of procarboxypeptidases 6-fold, and amylase 7 to 10-fold. Messenger RNA levels for trypsinogen, amylase and lipase were each increased 4 to 5-fold. At the ultrastructural level dexamethasone led to significant increases in rough endoplasmic reticulum (RER) (30-fold) and Golgi elements (1.5-fold) and to the de novo appearance of electron-opaque granules (0.1-0.5 microns) which were shown to contain amylase by immunolocalization techniques employing protein A-gold. Dexamethasone also led to the formation of gap junctions between AR4-2J cells. These findings indicate that AR4-2J cells provide a model for differentiation of pancreatic acinar cells which should also be studied for the differentiation markers for the regulated secretory pathway.

Amylases↗
PubMed2003

A multicompartment model of carboxyhemoglobin and carboxymyoglobin responses to inhalation of carbon monoxide.

We have developed a model that predicts the distribution of carbon monoxide (CO) in the body resulting from acute inhalation exposures to CO. The model includes a lung compartment, arterial and venous blood compartments, and muscle and nonmuscle soft tissues with both vascular and nonvascular subcompartments. In the model, CO is allowed to diffuse between the vascular and nonvascular subcompartments of the tissues and to combine with myoglobin in the nonvascular subcompartment of muscle tissue. The oxyhemoglobin dissociation curve is represented by a modified Hill equation whose parameters are functions of the carboxyhemoglobin (HbCO) level. Values for skeletal muscle mass and cardiac output are calculated from prediction formulas based on age, weight, and height of individual subjects. We demonstrate that the model fits data from CO rebreathing studies when diffusion of CO into the muscle compartment is considered. The model also fits responses of HbCO to single or multiple exposures to CO lasting for a few minutes each. In addition, the model reproduces reported differences between arterial and venous HbCO levels and replicates predictions from the Coburn-Forster-Kane equation for CO exposures of a 1- to 83-h duration. In contrast to approaches based on the Coburn-Forster-Kane equation, the present model predicts uptake and distribution of CO in both vascular and tissue compartments during inhalation of either constant or variable levels of CO.

Adult↗
PubMed2003

Minichromosome maintenance (Mcm) proteins, cyclin B1 and D1, phosphohistone H3 and in situ DNA replication for functional analysis of vulval intraepithelial neoplasia.

Vulval intraepithelial neoplasia (VIN) is defined histopathologically by distinctive abnormalities of cellular maturation and differentiation. To investigate the functional properties of VIN, the expression of several proteins involved in the regulation of the cell cycle as well as in situ DNA replication competence was analysed by immunohistochemistry. Snap-frozen vulval biopsies were graded as normal squamous epithelium (n=6), undifferentiated HPV positive VIN 1 (n=3), VIN 2 (n=8) and VIN 3 (n=20). Immunohistochemistry was performed using the following markers: cyclin D1 (expressed in middle/late G1), cyclin B1 (expressed in G2/early M), phosphorylated histone H3 (expressed during mitosis) and minichromosome maintenance (Mcm) proteins 2 and 5 (expressed during the cell cycle, but not in differentiated or quiescent cells). In situ DNA replication competence was used to identify S-phase cells. The percentage of positively stained nuclei in three representative microscopic fields was calculated per biopsy. In normal vulva, the expression of all markers was restricted to the proliferative compartment of the basal layer of the epithelium. In contrast in high-grade VIN, the majority of epithelial cells expressed the Mcm proteins from basal to superficial layer. The detection of cyclins B1 and D1, phospho-histone H3 and in situ DNA replication was also found through the full thickness of these lesions but by a lower proportion of the cells. This is consistent with these markers providing a series of 'snapshots' of the cell cycle status of individual cells. The low-grade VIN showed reduced expression of the cell cycle markers in relation to the level of dysplasia. The combination of these analyses establishes that the majority of VIN cells remain in a functional replicative or prereplicative state of the cell cycle. Clinical application of these analyses may provide a basis for improved diagnosis of VIN.

Biomarkers, Tumor↗
PubMed1994

Phosphatidylinositol-specific phospholipase C from Listeria monocytogenes contributes to intracellular survival and growth of Listeria innocua.

