The microsporidian spore invasion tube. II. Role of calcium in the activation of invasion tube discharge

A swelling response by the polaroplast organelle initiated microsporidian invasion tube extrusions by Glugea hertwigi spores. The tumescence was induced by the displacement of internal calcium. Sodium citrate, phosphate, and the calcium ionophore A23187 were effective in initiating polaroplast swelling and spore discharge; however, the addition of external CaCl2 switched the expanded polaroplasts to a contracted state and blocked spore discharge. Unlike CaCl2, equivalent concentrations of KCl, NaCl, MgCl2, and BaCl2 did not induced polaroplast contraction, and spore discharge was not blocked. 45CaCl2 readily incorporated into spores with expanded polaroplasts; however, little calcium uptake was apparent in spores with contracted polaroplasts. Metallochromic arsenazo III yielded a color spectrum characteristic of the dye-Ca++ complex in the polaroplast region; furthermore, a membrane association with calcium was indicated by strong chlorotetracycline fluorescence within the polaroplast; this fluorescence was extinguished by pretreating spores with ionophore {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187. An association of the membrane with calcium was also indicated by a potassium ferrocyanide-osmium tetroxide technique. All evidence indicates that an internal calcium displacement is an important initial step in the swelling response of the polaroplast organelle.     

Review: Toxoplasma gondii cellular invasion.

Toxoplasma gondii, the etiologic agent of toxoplasmosis, is a ubiquitous protozoan parasite that requires an intracellular site for growth and replication. The invasive process involves six steps: a) cellular recognition, b) parasite movements by means of a subpellicular microtubule cytoskeleton, c) cell to cell adhesion, d) rhoptry secretion of penetrating enhancing factor (PEF) with Ca++ and Ca++ activated ATPase dependence, e) conoid penetration, f) induction of a parasitophorous vacuole, a protective and exchange site, interiorization of the parasite. The invasion is an active, oriented and specific process depending on chemical factors as energy sources, cations, as well as microviscosity and membrane structures. Toxoplasma gondii stimulates T cell subsets and induces lymphokine (IFN gamma, IL2) release.     

Cell adhesion in invasion and metastasis.

Metastatic cells exhibit considerable flexibility in their adhesive interactions with other cells or components of the extracellular matrix. This review will describe the involvement of specific adhesion receptors, extracellular matrix molecules and cell dissociating cytokines in the metastatic cascade. We will particularly focus on disturbance of intercellular adhesion as a prerequisite for the release of invasive cells from carcinomas. We suggest that cell dissociation in these tumours is accomplished by loss of function or expression of the epithelial cell adhesion molecule E-cadherin, and through the activity of cell motility factors such as the scatter factor.     

Inhibition of tumor cell invasion by verapamil.

Verapamil, a calcium channel antagonist, inhibits murine B16 melanoma and colon adenocarcinoma C26 tumor metastasis by altering platelet aggregation [Tsuruo, T., et al. (1985) Cancer Chemother. Pharmacol., 14:30-33]. However, the role of calcium homeostasis in regulating several biochemical pathways implicated in other steps of the metastatic cascade suggests that calcium channel antagonists could also inhibit metastasis by other mechanisms. In this report, non-toxic doses of verapamil reversibly decreased human A375M and C8161 melanoma cell invasion and metastasis in a dose-dependent manner. Verapamil reduced cellular invasion and metastases by up to 96% (range 78-96%). Concomitantly, verapamil disrupts microtubule and microfilament organization and inhibits unidirectional cell migration but does not affect cellular adhesion to endothelial monolayers or reconstituted basement membranes. In addition, tumor cells treated with verapamil have a decrease in mRNA of type IV collagenase, a proteinase important in tumor cell degradation of basement membranes. Collectively, these data offer additional evidence regarding the mechanisms of action of verapamil as an anti-metastatic agent.     

Mechanism of control of trophoblast invasion in situ.

