Subnucleolar location of fibrillarin and NopA64 in Lepidium sativum root meristematic cells is changed in altered gravity.

Fibrillarin and the plant nucleolin homolog NopA64 are two important nucleolar proteins involved in pre-rRNA processing. In order to determine the effects of the altered gravity environment on the nucleolus, we have investigated the location of fibrillarin and NopA64 in nucleolar subcomponents of cress (Lepidium sativum L.) root meristematic cells grown under clinorotation, which reproduces an important feature of microgravity, namely, the absence of the orienting action of a gravity vector, and compared it to the location in control cells grown in normal 1 g conditions. Prior to these experiments, we report here the characterization of cress fibrillarin as a 41 kDa protein which can be isolated from meristematic cells in three nuclear fractions, namely, the soluble ribonucleoprotein fraction, the chromatin fraction, and the nuclear-matrix fraction. Furthermore, as reported for other species, the location of both fibrillarin and NopA64 in the cress cell nucleolus was in zones known to contain complex ribonucleoprotein particles involved in early pre-rRNA processing, i.e., processomes. Under altered gravity, a decrease in the quantity of both fibrillarin and NopA64 compared to controls was observed in the transition zone between fibrillar centers and the dense fibrillar component, as well as in the bulk of the dense fibrillar component. These data suggest that altered (reduced) gravity results in a lowered level of functional activity in the nucleolus.

[Alterations of nucleolar DNA localization caused by simulated microgravity].

The functions of the nucleolus responsible for the biosynthesis of ribosomes in altered gravity are still unclear. The location of nucleolar DNA both in control and simulated microgravity has been investigated using fluorescent and immunogold cytochemistry. We have determined the redistribution of rDNA in the nucleolar components accompanied by their ultrastructure changes. According to this the lowering of rDNA transcription level under simulated microgravity was supposed.

[Altered gravity affects subnucleolus localization of fibrillarin and NopA64, the most important proteins of rRNA processing].

Fibrillarin and plant nucleolin homologue NopA64 are two important nucleolar proteins involved in pre-rRNA processing. To understand better the effects of the altered gravity environment on the nucleolus functioning we have investigated the location of fibrillarin and NopA64 in nucleolar subcomponents of cress (Lepidium sativum L.) root meristematic cells grown under simulated microgravity that was compared to the control cells grown in normal conditions at I g. Cress fibrillarin was first shown to have the molecular weight 41 kDa. Both fibrillarin and NopA64 in the cress cell nucleolus are located in the zones known to contain processing pre-rRNA molecules as it has been previously reported in other species. The data confirm participation of these proteins in processomes--RNP complex particles involved in pre-rRNA processing. Under altered gravity a decrease in the quantity of both fibrillarin and NopA64 in the transition zone between fibrillar centres and the dense fibrillar component was observed, compared to control, which could point out to a lowering of the level of early pre-rRNA processing in these experimental conditions. This decrease was also detected in the bulk of the dense fibrillar component. These data support the idea that altered (reduced) gravity results in lowering the level of functional activity of the nucleolus.

Alterations of the intranucleolar DNA localization caused by simulated microgravity.

The functions of the nucleolus responsible for the biosynthesis of ribosomes in altered gravity are still unclear. The location of nucleolar DNA both in control and simulated microgravity with fluorescent and immunogold cytochemistry was investigated. We determined firstly the redistribution of rDNA in the nucleolar components accompanied with the changes in their ultrastructure. According to it, lowering the level of rDNA transcription in simulated microgravity was firstly proposed.

Nucleolar structure and function under clinorotation.

A nucleolus is the dynamic part of a cell nucleus where rDNA transcription, rRNA precursor processing and transport are proceeding. Investigations of the influence of microgravity and clinorotation on the structure and function of nucleolus began more than thirty years ago but, unfortunately, now there are only some articles concerning the nucleolar peculiarities in altered gravity. Available data about alterations of nucleolar morphology and function under the influence of high and low temperature, hypoxia and UV-microbeam irradiation allow us to put intent attention to the study of the nucleolus reactions on clinorotation. Therefore, the aim of our work was to investigate the ultrastructural organization and functional activity of nucleoli in Lepidium sativum root meristematic cells under clinorotation in order to understand whether a nucleolus can react to gravity.