Listeria monocytogenes is a facultative intracellular organism that is capable of replicating within macrophage and macrophage-like cells. The species secretes a phosphatidylinositol-specific phospholipase C (PI-PLC) encoded by the plcA gene. A plcA gene from L. monocytogenes was cloned downstream of a gram-positive promoter in the plasmid pWS2-2. To determine what effect plcA would have on intracellular survival when introduced into Listeria innocua, a species that does not growth intracellularly or contain plcA, transformation with the recombinant pWS2-2 plasmid was performed. Phospholipase C activity in Listeria innocua/pWS2-2 was confirmed on a brain heart infusion-phosphatidylinositol agar plate, whereas wild-type L. innocua did not produce PI-PLC activity. Intracellular growth of L. innocua/pWS2-2 was subsequently measured in the macrophage-like cell line J774 by Giemsa staining and viable count determinations at specific time points following infection. The J774 cells infected with wild-type L. innocua showed a falling viable count through 8 h postinfection. Although J774 cells infected with L. innocua/pWS2-2 also initially displayed reduced viable counts, the viable count rose after 6 h postinfection and increased further at 8 h postinfection before a subsequent decline again at 16 h postinfection. Giemsa staining revealed fewer than 6 bacteria in individual macrophage cells at 2 h postinfection, and yet approximately 15% of the J774 cells had 6 to 12 bacteria localized to one area of the macrophage cell after 6 h; moreover, electron micrographs showed that the L. innocua/pWS2-2 cells were replicating inside the phagosome of the host cell. Furthermore, Thoria Sol labeling demonstrated that lysosomes had fused with these phagosomes, and acridine orange staining revealed that the compartments were acidified. These results demonstrate that L. innocua cells transformed with the plasmid-borne plcA gene, and expressing functional PI-PLC, are able to grow intracellularly in what appear to be phagolysosomes, although between 3 and 6 h is needed for this to manifest itself. Intracellular growth specifically in L. innocua may be a secondary function associated with the plcA gene product. The addition of this one gene, plcA, to a species of Listeria that in the wild-type state does not replicate intracellularly apparently can now allow some of the bacteria to transiently multiply inside the phagosomes of host macrophage cells.

Amino Acid Sequence↗
PubMed1986

Steroidogenic and morphological characteristics of granulosa and thecal compartments of the differentiating rabbit corpus luteum in culture.

On the day after ovulation, the thecal tissue and associated mural granulosa lutein cells of the rabbit corpus luteum were separated from the granulosa lutein 'core' by dissection and these tissues were cultured separately or together (whole corpus luteum) in defined medium for 10 days on stainless-steel grids. The medium was changed completely every 24 h. Replicate tissues were cultured with testosterone (10 ng/ml), but no other hormones were added to the medium. Progesterone production increased during the first 2 days of culture for whole corpus luteum, granulosa lutein cells and the thecal compartment which also included granulosa lutein cells. After 3 days, the production of progesterone declined gradually, but was still detectable on Day 10. The production of the metabolite, 20 alpha-dihydroprogesterone, by whole corpus luteum was equal to or greater than that of progesterone. Without the addition of testosterone, the granulosa lutein cells produced little (10 pg/culture) oestradiol during 1 day of culture, but the thecal compartment and whole corpus luteum each produced about 100 pg/culture on Day 1 and declining quantities over the next 2 days. In the presence of testosterone added to the medium, the formation of oestradiol was greatly increased for all tissues for 5-6 days of culture, after which time oestradiol was no longer detectable with or without testosterone in medium. Transmission electron microscopy of cells after 10-12 days of culture revealed fine structure that is characteristic of luteal cells, including abundant smooth endoplasmic reticulum, lipid droplets, and junctions between the luteal cells. The corpus luteum in culture resembles the corpus luteum in situ in that steroidogenesis and differentiation can proceed for a period after ovulation without extrinsic hormonal stimulation.

20-alpha-Dihydroprogesterone↗
PubMed2005

Coexistence and error propagation in pre-biotic vesicle models: a group selection approach.

