Dormant Cysts Research Articles

Emergence of resistance to biocides during differentiation of Acanthamoeba castellanii.

A synchronous encystment method was used to study the order of development of resistance of Acanthamoeba castellanii to a range of biocides. The emerging resistance during encystation to short-term exposure to the minimum amoebicidal concentrations of each biocide tested was recorded during the first 36 h of the differentiation process. Hydrochloric acid and moist heat were tested as possible resistance markers. Development of the acid-insoluble, proteincontaining, ectocyst wall and the cellulose endocyst wall was followed by quantification of the acid- and alkali-insoluble residues of cell samples removed from synchronous encystment cultures up to 36 h. Resistance to chemical agents (polyhexamethylene biguanide, benzalkonium chloride, propamidine isethionate, pentamidine isethionate, dibromopropamidine isethionate, hydrogen peroxide) and to moist heat was seen to develop between 14 and 24 h after trophozoites were inoculated into the encystment media. Resistance to hydrochloric acid developed between 0 and 2 h and to chlorhexidine diacetate between 24 and 36 h. Levels of acid-insoluble residues began to increase after 8 h and alkali-insoluble residues (cellulose) were detected after 16 h and coincided with the emergence of resistance to all the agents tested except hydrochloric acid. The results suggest that resistance to the biocides tested probably results largely from the physical barrier of the cyst walls rather than as a consequence of a metabolically dormant cyst.

Cloning and Characterization of cDNAs Encoding Trehalase from Post-Dormant Embryos of the Brine Shrimp, Artemia franciscana

To investigate post-dormant regulation of trehalose metabolism in the brine shrimp, we cloned and characterized two trehalase cDNAs from embryos of Artemia franciscana using a PCR probe corresponding to a highly conserved region among trehalases. The cDNAs consisted of 2496 and 2485 nucleotides, and had almost the same open reading frame encoding 703 amino acids which showed 46.6–42.6% similarities to trehalases of animals. The calculated molecular mass of the trehalase was 79,995 Da. The deduced sequence had a cleavable signal peptide, a cell adhesion motif, four potential N-glycosylation sites, trehalase signatures and a unique, long carboxyl terminal polypeptide containing a predicted transmembrane region and a potential cAMP-dependent phosphorylation site. Phylogenetic analyses showed a large divergence among trehalases of arthropods. Northern blots revealed the presence of two mRNAs. One of them, a 2.6 kb mRNA, was abundant in the dormant cysts and prenauplii. The other 5.0 kb mRNA was newly synthesized during post-dormant development. Possible mechanisms of trehalase regulation are presented on the basis of the results shown by the Northern blots and developmental changes of trehalase activity.

Differentiation-associated surface antigen variation in the ancient eukaryote Giardia lamblia.

Encystation of Giardia lamblia is required for survival outside the host, whereas excystation initiates infection. The dormant cyst was considered an adaptation to external survival and passage through the stomach. However, we found previously that trophozoites which had recovered after completion of the life cycle had switched their major variant surface protein (VSP), called TSA 417, but neither the timing nor the molecular mechanism of switching had been elucidated. Here we demonstrate that TSA 417 predominates in cysts, but is downregulated during the stage of excystation that models cyst arrival in the small intestine. Transcripts of new VSPs appear late in encystation, and during and after excystation. Trophozoites appear to prepare for switching during encystation, when the major VSP on the cell surface diminishes and is internalized in lysosome-like vacuoles. As short-range DNA rearrangements were not detected, giardial VSP switching during differentiation appears to resemble the in situ switching of surface glycoproteins in African trypanosomes. We also report a unique extended 15 nucleotide polyadenylation signal in all VSP transcripts, but not in other known giardial genes. Antigenic variation during encystation-excystation may be a novel form of immune evasion that could help explain the common occurrence of reinfection by Giardia and other parasites with similar life cycles.

