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The article overviews some issues of the severe course of tropical malaria. In addition to the analysis of the ongoing situation with malaria in Russia, a general clinical picture of the severe course of tropical malaria is discussed. The main part of the overview includes a detailed analysis of current data on the molecular genetic aspects of the erythrocytes' adhesion in the case of tropical malaria. The main elements involved in the process of binding red blood cells and, as a result, in the process of their adhesion to other cells of the human body were considered in detail. Data were studied and summarized not only on protein interactions between an infected red cell and its cellular environment, but also on the genetic characteristics of the parasite leading to similar molecular-biological processes. In addition to the study of protein PfEMP1 role which is nowadays well-considered in the literature, the most up-to-date but less reported data on erythrocyte adhesion proteins STEVOR and RIFIN were also included. The team of authors hopes that this publication will help to get a deeper insight into the problem of erythrocyte adhesion in the course of complicated malaria infection forms and to summarize some of the available data on this issue.

Ran is an evolutionarily conserved GTPase crucial in regulating various cell divisions, including mitosis and meiosis. A previous study showed that the knockdown of RAN1 inhibited macronuclear amitosis with the abnormal organization of intramacronuclear microtubules in Tetrahymena thermophila. This study aimed to further investigate the effects of the inducible expression of wild-type Ran1 (Ran1WT), GTP-bound Ran1-mimetic (Ran1Q70L), and GDP-bound Ran1-mimetic (Ran1T25N) on cytoplasmic microtubule assembly during amitosis of T. thermophila, based on previous studies about their effects on intramacronuclear microtubule. The mutant strains of T. thermophila for inducible expression of Ran1WT/T25N/Q70L by Cd^(2+) were constructed. The inducibly expressed HA-Ran1Q70L/T25N distributed asymmetrically across the macronuclear envelope during amitosis. At the lower level of inducible expression, only Ran1T25N showed a significant decreasing effect on T. thermophila reproduction, macronuclear amitosis and cytokinesis. At the higher level of inducible expression, Ran1WT/Q70L/T25N inhibited T. thermophila reproduction, macronuclear amitosis and cytokinesis, and the inhibitive effect of Ran1T25N was the most significant. The inducible expression of Ran1WT/Q70L/T25N led to defects in amitosis and cytokinesis with abnormal cytoplasmic microtubule assembly. These results further confirmed the regulatory function of Ran1 on amitosis and suggested a novel role of Ran1 in cytokinesis and the alignment of cytoplasmic microtubules in T. thermophila.

Using the Physarum polycephalum, plasmodium, a giant amoeboid cell with the strongly pronounced auto-oscillatory mode of motility, which exhibits regularities of motile behavior common with those of tissue cells and has the same signal systems, the possibility of the participation of phosphatidylinositol-4,5-bisphosphate in the regulation of the contractile activity has been studied. The effect of neomycin as a substrate inhibitor of phospholipase C, which binds with high affinity to phosphatidylinositol-4,5-bisphosphate in the membrane, on force oscillations generated by plasmodial strands under isometric conditions and after the addition of the protein kinase C inhibitors staurosporine, UCN-01, and Ro-318220, separatelyand in combination with the calmodulin inhibitor calmidazolium has been examined. It has been shown that neomycin at pH 7.0 and concentrations of 0.1-5.0 mM stops contractile oscillations for 10-30 min but then they begin to gradually restore; the oscillation period at the initial stage of the restoration is.shorter than it was earlier and then increases due to the elongation of the contraction phase. Analysis of data obtained is in favor of the assumption that the plasmodial membrane contains MARCKS-like proteins and protein kinase C-controlled pools of phosphatidylinositol-4,5-bisphosphate, which can participate in the generation of auto-oscillations observed in the plasmodium.

Possible involvement of autocrine factors into the control of motile behavior via a receptor-mediated mechanism was investigated in Physarum polycephalum plasmodium, a multinuclear amoeboid cell with the auto-oscillatory mode of motility. Cyclic adenosine monophosphate (cAMP) and extracellular cAMP-specific phosphodiesterase, its involvement into the control of plasmodium motile behavior was proved by action of its strong inhibitor, were regarded as putative autocrine factors. It was shown that the plasmodium secreted cAMP. When it was introduced into agar support, 0,1-1 mM cAMP induced a delay of the plasmodium spreading and its transition to migration. When locally applied, cAMP at the same concentrations induced typical for attractant action the increase in oscillation frequency and the decrease of ectoplasm elasticity. The ability to exhibit positive chemotaxis in cAMP gradient and the dependence of its realization were shown to depend on the plasmodium state. Chemotaxis test specimens obtained from the migrating plasmodium, unlike those obtained from growing culture, generate alternative fronts which compete effectively with fronts oriented towards the attractant increment. The results obtained support our supposition stated earlier that advance of the Physarum polycephalum plasmodium leading edge is determined by local extracellular cAMP gradients arising from a time delay between secretion and hydrolysis of the nucleotide.