Compartmentalization of unlinked, competing templates is widely accepted as a necessary step towards the evolution of complex organisms. However, preservation of information by templates confined to isolated vesicles of finite size faces much harder obstacles than by free templates: random drift allied to mutation pressure wipe out any template that does not replicate perfectly, no matter how small the error probability might be. In addition, drift alone hinders the coexistence of distinct templates in a same compartment. Here, we investigate the conditions for group selection to prevail over drift and mutation and hence to guarantee the maintenance and coexistence of distinct templates in a vesicle. Group selection is implemented through a vesicle survival probability that depends on the template composition. By considering the limit case of an infinite number of vesicles, each one carrying a finite number of templates, we were able to derive a set of recursion equations for the frequencies of vesicles with different template compositions. Numerical iteration of these recursions allows the exact characterization of the steady state of the vesicle population-a quasispecies of vesicles-thus revealing the values of the mutation and group selection intensities for which template coexistence is possible. Within the main assumption of the model-a fixed, finite or infinite, number of vesicles-we find no fundamental impediment to the coexistence of an arbitrary number of template types with the same replication rate inside a vesicle, except of course for the vesicle capacity. Group selection in the form of vesicle selection is a must for compartmentalized primordial genetic systems even in the absence of intra-genomic competition of different templates.

Animals↗
PubMed2001

Effects of Elevated Carbon Dioxide and Nitrogen Fertilization on Mycorrhizal Fine Roots and the Soil Microbial Community in Beech-Spruce Ecosystems on Siliceous and Calcareous Soil.

To study the responses of forests to global change, model ecosystems consisting of beech and spruce trees were maintained in open top chambers for 4 years under four conditions, namely with normal and elevated CO2 and with low and high nitrogen input, each replicated four times. Each open top chamber (height 3 m, diameter 3 m, soil depth 1.5 m) contained two separate soil compartments containing nutrient-poor siliceous and nutrient-rich calcareous soil. Here, we focus on the fine roots and the soil microbial community in these model ecosystems. At the time of planting, the fine roots were cut back according to forestry practice, and the newly formed roots were colonized by the indigenous soil microflora. After 4 years, the total biomass of fine roots, when averaged over all treatments, was almost the same in each of the two soil types; it was highest in the top 100 mm of soil (60%) and decreased sharply in deeper soil layers. Fungal biomass associated with the fine roots, consisting mainly of ectomycorrhizal fungi, was estimated using the ergosterol content as a marker. It was much higher in fine roots in the siliceous than in the calcareous soil, indicating considerably enhanced ectomycorrhiza formation in the nutrient-poor siliceous soil. Elevated atmospheric CO2 stimulated fine root production by ca. 85% and 43% in the top 100 mm of calcareous and siliceous soils respectively. Increased nitrogen input caused a slightly reduced production of fine root biomass in the calcareous soil but increased it by 33% in the siliceous soil. Marker substances for microorganisms were analyzed in the root-free soil. The amounts of carbon released by fumigation/extraction (a general marker for microbial biomass) and chitin (a marker for fungal biomass) were significantly higher in the top layer of the siliceous than of the calcareous soil, but they did not respond significantly to the treatments with elevated CO2 or the nitrogen fertilizer. The total concentration of the phospholipid fatty acids (PLFAs) and the number of the PLFAs did not differ between the two soil types. However, four of the eight most abundant PLFAs differed significantly between the two soil types. Principal component analysis revealed clearly separated clusters for the two soil types. Although analysis did not reveal any significant changes in response to the treatments, the concentration of the PLFA typical for ectomycorrhizal fungi was significantly higher under conditions of elevated CO2 in the nutrient-rich calcareous soil.

Journal Article↗
PubMed1995

Transcriptional inhibition of the parvovirus minute virus of mice by constitutive expression of an antisense RNA targeted against the NS-1 transactivator protein.