Cloning and sequencing of an α-tubulin cDNA from Artemia franciscana: evidence for translational regulation of α-tubulin synthesis

The brine shrimp, Artemia franciscana, exhibits a limited number of tubulin isotypes which change little during early postgastrula growth. In order to better understand the synthesis of α-tubulins during Artemia development, a cDNA termed αAT1 was cloned and sequenced. Alignment analyses revealed that the polypeptide encoded by αAT1 is similar to α-tubulins from other species. Hybridization of αAT1 to restriction-digested DNA on Southern blots produced a simple banding pattern, indicating that Artemia have a small number of α-tubulin genes. Probing of Northern blots demonstrated an abundant supply of α-tubulin mRNA in dormant cysts, emerging nauplii and instar I larvae. However, it was not until instar I larvae were produced that the amount of polysomal α-tubulin mRNA increased, suggesting that synthesis of the tubulin corresponding to αAT1 is translationally controlled. This work provides one of the few examples where tubulin synthesis is thought to be translationally regulated. Moreover, when considered in the light of previous analyses, the findings imply that cell differentiation in postgastrula Artemia and the diversification of microtubule function certain to accompany this process occur with little or no change in α-tubulin composition.

The effect of seasonal variability on the germination and vertical transport of a cyst forming dinoflagellate, Gyrodinium sp., in the Chesapeake Bay

Seasonal dinoflagellate blooms frequently occur in estuaries such as the Chesapeake Bay. Studies have shown that environmental factors such as temperature, salinity, light intensity, nutrient availability and physical mixing can affect the seasonal and vertical distribution of motile dinoflagellates. Furthermore, these environmental factors can affect the life history of cyst forming dinoflagellates by inducing the formation of dormant cysts in unfavorable conditions and by inducing cyst germination in favorable conditions. A generalized, dynamic model, developed in Stella II©, is proposed to examine combined effects of environmental processes on the life history and transport of a cyst forming dinoflagellate, Gyrodinium sp., in the Chesapeake Bay. The proposed model is arranged as a one-dimensional vertical column with no lateral migration or emigration to examine the role of local cysts in the formation of blooms. Two sampling stations provided by the EPA Chesapeake Bay Program, one at the mouth of the Potomac River (LE2.3) and the other in the main stem of the Chesapeake Bay (CB5.2), were used for comparison. Calibration was attempted using the Maryland Phytoplankton Taxon Survey, also provided by the EPA Chesapeake Bay Program, at MLE2.2 (a location upstream from station LE2.3). Observed cell concentrations for the winter and fall did not coincide with the cell concentrations calculated from the model for station LE2.3. This discontinuity suggests that cell migration from other portions of the Chesapeake Bay and the Potomac river could have contributed to the majority of observed winter and fall concentrations. Furthermore, observed cell concentrations at CB5.2 were the result of cell migration alone, due to the presence of a permanent pycnocline that inhibited cyst transport to the surface. The results of the proposed vertical transport model suggest that a three-dimensional model incorporating migration and emigration may be necessary in examining bloom formation.

Cellular and Transcriptional Changes during Excystation ofGiardia lamblia in Vitro

Hetsko, M. L., McCaffery, J. M., Svärd, S. G., Meng, T.-C., Que, X., and Gillin, F. D. 1998. Cellular and transcriptional changes during excystation ofGiardia lamblia in vitro. Experimental Parasitology88, 172–183. Excystation ofGiardia lambliaentails differentiation of dormant cysts into parasitic trophozoites. Despite its importance for infection, this transformation is not understood at the cellular or molecular levels. In these studies, we report that excystation entails detection of environmental stimuli across the tough extracellular cyst wall leading to highly coordinated physiological, structural, and molecular responses. We found that novel cytoplasmic rearrangements and changes in individual species of mRNA and in cytoplasmic pH occur within the cyst wall in the earliest stage of excystation, in response to conditions modeling cyst ingestion and passage into the human stomach. This suggests that cysts do not contain all the mRNA needed for excystation and emergence and supports ourhypothesisthat external stimuli, including hydrogen ions, may penetrate or be perceived across the cyst wall. In contrast, changes in cyst wall structure or proteins were detected only later in excystation, in the stage that models passage into the human small intestine, where trophozoites can emerge and survive. These findings show that excystation ofG. lambliais a highly complex and active process and provide important insights into its cellular and molecular components.

Mechanisms of innate resistance to Toxoplasma gondii infection.

