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Heat shock proteins have been revealed for the first time by the methods of Western blotting using alkaline phosphatase and ECL in the cells of Porphyra purpurea from Kattegat area of the Baltic Sea in normal and experimental stress conditions. It was demonstrated with application of monoclonal anti-Hsp70 antibodies that a slight band about 70 kDa is present constitutively at the film; additionally the polypeptide of about 40 kDa ("Hsp40") has been detected. After heat shock at 28 degrees C during 1 hr significant "expenditure" of Hsp70 was observed, as well as the pronounced induction of "Hsp40"; the induction was expressed especially strongly in 24 hr after the stress application.

The composition and the level of heat shock (stress) proteins content were studied by the method of immunobloting in the gill epithelium cells of mussels Mytilus edulis L. from the White sea, treated by water of different salinities. Stress proteins of about 70 and 40 kDa were revealed at Western blots. After long-term (11-14 days) acclimation to 14 and 35 per thousand the level of Hsp70 in gill epithelium cells increased to compare with that in control mussels. Hsp70 induction was observed in the cells of isolated gills after salinity shock at 14 per thousand during 3 and 24 hours.

Our own studies of alterations in the level of constitutive heat shock protein of 70 kDa family (Hsp70) in freshwater (Paramecium jenningsi), meta-freshwater (Tetrahymena pyriformis) and curyhaline (P. nephridiatum) ciliates acclimated to salt-water and fresh-water medium were reviewed. It has been shown that the level of constitutive Hsp70 content correlates with the salinity-resistance of ciliate species: in P. jenning i it was lower in freshwater, than in marine water, in euryhaline P. nephridiatum it was higher in freshwater, than in marine water, and was more or less stable in T. pyriformis, the ciliate that occupies intermediate position between two mentioned above species according its salinity-resistance. The ecological importance of constitutive heat shock protein level is discussed in the context of salinity adaptations studies.

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.

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.

Expression of HSP70 is induced by stress factors, including sub-lethal chilling, but the role of HSP70 for overcoming consequenses of cold stress is not clear. If the role is positive, the level of HSP70 expression might be higher in populations from colder climates. By use of immunoblotting we investigated dynamics of HSP70 expression in response to cold stress in two Myrmica species from three localities of different latitude (50, 60 and 67 degrees N). The results showed that in the more thermophilic species Myrmica rubra expression of HSP70 after cold stress was more intense. Expression traits within both species did not show direct relationship with latitude. In M. rubra the southernmost population and in M. ruginodis the northernmost one displayed the fastest and most intense response. However, in M. rubra the two other studied populations were similar in timing and intensity of the response, while in M. ruginodis the intermediate population showed the slowest and weakest response. The data suggest that the expression of HSP70 might play the most considerable role in the adaptation to cold in the southernmost population of M. ruginodis.

The level of Hsp70 was studied in the cells of eurihaline ciliate Paramecium nephridiatum after the environmental salinity changes. Two types of treatment were applied. "Shock": ciliates were placed for 1 h to the medium with stress salinity, then transferred back to the medium, they were acclimated to, for 2 h; "adaptation": ciliates were placed for 3 h into stress salinity. It has been shown, that ciliates, acclimated to fresh water (0%) have the higher constitutive level of Hsp70, than those, acclimated to 10%. Transfer from fresh water to 10% does not cause the increase of Hsp70 synthesis in protists, whereas the reciprocal transfer results in induction of Hsp70 in the cells. "Adaptation" results in induction of Hsp70 in both "directions" of salinity changes. The results obtained allow to presume that the possibility to survive in the media of various salinity in eurihaline ciliates is somehow determined by the higher initial level of Hsp70 in their cells, than in stenohaline representatives of the same genus.

Changes in the level of heat shock proteins (HSP) in cells of freshwater protists, amoebae Amoeba proteus and ciliates Paramecium jenningsi, in response to changes in the environmental salinity were investigated. Changes in salinity levels were considered as a stress factor. The immunoblotting method revealed a polypeptide antigen cross-reacting with antibodies against bovine HSP70 in total protein extracts of both intact cells and cells subjected to salinity stress. The same polypeptide antigen was revealed in A. proteus cells subjected to heat shock. Therefore, it may be supposed that the polypeptide revealed after salinity shock is a heat shock protein related to the vertebrate HSP70. Under the impact of stress factor, well acclimated protists mostly spend their own previously accumulated HSP70. A conclusion is made that freshwater protists, living under conditions of increased salinity, appear to be preadapted to changes in environmental factors.

A heat shock protein of HSP70 family was revealed for the first time in trophozoites of Acanthamoeba sp. (strain Am8) excysted from cysts previously isolated from samples of permafrost aging 30,000-35,000 years. The constitutive level of this HSP, shown by immunnoblotting in unstressed trophozoites of the ancient acanthamoebae, much surpassed that in unstressed cells of the three examined species of contemporary freshwater amebae of the genus Amoeba.

The unique structural and functional organization of macronuclear (somatic nucleus) genome of the spirotrichous ciliates, exemplified by Stylonychia lemnae, has been reviewed. Data on the architecture of S. lemnae nuclear apparatus at interphase and during vegetative cell division, conjugation or autogamy are summarized. Special attention being paid to the structural and functional peculiarities of short macronuclear minichromosomes known to contain protein-coding regions, 5'- and 3'-flanking nontranslated regions, and telomeres. A hypothesis, previously put forward, according to which in the spirotrichous ciliates the telomeres themselves may serve as starting points of replication in minichromosomes, has now received its further substantiation. The recent experimental data, which confirm that 5'-nontranscribed DNA leader sequence of alpha1- and alpha2-tubulin-encoding minichromosomes display at least several regulatory elements typical for eukaryote promoter (TATA-box, CAAT-box, transcriptional initiator), are discussed. Up to now, there is no confirmation with regard to a possible existence in the spirotrichous minichromosomes of specific regulatory sequences capable of controlling both replication and transcription processes.

Glucose-6-phosphate and 6-phosphogluconate dehydrogenases (G6PD and 6PGD) are revealed in Amoeba proteus by electrophoresis in polyacrylamide gels, thus proving the availability of the phosphogluconic pathway in amoebae. 6PGD is marked as a single band, and G6PD shows multiple banding. When an amoebic homogenate is obtained using Triton-100, a supplementary form of G6PD extracted from membranes of some cell organelles (presumably mitochondria) becomes apparent. Hexose-6-phosphate dehydrogenase seems to be absent and therefore all the G6PD forms found may be specific G6PDs proper.