Jalal Soltani; Jonathan A. Lal; G. Paul H. van Heusden; Paul J.J. Hooykaas
Abstract
Agrobacterium tumefaciens is capable of gene transfer to both plant and non-plant organisms. Indeed, upon infection of eukaryotic cells, Agrobacterium tumefaciens transfers a piece of its tumor inducing (Ti)-plasmid, called T-DNA, to the host cell nucleus, which subsequently integrates into the host ...
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Agrobacterium tumefaciens is capable of gene transfer to both plant and non-plant organisms. Indeed, upon infection of eukaryotic cells, Agrobacterium tumefaciens transfers a piece of its tumor inducing (Ti)-plasmid, called T-DNA, to the host cell nucleus, which subsequently integrates into the host genome. The VirD2 virulence protein which has relaxase endonuclease activities covalently binds to the 5'end of T-DNA and facilitates its transfer, nuclear localization and integration into the host genome in collaboration with the interacting proteins of the host cell. The VirD2 is essential for Agrobacterium–mediated transformation of both plants and non-plant cells. Here, using yeast Green Flourescent Protein (yGFP) technology, we studied the subcellular localization of VirD2, expressed in the model eukaryote Saccharomyces cerevisiae. Fluorescence microscopy showed that an N-terminal yGFP fusion of VirD2 (i.e. 5' GFP-VirD2 3'), was located in the nucleus of yeast. With C-terminal fusions of VirD2 to yGFP (i.e. 5' VirD2-GFP 3'), no particular subcellular concentration of fluorescence was seen. This further confirms nuclear localization of VirD2 in eukaryotic cells and more importantly highlights the role of Nuclear Localization Signal sequences (NLS) of the C-terminal of VirD2 in this phenomenon.
Roya Lari; Jameel A. Khan; Peter D. Kitchener
Abstract
The exact developmental origin of microglia is still under debate. In the present study we investigated which
heamatopoietic tissues and which features of the organotypic brain slice culture promoted microglia
ramification. The potential of cells derived from embryonic yolk sac, embryonic aorta-gonad-mesonephros ...
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The exact developmental origin of microglia is still under debate. In the present study we investigated which
heamatopoietic tissues and which features of the organotypic brain slice culture promoted microglia
ramification. The potential of cells derived from embryonic yolk sac, embryonic aorta-gonad-mesonephros and
adult blood monocytes was examined. These tissues were co-cultured with brain slices after the brain slices
had first been maintained in vitro for 1 day, 5 days and 9 days. When brain slices had been maintained in
culture for 1 day before the donor cells were added, the donor cells took several days to ramify. However,
when donor tissues were added to brain slices that had been 5 or 9 days maintained in culture, the donor cells
exhibited a ramified morphology within a day. Therefore changes in organotypic brain slices had an effect on
the transformation of cells to the microglial morphology. When adult blood monocytes were added to brain
slice cultures there was no evidence of any tendency to ramify over 6 days of co-culture. This study did not
support the suggestion that microglia cells derive from bone-marrow (BM) cells or from circulating
monocytes.