The interaction of protozoan parasites with innate host defences is critical in determining the character of the subsequent infection. The initial steps in the encounter of Toxoplasma gondii with the vertebrate immune system provide a striking example of this important aspect of the host-parasite relationship. In immuno-competent individuals this intracellular protozoan produces an asymptomatic chronic infection as part of its strategy for transmission. Nevertheless, T. gondii is inherently a highly virulent pathogen. The rapid induction by the parasite of a potent cell-mediated immune response that both limits its growth and drives conversion to a dormant cyst stage explains this apparent paradox. Studies with gene-deficient mice have demonstrated the interleukin-12 (IL-12)-dependent production of interferon gamma (IFN-gamma) to be of paramount importance in controlling early parasite growth. However, this seems to be independent of nitric oxide production as mice deficient in inducible nitric oxide synthase (iNOS) and tumour necrosis factor receptor were able to control early growth of T. gondii, although, they later succumbed to infection. Nitric oxide does, however, seem to be important in controlling persistent infection; treating chronic infection with iNOS metabolic inhibitors results in disease reactivation. Preliminary evidence implicates neutrophils in effector pathways against this parasite distinct from that described for macrophages. Once initiated, IL-12-dependent IFN-gamma production in synergy with other proinflammatory cytokines can positively feed back on itself to induce 'cytokine shock'. Regulatory cytokines, particularly IL-10, are essential to down-regulate inflammation and limit host pathology.

Cellular immune reactions directed against Toxoplasma gondii with special emphasis on the central nervous system.

Toxoplasma gondii is an obligate intracellular parasite which, after primary infection of humans, is maintained in a dormant state by the host cellular immune system. In the event of an acquired immunosuppression, those parasites surviving as dormant cysts in the host may undergo a change in status, proliferate and cause a life-threatening toxoplasmic encephalitis. Over the last decade much knowledge has accumulated concerning the immune response against T. gondii. This review focuses attention particularly on the anti-parasitic effector mechanisms and the cellular immune reactions in the central nervous system during the course of reactivated toxoplasmic encephalitis.

Segmentation in the crustacean Artemia: engrailed staining studied with an antibody raised against the Artemia protein.

We have studied the process of post-embryonic segmentation in the anostracan crustacean Artemia franciscana using a specific antibody raised against the engrailed protein of this organism. Three cephalic segments are specified during embryonic development, before larval hatching, whilst trunk (thoracic) segmentation begins after the first stage free-swimming nauplius larva has emerged from the dormant cyst. Thus, cephalic and trunk segmentation seem to be at least in part independent and superimposed processes. Trunk stripes of engrailed expression are added one at a time as segments are generated from the posterior growth zone. The first detectable decision in engrailed expression is the establishment of a line of engrailed-expressing cells, interpretable as delineating the parasegmental boundary. The subsequent widening of engrailed stripes is not correlated with cell lineage events but is probably mediated by the combination of inheritance of the active state and recruitment of new cells into the engrailed-expressing stripe.

Disturbance and the Species Composition of Rain Pool Microbial Communities

Ephemeral rain pools contain diverse assemblages of protists and small metazoans. Rain pool organisms survive periods of desiccation as dormant cysts or resting stages, and become active shortly after the pools fill with water. Non-equilibrium mechanisms related to the frequent drying of pools could promote the observed coexistence of over 15 species of ecologically similar microorganisms. We used dormant organisms from the substrate of a well-characterized ephemeral pool to create 20 microcosms that experienced two different disturbance regimes. Undisturbed microcosms held a constant volume of medium over the entire 30-d study period. Disturbed microcosms dried periodically, at approximately five-d intervals, and were then refilled with medium to initiate new rounds of community development. Daily monitoring of the presence or absence of active organisms provided information about the number and identity of species in disturbed and undisturbed microcosms. Undisturbed microcosms contained a greater number of active species, primarily because some predatory species only became active after at least five to ten d of uninterrupted community development. Species composition diverged rapidly after the first drying episode as communities in undisturbed and disturbed microcosms developed along different successional trajectories. If non-equilibrium mechanisms contribute to diversity in this system, their contribution must occur over more episodes of disturbance than we observed.

Co-infection of Toxoplasma gondii with other pathogens: pathogenicity and chemotherapy in animal models.