In some unicellular eukaryotes, cAMP performs functions not only of the second messenger, but also of hormone, the primary messenger. We have found that cAMP binds to surface receptors of the free-living infusorian Dileptus anser and stimulates activity of the adenylyl cyclase signaling system (AC-system) including heterotrimeric G-proteins and enzyme adenylyl cyclase (AC). The binding of cAMP to receptor is performed with a high affinity (K(D), 27 nM) and is highly specific, as cGMP and adenosine do not produce a marked effect on it. The infusorian cAMP-receptors have been shown to be coupled to G-proteins, which is indicated by a decrease of their affinity to the ligand in the presence of GTP, stimulation of the GTP-binding of G-proteins with the cyclic nucleotide, and block of the cAMP regulatory effects with suramin, an inhibitor of heterotrimeric G-proteins. cAMP stimulates dose-dependently the AC activity, its effect remaining virtually unchanged in the presence of cGMP, AMP, GMP, and adenosine. N6,O2-dibutyryl-cAMP, a non-hydrolyzed cAMP analog, only at comparatively high concentrations competes with cAMP for binding sites and decreases the cAMP stimulating effects on the AC activity and GTP binding. Thus, we have shown for the first time that the AC system of the infusorians D. anser is stimulated with the extracellular cAMP that in this case functions as the external signal regulates activity of extracellular cAMP-dependent effector systems.

Calcium-sensitive forms of adenylyl cyclase (AC) were revealed in most vertebrates and invertebrates and also in some unicellular organisms, in particular ciliates. We have shown for the first time that calcium cations influence the AC activity of ciliate Tetrahymena pyriformis. These cations at the concentrations of 0.2-20 microM stimulated the enzyme activity, and maximum of catalytic effect was observed at 2 microM Ca2+. Calcium cations at a concentrations of 100 microM or higher inhibited the AC activity. Calmodulin antagonists W-5 and W-7 at the concentrations of 20-100 microM inhibited the catalytic effect induced by 5 microM Ca2+ and blocked the effect at higher concentrations of Ca2+. Chloropromazine, another calmodulin antagonist, reduced Ca2+-stimulated AC activity only at the concentrations of 200-1000 microM. AC stimulating effects of serotonin, EGF and cAMP increased in the presence of 5 microM Ca2+. AC stimulating effects of EGF, cAMP and insulin decreased in the presence of 100 microM Ca2+, and AC stimulating effect of cAMP decreased also in the presence of calmodulin antagonists (1 mM). At the same time, stimulating effect of D-glucose in the presence of Ca2+ and calmodulin antagonists did not change essentially. The data obtained speak in favor of the presence of calcium-sensitive forms of AC in ciliate T. pyriformis which mediate enzyme stimulation by EGF, cAMP, insulin, and serotonin.

The content of constitutive from of 70 kDa family heat shock pritein (Hsp70) was determined by the method of immunoblotting. 9 strains of representatives of the genus Acanthamoeba including 8 amphizoic (facultative parasitic) strains and one free-living (isolated from upper horizons of Arctic soils) were studied. We also examined 15 strains of free-living freshwater amoebae of various geographic origin, age and species. 14 of them belonging to the genus Amoeba and one to the genus Trichamoeba. The presence of Hsp70 was demonstrated in the cells of all 25 freshwater amoeba strains, whereas it was shown only for 2 of amphizoic acanthamoebae strains. In all these cases, the position of zone at the blot, revealed by monoclonal anti-HSP70 antibodies, corresponded to polypeptide with molecular mass about 70 kDa. We also found rather high level of constitutive Hsp70 in the cells of contemporary free-living tundra soil representative. However, in this case, the stained zone occupied the position corresponding to MW about 60 kDa which was just the same as earlier obtained for the ancient tundra acanthamoebae strain from permafrost.

