Ubiquitin and ubiquitin-conjugated proteins in the olive (olea europaea l.) pollen

A study of free ubiquitin and ubiquitinated proteins has been carried out in olive (Olea europaea L.) during the late stages of pollen development and during in-vitro pollen hydration. As described in relatively few plant species, immunoblot experiments carried out using crude protein extracts have shown a reduction ill the free ubiquitin level taking place during the mature pollen stage. This reduction does not coincide with a pal allel reduction in the level of the corresponding transcripts, as detected by RT-PCR analysis, suggesting the existence of a post-transcriptional regulatory mechanism. Relatively high levels of free ubiquitin are quickly restored upon in-vitro pollen hydration. TEM immunocytochemical localisation of ubiquitin-conjugated compounds in the mature pollen grain showed the antibodies bound to the cytoplasm, the chromatin and the nucleolus of the vegetative and generative cells. Within the cytoplasm, labelling was concentrated in the cytosol, the endoplasmic reticulum membranes and minuscule vesicles associated to the Golgi apparatus. The fibrillar material adhering to the exine, and probably derived from the tapetum, also showed Sold particles. Lipid bodies, aperture regions, mitochondria, the intine and the cell wall were devoid of labelling. The changes observed throughout pollen development, as well as the specific localisation, reflect the involvement of the ubiquitin-mediated degradative pathway in regulation of pollen metabolism.