Toxoplasma gondii is a protozoan parasite that is normally controlled by the host immune system and results in an asymptomatic chronic infection maintained by dormant cysts. Actually, animals and humans are simultaneously infected with T. gondii and by various micro-organisms during their life, and raise a specific protective immune response against each pathogen. The advent of immunosuppression related to concurrent viral infection such as human immunodeficiency virus (HIV) in humans or to the use of intensive immunosuppressive therapies is responsible for a profound alteration of protective immunity, and this often results in the recurrence of latent infections. Cerebral toxoplasmosis is one of the most common infections occurring in AIDS patients, especially in countries where T. gondii is highly endemic. However, other opportunistic infections due to protozoa, bacteria, and fungi are also commonly observed in these patients. Thus, co-infection with various pathogens is usual in patients with acquired immunodefiency syndrome (AIDS), and several lines of evidence suggest that interactions between the immune responses generated by each pathogen, due to cross-regulation of Th1- and Th2-activated CD4+ T lymphocytes, macrophages, neutrophils and natural killer (NK) cells can result in an aggravation of one or more infections (Mosier 1994). Furthermore, several of the pathogens infecting AIDS patients are intracellular (including HIV, T. gondii, Mycobacterium avium-intracellulare, MAI, and cytomegalovirus, CMV), giving rise to the hypothesis that possible interactions between these pathogens within the cell may occur and result in alterations of cell functions.

Acanthamoeba

Acanthamoeba is a small, free-living amoeba which is characterized by a feeding and replicating trophozoite and dormant cyst stage. The resistance of acanthamoeba cysts to extremes of temperature, disinfection and desiccation accounts for the ubiquitous distribution of the organism in our environment. Acanthamoeba produces two distinct forms of disease in man. Granulomatous amoebic encephalitis (GAE) is a chronic and invariably fatal infection occurring in immunocompromized persons. Since GAE was first recognized in 1971, approximately 50 cases have been reported worldwide. In contrast, keratitis due to Acanthamoeba affects previously healthy persons and occurs with far greater frequency. Since the disease was first described in 1973, many hundreds of cases of acanthamoeba keratitis have been reported worldwide. Approximately 80–90% of these have been in wearers of contact lenses. Acanthamoeba keratitis is perhaps the most difficult ocular infection to manage successfully and can result in permanently impaired vision or blindness. Acanthamoeba keratitis continues to be reported with increasing frequency, at least 90 cases having occurred in the United Kingdom since 1974 and, of these, 63 were recorded in the past 3 years.

Life history divergence of sympatric diploid and polyploid populations of brine shrimp Artemia parthenogenetica.

In order to study how polyploidy affects life history patterns in animals, we have examined sympatric diploid and polyploid brine shrimp (Artemia parthenogenetica) from China, Italy and Spain under laboratory conditions. At optimal temperature and salinity (25°C and 90 ppt), diploids from the three populations had much higher intrinsic rates of increase, higher fecundity, faster developmental rates, and larger brood sizes than their sympatric polyploids. The Chinese and Italian populations were selected for further analysis to determine the life history responses of diploids and polyploids to temperature and salinity changes. Under intermediate and high salinities, Chinese and Italian polyploids produced most of their offspring as dormant cysts while their sympatric diploids produced most of their offspring as nauplii. This relationship is reversed in the Spanish diploid-polyploid complex. For the Chinese population at 25° C, pentaploid clones had higher developmental rates than diploid clones at 35 ppt; at 90 ppt, diploid clones had higher developmental rates than the pentaploids. Italian diploids and tetraploids had different responses to variation in both temperature (25° C and 31° C) and salinity (30 ppt and 180 ppt). Our results demonstrate that relative fitness of the two cytotypes is a function of environmental conditions and that sympatric diploids and polyploids respond differently to environmental changes. Chinese and Italian polyploids are expected to have lower fitness than their sympatric diploids when the physical environment is not stressful and when intraspecific competition is important. However, polyploids may have advantages over sympatric diploids in stressful habitats or when they encounter short-term lethal temperatures. These results suggest that polyploid Artemia have evolved a suite of life-history characteristics adapting them to environments that contrast to those of their sympatric diploids.

Intracellular water in Artemia cysts (brine shrimp): Investigations by deuterium and oxygen-17 nuclear magnetic resonance