The influence of dibromothymoquinone on chlorophyll fluorescence was studied in Chlamydomonas reinhardtii cells using PAM and PEA fluorometers. The reagent affected differently control cells incubated in complete medium and S-starved cells. Thus, the fluorescence yield in the control essentially increased in the presence of dibromothymoquinone, which can be due to the inactivation of light-harvesting complex II protein kinase, followed by the suppression of membrane transition from high-fluorescence state 1 to low-fluorescence state 2. On the contrary, S-starved cells with membranes in state 2 showed a lower fluorescence yield in the presence of dibromothymoquinone than without it. The JIP test of OJIP fluorescence transients suggests that dibromothymoquinone inhibits both light-harvesting complex II kinase and photosynthetic electron transport when added to control, while in starved cells, it acts predominantly as an electron acceptor.

Alterations of Hsp70 level were studied in the cells of freshwater ciliate Tetrahymena pyriformis after medium salinity changes. It is shown that ciliates, acclimated to fresh water (0 per thousand) and to salt water of 2 and 10 per thousand have similar constitutive levels of Hsp70 in their cells. Neither pronounced induction of Hsp70, was not decrease of its level, revealed in ciliates after salinity stresses. These data differ from the results obtained while studying euryhaline ciliate Paramecium nephridiatum and strongly freshwater one Paramecium jenningsi. We presume that the differences in the mode of chaperone system reaction of these ciliates species might be connected with different extents of salinity persistence - the least in P. jenningsi, intermediate in T. pyriformis and the most pronounced in P. nephridiatum.

The review considers the up to date achievements about posttranslational regulation of programmable cell events: growth, development, and apoptosis in unicellular eukaryotes mediated by histone modifications and DNA metylation. Special attention is given to the evolution aspects of the problem.

In universal-code eukaryotes, a single class-1 translation termination factor eRF1 decodes all three stop codons, UAA, UAG, and UGA. In some ciliates with variant genetic codes one or two stop codons are used to encode amino acid(s) and are not recognized by eRF1. In Stylonychia, UAG and UAA codons are reassigned as glutamine codons, and in Euplotes, UGA is reassigned as cysteine codon. In omnipotent eRF1s, stop codon recognition is associated with the N-terminal domain of eRF1. Because variant-code ciliates most likely evolved from universal code ancestor(s), structural features should exist in ciliate eRF1s that restrict their stop codon recognition. To find out amino acid residues which confer UAR-only specificity to Euplotes aediculatus eRF1, eRFI chimeras were constructed by swapping eRF1 E. aediculatus N-terminal domain sequences with the matching ones from the human protein. In these chimeras the MC-domain was from human eRF1. Functional analysis of these chimeric eRFI highlighted the crucial role of the two regions (positions 38-50 and 123-145) in the N-terminal domain of E. aediculatus eRF1 that restrict E. aediculatus eRF1 specificity toward UAR codons. Possibly, restriction of eRF1 specificity to UAR codons might have been an early event occurring in independent instances in ciliate evolutionary history, possibly facilitating the reassignment of UGA to sense codons.

Using electrophoretic mobility shift assay we first revealed in the nuclear extracts of the rodent malaria parasite Plasmodium berghei (P. berghei) proteins, which bind specifically to the double-stranded oligonucleotides reproducing the binding sites of the transcription factors of AP1 family, NF-IL6 and SP1 involved in the up-regulation of human multidrug resistance (mdr1) gene and to the oligonucleotide corresponding to the element responsive for the stimulation by serum (SRE). The nuclear proteins isolated from the P. berghei strains with various chloroquine sensitivity bound differently to the most of the oligonucleotide probes used. Mutations in the consensus sequences of AP1, NF-IL6 and SRE led to the loss of some DNA-protein complexes, suggesting the existence of malaria parasite nuclear proteins, whose DNA-binding domains are similar to DNA-binding domains of NF-IL6, SRF1, and AP1 family members. These proteins exhibit greater activities in chloroquine resistant strains. The results obtained denote profound alterations in the plasmodium regulatory apparatus occurred as the result of selection on chloroquine resistance.