We have assessed a genetic resistance approach based on antisense RNA to interfere with the prototype Minute Virus of Mice (MVMp), an autonomous parvovirus. MVMp is a cytolytic virus when infecting the permissive A9 mouse fibroblast cell line, and its gene expression is largely regulated at the level of transcription initiation by the nonstructural transactivator NS-1 protein, a multifunctional polypeptide also involved in viral DNA replication and cytotoxicity. An NS-1 specific antisense RNA constitutively expressed in transfected A9 clones increased several fold the proliferative viability of the cells upon high multiplicity virus infection, and cultures infected at low multiplicity reached confluence overcoming virus progression. All clones shared a common phenotype of resistance characterized by a lowered synthesis of viral DNA replicative intermediates and genomic forms, a significant reduction in the accumulation of the three viral messengers in the cytoplasmic and nuclear compartments, and a specific inhibition in viral protein synthesis. These results indicate that the constitutive antisense RNA mediates an overall repression of viral macromolecular synthesis by preventing the onset of NS-1 functions. Therefore, cytocidal parvoviruses may be hampered by engineered antisense RNA targeted against early regulators of virus growth.

Animals↗
PubMed1996

Cell cycle regulation and cell type-specific localization of the FtsZ division initiation protein in Caulobacter.

Many genes involved in cell division and DNA replication and their protein products have been identified in bacteria; however, little is known about the cell cycle regulation of the intracellular concentration of these proteins. It has been shown that the level of the tubulin-like GTPase FtsZ is critical for the initiation of cell division in bacteria. We show that the concentration of FtsZ varies dramatically during the cell cycle of Caulobacter crescentus. Caulobacter produce two different cell types at each cell division: (i) a sessile stalked cell that can initiate DNA replication immediately after cell division and (ii) a motile swarmer cell in which DNA replication is blocked. After cell division, only the stalked cell contains FtsZ. FtsZ is synthesized slightly before the swarmer cells differentiate into stalked cells and the intracellular concentration of FtsZ is maximal at the beginning of cell division. Late in the cell cycle, after the completion of chromosome replication, the level of FtsZ decreases dramatically. This decrease is probably mostly due to the degradation of FtsZ in the swarmer compartment of the predivisional cell. Thus, the variation of FtsZ concentration parallels the pattern of DNA synthesis. Constitutive expression of FtsZ leads to defects in stalk biosynthesis suggesting a role for FtsZ in this developmental process in addition to its role in cell division.

Amino Acid Sequence↗
PubMed1997

Characteristics of the CD8+ lymphocytosis during primary simian immunodeficiency virus infections.

OBJECTIVE: To investigate the source of the expanded blood CD8+ subsets during an acute primary simian immunodeficiency virus (SIV) infection of macaques and the potential role of these cells in disease progression. DESIGN AND METHODS: The primary CD8+ lymphocytosis, which occurs at 1-2 weeks following infection with SIVsmm/PBj-14, was examined in rhesus and cynomolgus macaques. Extensive subset analysis of the expanded blood CD8+ cell pool in a rhesus macaque was compared phenotypically with those in thymus, lymph nodes, spleen, ileum and lung washouts obtained at necropsy during blood lymphocytosis. The influence of the primary CD8+ cells expansion on disease progression was assessed at days 175-679 post-infection in long-term PBj-14 survivors staged according to immunological, virological and histopathological changes in their lymphoid organs. RESULT: The very rapid and transient blood lymphocytosis following infection consisted of two distinct CD45RA(low), CD8+ and CD28-, lymphocyte function-associated antigen (LFA)-1(high), CD45RA(high), CD8+ populations. These populations were present in low levels in thymus, lymph and spleen but were highly represented in mucosal tissues, such as long washout, in which CD28- LFA-1(high) CD45RA(high) CD8+ cells comprised 86% of CD8+ cells, and gut, which was predominantly CD45RA(low) CD28- CD8+ cells. A comparison of progressor and non-progressor PBj-14-infected rhesus and cynomolgus macaques also indicated that the existence or magnitude of a blood CD8+ lymphocytosis during the acute phase of infection did not by itself appear to influence or be predictive of disease progression. CONCLUSION: The marked blood CD8+ lymphocytosis observed during acute SIV infection did not result from expansion of virus-specific precursors in peripheral lymph node and did not appear to influence the rate of disease progression. The findings provide a novel explanation for the primary CD8+ cell lymphocytosis and invoke a mechanism whereby virus-induced cytokine/chemokine production in mucosal sites initiate the transient migration of a pre-existing CD8+ population into the blood from compartments such as lung and gut. Such results suggest that the magnitude of lymphocytosis may depend on the level of viral replication in mucosal tissues and the presence of other infections, for example, cytomegalovirus.

Animals↗