The dormant cysts of Artemia undergo cycles of hydration-dehydration without losing viability. Therefore, Artemia cysts serve as an excellent intact cellular system for studying the dynamics of water-protein interactions as a function of hydration. Deuterium spin-lattice (T(1)) and spin-spin (T(2)) relaxation times of water in cysts hydrated with D(2)O have been measured for hydrations between 1.5 and 0.1 g of D(2)O per gram of dry solids. When the relaxation rates (I/T(1), I/T(2)) of (2)H and (17)O are plotted as a function of the reciprocal of hydration (1/H), an abrupt change in slope is observed near 0.6 g of D(2)O (or H(2) (17)O)/gram of dry solids, the hydration at which conventional metabolism is activated in this system. The results have been discussed in terms of the two-site and multisite exchange models for the water-protein interaction as well as protein dynamics models. The (2)H and (17)O relaxation rates as a function of hydration show striking similarities to those observed for anisotropic motion of water molecules in protein crystals.It is suggested here that although the simple two-site exchange model or n-site exchange model could be used to explain our data at high hydration levels, such models are not adequate at low hydration levels (<0.6 g H(2)O/g) where several complex interactions between water and proteins play a predominant role in the relaxation of water nuclei. We further suggest that the abrupt change in the slope of I/T(1) as a function of hydration in the vicinity of 0.6 g H(2)O/g is due to a change in water-protein interactions resulting from a variation in the dynamics of protein motion.

Intracellular polyamine patterns during encystment of Physarum flavicomum

In Physarum flavicomum Berk., growing amoebae convert to dormant cysts under conditions of nutrient imbalance. Exogenous adenine inhibits the process and the cells produce an elevated intracellular concentration of S-adenosylmethionine. Evidence indicates that the increased level of S-adenosylmethionine is responsible for the disruption of the normal developmental process. One of the biological functions of S-adenosylmethionine is in polyamine synthesis and it is known that a well-controlled intracellular concentration of polyamines is essential for normal cell growth and differentiation. In this study, high-performance liquid chromatography was used to determine the intracellular polyamine patterns in growing cells, adenine-treated and normal encysting cells, and dormant cysts. Putrescine and spermidine were the most abundant polyamines found in the cells; growing cells had the highest level, adenine-treated cells had a 1.5 to 2.0 times higher level than normal encysting cells, while cysts had the lowest (only 3 and 12% of that of growing cells). Cadaverine and N1-acetylspermidine were found in all the cells and their levels decreased during encystment. Acetylputrescine was found in growing cells only and acetylspermine was found in all cells except cysts. Acetylcadaverine, N8-acetylspermidine, 1,3-diaminopropane, and spermine were not detected in any of the cells.

Dolichol phosphorylation occurs via a CTP‐dependent reaction in Artemia larvae

The phosphorylation of dolichol in larval stages of the brine shrimp, Artemia salina, has been investigated. The dolichol kinase has been assayed in crude microsomes; the enzyme requires CTP as phosphoryl donor and calcium as divalent cation. Activity increases with both incubation time and added microsomal protein. The product of the reaction has been characterized by chromatographic and enzymatic procedures. With gamma-32P CTP as substrate, the apparent Km for CTP is 24 microM. Enzymatic activity is stimulated fivefold by exogenous dolichol. The specific activity of the enzyme increases with the frequency of molting. Dolichol kinase activity was detectable in membranes prepared from dormant Artemia cysts. The low level in dormancy may anticipate the critical role of the enzyme during hatching.

Molecular Biology of DNA inAcanthamoeba, Amoeba, Entamoeba, and Naegleria

This chapter discusses the molecular biology of DNA in acanthameba, ameba, entameba, and naegleria. The chapter reviews a reasonably current review about gene structure and the overall organization of the DNA and its metabolism. The four genera of amebas that have been studied most extensively are described. The genus ameba includes large free-living organisms. The cell cycle is simple; replication is by binary fission and differentiation is unknown. These amebas are large enough to manipulate by various surgical procedures, and this attribute has been exploited extensively. Cultures typically are grown using the ciliated protozoan tetrahymena as the major food organism. The genus acanthameba includes small amebas that are ordinarily free-living, but can be opportunistic pathogens of animals and humans. Such organisms are considered amphizoic. In a few cases, infection has been lethal, but these organisms are ubiquitous in nature and most humans appear to have good resistance to infections. The taxonomy of acanthameba is confusing, and it is unclear whether pathogenicity is restricted to certain species or whether all strains are capable of it. acanthameba replicate by binary fission. They differentiate into dormant cysts (that is, they encyst) during adverse environmental conditions. The genus naegleria includes small amebas that can differentiate either into a cyst or a flagellated stage. In naegleria, it is clear that certain species are pathogenic and that other species are not. The pathogenic Naegleria fowleri is highly virulent and fatal to humans. The genus entameba is distinct from the rest because it includes anaerobes and parasites. The life cycle includes a phase of replication by binary fission and encystment, which occurs in response to environmental factors. Nuclear division can continue in the cyst in the absence of cytoplasmic division. Species are typically identified according to their hosts. Entameba histolytica, the human parasite, and Entameba invadens, a parasite of reptiles, are the two most commonly studied species. Both can be grown axenically under anaerobic conditions. Entameba moshkovskii is interesting because it appears to be free-living.