This review summarizes modem data on Golgi apparatus of parasitic protists and demonstrates how the parasitic lifestyle determines functional and structural peculiarities of secretory systems in unrelated groups of unicellular parasites, in comparison to ones of "model systems", mammalian and yeast cells. The review covers the most well-studied protists, predominantly of high medical importance, belonging to following taxons: Parabasalia (Trichomonas), Diplomonada (Giardia), Entamoebidae (Entamoeba), parasitic Alveolata of the phyllum Apicomplexa (Toxoplasma and Plasmodium), and Kinetoplastida (Trypanosoma and Leishmania). Numerous recent publications demonstrated that studies on intracellular traffic in the mentioned above parasites essentially advanced our knowledge of Golgi function, traditionally based on research of cultured mammalian and yeast cells. Morphology of Golgi organelle in eukaryotes from various taxonomic groups has been compared. Within three of total six the highest taxons of Eukatyota (Adl et al., 2005) there exist at minimum eight groups represented by species lacking Golgi dictiosomes. However, biochemical and (or) molecular (genomic) evidences indicate that the organelle with functions of Golgi was present in every studied so far lineage of eukaryotes. Loss of Golgi organelle is a secondary event, which has been proven by identification of Golgi genes in the genomes of Golgi-lacking lineages. This loss might have occurred independently several times in the course of evolution. Neither the number of stacks, nor the size of the organelle correlates with intensity of secretion, or the position of the species on the evolutionary tree (in terms of presumably early/lately diverged lineages).

The review summarizes current data about transduction mechanism of apoptotic signals by caspase-like enzymes and mitochondrial apoptogenic proteins in unicellular eukaryotes. The role of receptor-dependent and receptor-independent caspase cascades is rewieved. The special attention is given to evolution aspects of problem of apoptosis.

Three different phosphatases ("slow", "middle" and "fast") were found in Amoeba proteus (strain B) after PAGE and a subsequent gel staining in 1-naphthyl phosphate containing incubation mixture (pH 9.0). Substrate specificity of these phosphatases was determined in supernatants of homogenates using inhibitors of phosphatase activity. All phosphatases showed a broad substrate specificity. Of 10 tested compounds, p-nitrophenyl phosphate was a preferable substrate for all 3 phosphatases. All phosphatases were able to hydrolyse bis-p-nitrophenyl phosphate and, hence, displayed phosphodiesterase activity. All phosphatases hydrolysed O-phospho-L-tyrosine to a greater or lesser degree. Only little differences in substrate specificity of phosphatases were noticed: 1) "fast" and "middle" phosphatases hydrolysed naphthyl phosphates and O-phospho-L-tyrosine less efficiently than did "slow" phosphatase; 2) "fast" and "middle" phosphatases hydrolysed 2- naphthyl phosphate to a lesser degree than 1-naphthyl phosphate 3) "fast" and "middle" phosphatases hydrolysed O-phospho-L-serine and O-phospho-L-threonine with lower intensity as compared with "slow" phosphatase; 4) as distinct from "middle" and "slow" phosphatases, the "fast" phosphatase hydrolysed glucose-6-phosphate very poorly. The revealed broad substrate specificity of "slow" phosphatase together with data of inhibitory analysis and results of experiments with reactivation of this phosphatase by Zn2+-ions after its inactivation by EDTA strongly suggest that only the "slow" phosphatase is a true alkaline phosphatase (EC 3.1.3.1). The alkaline phosphatase of A. proteus is secreted into culture medium where its activity is low. The enzyme displays both phosphomono- and phosphodiesterase activities, in addition to supposed protein phosphatase activity. It still remains unknown, to which particular phosphatase class the amoeban "middle" and "fast" phosphatases (pH 9.0) may be assigned.

In free-living Amoeba proteus (strain B), 3 phosphatase were found after disc-electrophoresis of 10 microg of protein in PAGE and using 1-naphthyl phosphate as a substrate a pH 9.0. These phosphatases differed in their electrophoretic mobilities - "slow" (1-3 bands), "middle" (one band) and "fast" (one band). In addition to 1-naphthyl phosphate, "slow" phosphatases were able to hydrolyse 2-naphthyl phosphate and p-nitrophenyl phosphate. They were slightly activated by Mg2+, completely inhibited by 3 chelators (EDTA, EGTA and 1,10-phenanthroline), L-cysteine, sodium dodecyl sulfate and Fe2+, Zn2+ and Mn2+ (50 mM), considerably inactivated by orthovanadate, molybdate, phosphatase inhibitor cocktail 1, p-nitrophenyl phosphate, Na2HPO4, DL-dithiothreitol and urea and partly inhibited by H2O2, DL-phenylalanine, 2-mercaptoethanol, phosphatase inhibitor cocktail 2 and Ca2+. Imidazole, L-(+)-tartrate, okadaic acid, NaF and sulfhydryl reagents -p-(hydroxy-mercuri)benzoate and N-ethylmaleimide - had no influence on the activity of "slow" phosphatases. "Middle" and "fast" phosphatases, in contrast to "slow" ones, were not inactivated by 3 chelators. The "middle" phosphatase differed from the "fast" one by smaller resistance to urea, Ca2+, Mn2+, phosphates and H2O2 and greater resistance to dithiothreitol and L-(+)-tartrate. In addition, the "fast" phosphatase was inhibited by L-cysteine but the "middle" one was activated by it. Of 5 tested ions (Mg2+, Cu2+, Mn2+, Ca2+ and Zn2+), only Zn2+ reactivated "slow" phosphatases after their inactivation by EDTA treatment. The reactivation of apoenzyme was only partial (about 35 %). Thus, among phosphatases found in amoebae at pH 9.0, only "slow" ones are Zn-metalloenzymes and may be considered as alkaline phosphatases (EC 3.1.3.1). It still remains uncertain, to which particular phosphatase class "middle" and "fast" phosphatases (pH 9.0) may belong.

