Search

Survival and morphogenesis of the enucleated fragments of Acetabularia has been studied after their reconstruction from the cell membrane and the endoplasm. Membranes and endoplasms of different species as well as of the same species but from plants in different functional states and with different regenerative abilities were combined. It was shown that inability of plants for growth and morphogenesis was due to the membrane state. When reconstructing anuclear fragments from membranes and endoplasms of different species or the same species but from plants in different functional states, a phenomenon of membrane-endoplasm incompatibility was found.

The ribosomal RNA transport from a nucleus to a perinuclear cytoplasm and its following distribution in the cytoplasm of Acetabularia mediterranea cells were studied using transplantation of RNA-labeled rhizoid into unlabeled stalk. In addition rifamycin treatment was used for inhibition of cytoplasmic RNA synthesis. Acetabularia nuclei contain the stable RNA fractions similar to those present in some other eukaryotes. Nuclear 25S and 17S ribosomal RNA rapidly enter the rhizoid cytoplasm whereas the following trasfer of them to other regions of the cell is a very slow process. Within two days only an insignificant part of 25S and 17S ribosomal RNA is transferred from the rhizoid to the stalk and is distributed there over the base-apical gradient. No preferential transfer of the nuclear ribosomal RNA to the apical region was observed.

The biosynthesis of nuclear RNA, its transport from the nucleus to the cytoplasm and distribution in the cytoplasm were studied in Acetabularia mediterranea under different light conditions. It was shown that the nuclear RNA incorportate 3H-uracil more rapidly in the darkness and the transport of labeled RNA from the nucleus slowed down after the transfer of plants in the cold medium in the darkness. To study the distribution of nuclear RNA in the cytoplasm, the 3H-uracil labeled nuclei were transplanted in the rhizoids of unlabeled plants, the dikaryons obtained were kept for different time in the light and in the darkness and the content of 3H-RNA was determined in different stem regions. It was shown that the transport of 3H-RNA in the cytoplasm is slowed down in the darkness and it is distributed by the basal-apical gradient. RNA is rapidly accumulated in the apical stem zone in the light and redistributed afterwards in the basal stem zones. The problem of relationship between the polarity and nuclear RNA distribution in Acetabularia is discussed.

The distribution of total RNA and its individual fractions in two regions of Acetabularia mediterranea stem during regeneration was investigated. During regeneration of both the nuclear and enucleated cells, RNA concentration increases in the cytoplasm of growth zone whereas it changes insignificantly in the central stem region. A study of the qualitative RNA composition in the same stem regions has shown that during regeneration high molecular weight RNA fractions (main peaks - 0,56-10(6) and 1.04-10(6) Dalton) are found in the growth zone and are practically absent from the central cell region. Low molecular weight RNA (supposedly, tRNA and products of RNA destruction) are present in both the cell regions under study.

The intensity of RNA synthesis was studied in different regions of anuclear fragments of the Acetabularia mediterranea stem under their local illumination. The local illumination was shown to activate RNA synthesis and formation of distal-medial gradients of this synthesis in the illuminated regions of the fragments to a much greater extent than in their darkened regions. The formation of the gradient in the illuminated region took place even if the growth in this region was insignificant and was not accompanied by the cap formation. The results obtained suggests the absence of obligatory correspondence between the ability of morphogenesis and the gradient of RNA synthesis.

The process of induction of polarity in enucleated fragments (EF) of Acetabularia mediterranea has been studied by the method of local illumination. A device is described which allows to illuminate one part of EF at 2, 500 lx and another at 5 lx. The local illumination of a part of EF during 10-24 hrs was shown to polarize with a high probability the subsequent growth and formation of caps at the subsequent uniform illumination. Using the method of successive illumination and darkening of apical and basal portions of EF, it was shown that the initial stages of induction were partially reversible and the complete determination of polarity took place after the initiation of the cap anlage. After the short-term induction by light (2500 lx), the darkening of the growth zone inhibits completely the process of cap initiation, but already at 100 lx the EF previously induced at 2 500 lx are initiated and grow. While using the local illumination of grafts of EF from two species, A. crenulata and A. mediterranea, with their subsequent fragmentation and the analysis of cap morphology, it was shown that the induction of polar growth did not provide the directed transport of the factors controlling morphogenesis to the future growth zone.

The possibility to synthesize the viral-specific products after microinjection of Tobacco mosaic virus (TMV) preparations and the TMV RNA into the single-celled seaweed Acetabularia was studied. The accumulation of the newly synthesized protein and double-stranded RNA 24 hours after injection of TMV RNA and native virus preparations was demonstrated by immunological and immunofluorescent methods. The virus titer sharply dropped 3--4 hours after introduction into Acetabularia and in 48 hours it reached a maximum level. The presented data showed the possibility of TMW RNA replication and translation involving formation of viral-specific proteins and the production of a virus of full value in the Acetabularia cell.

RNA synthesis and distribution in cytoplasm of Acetabularia mediterranea cell were studied by radiochemical and ultramicrobiochemical methods. Existance of an apico-basal gradient of RNA concentration in the cytoplasm was shown. Variations in the rate of the RNA synthesis in different regions of the cytoplasm are of great importance for the formation of this gradient whereas a contribution of nuclear RNA transport is insignificant. Sedimentation coefficients of about 23S and 16S were detected for the most part of the synthesized cytoplasmic RNAs. Evidently these RNAs represent rRNA of chloroplast ribosomes. Basal region of cytoplasm contains a relatively large amount of low molecular weight RNA. The differences in stability of newly-synthesized RNA were found in cytoplasmic regions under examination.

The species specificity of the factors controlling the cap development was established in the experiments with the transplantation of both the intact and centrifuged in the basal direction apical regions of Acetabularia meditteranea on nuclear basal regions of A. crenulata. These factors are found at the stage of 72 hrs of regeneration primarily in the cytoplasmic zone adjacent to the cell membrane which is not displaced during centrifugation. Using direct measurements and radiochemical method, we have shown that the accumulation of proteins proceeded in this zone due, mainly, to their transition from the cytoplasmic zone displaced during centrifugation.

Using immunochemical and, subsequently, autoradiographic methods, the authors have shown that after the injection of the total fraction of polyribosomes isolated from the rat liver in the Acetabularia cytoplasm a protein interacting with the antiserum against rat thyrosine aminotransferase is synthesized. Prospects of using Acetabularia as a test-system for the translation of heterogenous mRNA and mRNP are discussed.