Molecular cloning and analysis of cDNA sequences for two ribosomal proteins from Artemia. The coordinate expression of genes for ribosomal proteins and elongation factor 1 during embryogenesis of Artemia.

The large subunit of eukaryotic ribosomes contains acidic phosphoproteins which are related to L7/L12 from Escherichia coli. In the brine shrimp Artemia these proteins are designated eL12 and eL12'. We have isolated cDNA clones for these proteins from a cDNA bank that was constructed by the use of size-fractionated poly(A)-rich RNA (8-10S fraction) from Artemia and a synthetic oligonucleotide as primer. Clones containing DNA sequences coding for eL12 and eL12 were characterized by hybrid-selected translation and DNA sequencing. The proteins eL12 and eL12' share an identical peptide of 22 amino acids at their carboxy termini whereas the remaining part of the protein shows little sequence homology. The nucleotide sequences show a different codon use for the amino acids in the common carboxy terminus, thereby excluding a common exon coding for this part of both proteins. Despite the differences in amino acid sequence in the major part of eL12 and eL12' the proteins have a considerable degree of homology on the basis of the distribution of hydrophobic and hydrophilic amino acids over the polypeptide chains, in agreement with a related folding and function of both proteins. Relative levels of mRNA coding for eL12, eL12' and elongation factor 1 alpha were determined during the development of Artemia from a dormant cyst to a nauplius. The data show a coordinate expression of the genes for EF-1 alpha and both ribosomal proteins, excluding a differential expression of the genes for these related ribosomal proteins during embryogenesis. Analysis of the gene copy number for eL12 and eL12' indicates the presence of a few genes for each protein.

Purification and characterization of a cytosol protease from dormant cysts of the brine shrimp Artemia.

A thiol protease has been isolated and purified from the postribosomal fraction of encysted embryos of the brine shrimp Artemia using a six-step procedure. The purified enzyme has a molecular weight of 55,000 +/- 4,200 and is composed of subunits of Mr 31,500 +/- 559 and 25,867 +/- 1,087. Isoelectric focusing revealed two discrete bands, one at pH 4.6 and the other at pH 5.1. The protease appears to be a member of the thiol group of proteases based on its inhibition by leupeptin, antipain, chymostatin, Ep-475, and several other thiol protease inhibitors. The enzyme was stimulated by heavy metal chelators and thiol reagents. At pH 3.5-4.0 the thiol protease hydrolyzed a wide range of proteins including bovine serum albumin, hemoglobin, Artemia embryo soluble proteins, Artemia lipovitelline, and protamine, whereas at pH 6.0-6.5 the enzyme showed a high degree of specificity for Artemia elongation factor 2 and lipovitelline alpha 1. The total amount of protease activity in crude homogenates of Artemia embryos decreased by about 50% during the first 24 h of development, while the amount of free, active enzyme decreased proportionally for 9 h of development then remained constant during the next 26-27 h of development. These changes in protease activity appear to reflect changing levels of an endogenous protease inhibitor during development.

Differentiation of pathogenic amoebae: encystation and excystation ofAcanthamoeba culbertsoni — A model

Differentiation into dormant cysts and vegetative trophozoites is an inherent character intimately associated with the life cycle and infectivity of pathogenic amoebae. In the case of human intestinal amoebiasis encystation and excystation are of immediate relevance to the process of transmission of the disease from healthy carriers to susceptible individuals. Using a pathogenic free living amoebaAcanthamoeba culbertsoni as a model, considerable progress has been achieved in understanding the mechanism and control of the process of differentiation. The turnover of the regulatory molecule cyclic 3: ’5′ adenosine monophosphate is responsible for triggering the process of encystation. Amoebae bind effector molecules such as biogenic amines to a membrane localized receptor which itself resembles the β-adrenergic receptor of mammalian organisms. The activation of adenylate cyclase or inhibition of cyclic AMP phosphodiesterase maintain the dynamic intracellular cyclic AMP. The cytosol fraction of amoebae has a cyclic AMP binding protein. During encystation, enzymes needed for synthesis of cellulose and glycoproteins are induced. Control is exercised at transcriptional level and the process is subject to catabolic repression.