We have previously shown that first trimester human trophoblast cells share in vitro invasive properties with malignant cells. In this study we show that the in situ control of trophoblast invasion is provided by the uterine microenvironment. Trophoblast cells were labeled with 125I-deoxyuridine and examined for their ability to invade an epithelium-free human amniotic membrane in vitro under various conditions. The degree of invasion was determined as the percentage of the radioactivity retained within the membrane. Conditioned media from first trimester human decidual cells (DCM) suppressed invasion of trophoblast cells in the amnion invasion assay. This suppression was prevented by addition of neutralizing anti-TGF beta antibody or neutralizing antibody to tissue inhibitor of metalloproteinases (TIMP-1) to the DCM, and mimicked by TGF beta 1. These antibodies also augmented invasion beyond control levels, suggesting that trophoblast cells may also produce these factors. A bioassay for TGF beta activity, measured by antiproliferative effect on the mink lung epithelial cell line Mv 1 Lu, revealed that decidual cells produced this factor only in the latent form, whereas the active form was produced by the trophoblast. A decrease in collagenase type IV activity in the conditioned media of trophoblast cultures was observed when TGF beta 1 was added to these cultures. Removal of endogenous TGF beta in trophoblast cultures by addition of anti-TGF beta antibody resulted in down-regulation of TIMP message as determined by Northern analysis. These results indicate that a) decidua-derived (and to a minor extent trophoblast-derived) TGF beta is the prime mediator in the control of invasion by first trimester trophoblast, the latent form of TGF beta likely being activated by trophoblast-derived proteinases; b) induction of TIMP by TGF beta in both trophoblast and decidua is the final pathway in this control.     

Streptococcal invasion

The genus Streptococcus consists of large number of species many of which are pathogenic to humans and animals. Although streptococci have long been considered as extracellular pathogens, they are capable of causing serious invasive infections such as necrotizing fasciitis and meningitis. Streptococcal invasion, therefore, has been a focus of many studies in recent years. Streptococci are efficiently internalized by nonprofessional phagocytes and the current research interest has shifted to determine the role of this invasion in the natural infection process. Moreover, characterization of bacterial and eukaryotic components involved in the uptake process might be useful in developing new strategies for combating streptococcal infections.     

沙门菌侵袭研究进展

鼠伤寒沙门菌表达两个不同的Ⅲ型分泌系统(typeⅢsecretion/translocation systems, TTSS),分别由致病岛1和2(pathogenicityi slands 1 and 2, SPI-1 and SPI-2)编码。细菌依赖TTSS将效应蛋白转运至宿主细胞,通过“触发”机制诱导细菌进入宿主细胞。这些效应蛋白可诱导细胞骨架重排,导致“巨吞饮”,促使细菌入侵。本综述依据多种沙门菌效应蛋白的功能,建立沙门菌侵袭模型。TTSS活化并转运效应蛋白进入宿主细胞发挥功能(Ⅰ)。小G蛋白交换因子SopE和肌醇磷酸酯酶SopB通过激活CDC42和Rac1,诱导内陷相关的蛋白聚集(Ⅱ)。SipA和SipC通过降低肌动蛋白临界浓度、刺激网素成束、稳定纤维状肌动蛋白(fibrousactin, F-actin)以及使肌动蛋白核化等功能,促使细菌入侵(Ⅲ)。SopB可使膜内陷区PIP2的浓度降低以及VAMP8聚集,促使细胞膜分裂(Ⅳ)。这些效应蛋白的联合作用,使膜皱褶在局部向外显著延伸,使沙门菌被细胞内形成的特殊膜结构包裹。沙门菌的另一种效应蛋白SptP,通过刺激小G蛋白内源性GTPase的活性,抑制小G蛋白的活化,使细胞膜恢复至原有状态(Ⅴ)。     

The microsporidian spore invasion tube. III. Tube extrusion and assembly

The polar filaments within microsporidian spores discharges as tubes with subsecond velocity. Populations of discharging tubes of Glugea hertwigi spores pulse-labeled with latex particles for 1-3 s were consistently devoid of label at the distal ends; discharging tubes were completely labeled after 30- to 60-s exposure to latex. This experiment indicates that discharge tubes grow at the tip. Completely assembled discharge tubes consisted of single, empty cylinders; however, incompletely discharged tubes had a cylinder-within-a-cylinder profile at the distal ends. This observation indicates that the discharge tube material emerges at the distal end by an eversion process. Finally, studies with cinematic Nomarski interference optics of spore tubes extruding across a water-air interphase indicate that all the material emerging from the growing tip of the tube is incorporated into the wall of the discharge tube. Evidence indicates that the polar filament of undischarged spores is a homogeneous coil of polar tube protein equivalent to the polar tube protein in discharged tubes.     