Data on parasitophorous vacuole (PV) formation in host cells (HC) harbouring different intracellular protozoan parasites have been reviewed and critically analysed, with special reference to the main representatives of the Coccidia. The vacuole membrane (PVM) is the interface between host and parasite, playing a role in nutrient acquisition by the parasite from the HC. The PV phenomenon is regarded as a generalized HC response to the introduction of alien bodies (microorganisms), which eventually reflects the evolutionary established host-parasite relationships at cellular, subcellular and molecular levels. Special attention has been paid to the existing morpho-functional diversity of the PVs within the same genera and species of parasites, and even at different stages of the parasite life cycle. The PVM is generally considered to derive from the HC plasmalemma, whose biochemical composition undergoes significant changes as the intravacuolar parasite grows. The original HC proteins are selectively excluded from the PVM, while those of the parasite are incorporated. As the result, the changed PVM becomes not fusigenic for HC lysosomes. For Toxoplasma gondii and other cyst-forming coccidia (Isospora, Sarcocystis), a definite correlation has been noticed between the extent of rhoptry and dense granule secrets released by a zoite during HC internalization, on the one hand, and the pattern of the PV that forms, on the other one. In T. gondii, tachyzoites, known to discharge abundant secrets, commonly force the development of PVs limited with a single unit membrane and equipped with a tubulovesicular network in the lumen. Unlike, bradyzoites known to be deficient in secretory materials trigger the formation of PVs with a three-membrane lining composed of the changed invaginated plasmalemma in addition to two membranes of endoplasmic reticulum. The two different types of PV harbour, respectively, exoenteric and enteric stages of T. gondii, the latter being confined to the cat intestine only. Unlike, all endogenous stages of the classic intestinal coccidia (Eimeria spp.) develop within PVs limited with a single membrane, with some invaginations extending into the PV lumen. Unusual PV patterns are characteristic of the extracytoplasmic eimerian coccidia (Cryptosporidium, Epieimeria) and adeleid haemogreagarines (Karyolysus). In cyst-forming coccidia, the PVM is actively involved in tissue cyst wall formation, thus protecting the encysted parasites from recognition by the host immune system. All this strongly suggests that the PV is far from being an indifferent membraneous vesicle containing a parasite, but represents a metabolically active compartment in infected cells. Since all the coccidia are obligate intracellular parasites, the mode of their intimate interaction with the HC, largely accomplished via the PV and its membrane, is vital for their survival as biological species.

An electron microscope study of sulfatized glycosaminoglycans (SG) was made for cyst stages of S. muris. The polysaccharides were detected in the submembranous and subwall layers of the sarcocysts, in addition to the ground substance and septae. Moreover SG were discovered in the cyst stages themselves--metrocytes, intermediate cells and merozoites (gamonts). SG were discernible as electron dark spots in vacuoles of the metrocytes. SG shaped as granules were scattered in the cytoplasm of both intermediate cells and merozoites. More granules of SG were seen in the cytoplasm of the merozoites compared to the intermediate cells. Thus, the quantity, localization and structure of SG are seen to follow the process of differentiation in muscle cysts of S. muris.

Amino acid composition of water-soluble and water-insoluble proteins of 8 strains of Tr. vaginalis is studied. 17 amino acids are found in both protein hydrolyzates. Despite the complete coincidence of their qualitative compositions there are reliable differences in the quantitative contents of some amino acids. Differences in the contents of main amino acids of water-soluble proteins of different strains reflect the belonging of the latter to different sero-groups. No reliable differences in the quantitative contents of amino acids of both water-soluble and water-insoluble proteins in strains belonging to one sero-group are recognised.