Assay for adhesion and agar invasion in S. cerevisiae

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, biofilms can reduce industrial productivity, destroy structures, and threaten human life. 1-3 On the other hand, harnessing the power of biofilms can help clean the environment and generate sustainable energy. 4-8 The ability of S. cerevisiae to colonize surfaces and participate in complex biofilms was mostly ignored until the rediscovery of the differentiation programs triggered by various signaling pathways and environmental cues in this organism. 9, 10 The continuing interest in using S. cerevisiae as a model organism to understand the interaction and convergence of signaling pathways, such as the Ras-PKA, Kss1 MAPK, and Hog1 osmolarity pathways, quickly placed S. cerevisiae in the junction of biofilm biology and signal transduction research. 11-20 To this end, differentiation of yeast cells into long, adhesive, pseudohyphal filaments became a convenient readout for the activation of signal transduction pathways upon various environmental changes. However, filamentation is a complex collection of phenotypes, which makes assaying for it as if it were a simple phenotype misleading. In the past decade, several assays were successfully adopted from bacterial biofilm studies to yeast research, such as MAT formation assays to measure colony spread on soft agar and crystal violet staining to quantitatively measure cell-surface adherence. 12, 21 However, there has been some confusion in assays developed to qualitatively assess the adhesive and invasive phenotypes of yeast in agar. Here, we present a simple and reliable method for assessing the adhesive and invasive quality of yeast strains with easy-to-understand steps to isolate the adhesion assessment from invasion assessment. Our method, adopted from previous studies, 10, 16 involves growing cells in liquid media and plating on differential nutrient conditions for growth of large spots, which we then wash with water to assess adhesion and rub cells completely off the agar surface to assess invasion into the agar. We eliminate the need for streaking cells onto agar, which affects the invasion of cells into the agar. In general, we observed that haploid strains that invade agar are always adhesive, yet not all adhesive strains can invade agar medium. Our approach can be used in conjunction with other assays to carefully dissect the differentiation steps and requirements of yeast signal transduction, differentiation, quorum sensing, and biofilm formation.     

Strand invasion by DNA-peptide conjugates and peptide nucleic acids.

Peptide nucleic acids (PNAs) and conjugates between oligonucleotides and cationic peptides possess superior potential for strand invasion at complementary sequences. We discovered that oligonucleotide-peptide conjugates and PNAs fall into three classes based on their hybridization efficiency; i) those complementary to inverted repeats within AT-rich region hybridize with highest efficiency; ii) those complementary to areas adjacent to inverted repeats or near AT-rich regions hybridize with moderate efficiency; and iii) those complementary to other regions do not detectably hybridize. The correlations between oligomer chemistry, DNA target sequence, and hybridization efficiency that we report here have important implications for the recognition of duplex DNA.     

A silent invasion

Invasions mediated by humans have been reported from around the world, and ships’ ballast water has been recognized as the main source of marine invaders worldwide. Some invasions have dramatic economic and ecological consequences. On the other hand, many invasions especially in the marine realm, can go unnoticed. Here we identify a human mediated, worldwide introduction of the hydrozoan species Turritopsis dohrnii. The normal life cycle of hydrozoans involves the asexual budding of medusae from colonial polyps. Medusae of Turritopsis, however, when starved or damaged, are able to revert their life cycle, going back to the polyp stage through a process called transdifferentiation. They can thus easily survive through long journeys in cargo ships and ballast waters. We have identified a clade of the mitochondrial 16S gene in Turritopsis which contains individuals collected from Japan, the Pacific and Atlantic coasts of Panama, Florida, Spain, and Italy differing from each other in only an average of 0.31% of their base-pairs. Fifteen individuals from Japan, Atlantic Panama, Spain, and Italy shared the same haplotype. Turritopsis dohrnii medusae, despite the lack of genetic differences, are morphologically different between the tropical and temperate locations we sampled, attesting to a process of phenotypic response to local conditions that contributes to making this grand scale invasion a silent one.     

Small-step invasion research

Recent invasion research can be categorized broadly into two types: studies of the spatial spread, and those of the biological impact the invader has on the native biota. The first type is the most factual, and the second the most theoretical. So far, however, it is difficult to connect the two, implying that neither the spatial spread nor the species interactions can be explained in terms of each other. Recent models, analysing spatial spread, progress steadily by making small steps.