Emerging areas of nanotechnology hold the promise of overcoming the limitations of existing technologies for intracellular manipulation. These new developments provide approaches for the creation of chemical-biological hybrid nanocomposites that can be introduced into cells and subsequently used to initiate intracellular processes or biochemical reactions. Such nanocomposites would advance medical biotechnology, just as they are improving microarray technology and imaging in biology and medicine, and introducing new possibilities in chemistry and material sciences. Here we describe the behaviour of 45-A nanoparticles of titanium dioxide semiconductor combined with oligonucleotide DNA into nanocomposites in vivo and in vitro. These nanocomposites not only retain the intrinsic photocatalytic capacity of TiO2 and the bioactivity of the oligonucleotide DNA (covalently attached to the TiO2 nanoparticle), but also possess the chemically and biologically unique new property of a light-inducible nucleic acid endonuclease, which could become a new tool for gene therapy.

Proliferating cell nuclear antigen (PCNA) protein is one of the central molecules responsible for decisions of life and death of the cell. The PCNA gene is induced by p53, while PCNA protein interacts with p53-controlled proteins Gadd45, MyD118, CR6 and, most importantly, p21, in the process of deciding cell fate. If PCNA protein is present in abundance in the cell in the absence of p53, DNA replication occurs. On the other hand, if PCNA protein levels are high in the cell in the presence of p53, DNA repair takes place. If PCNA is rendered non-functional or is absent or present in low quantities in the cell, apoptosis occurs. The evolution from prokaryotes to eukaryotes involved a change of function of PCNA from a 'simple' sliding clamp protein of the DNA polymerase complex to an executive molecule controlling critical cellular decision pathways. The evolution of multicellular organisms led to the development of multicellular processes such as differentiation, senescence and apoptosis. PCNA, already an essential molecule in the life of single cellular organisms, then became a protein critical for the survival of multicellular organisms.

While few publications have documented the uptake of nanoparticles in plants, this is the first study describing uptake and distribution of the ultra-small anatase TiO 2 in the plant model system Arabidopsis. We modified the nanoparticle surface with Alizarin red S and sucrose, and demonstrated that nanoconjugates traversed cell walls, entered into plant cells, and accumulated in specific subcellular locations. Optical and X-ray fluorescence microscopy co-registered the nanoconjugates in cell vacuoles and nuclei. KeywordsAnatase TiO 2 nanoparticles; TiO 2 nanoconjugates; Arabidopsis thaliana; X-ray fluorescence microscopy (XFM)The application of nanotechnology to plant systems has lagged behind nanomedicine and nanopharmacology in spite of its potential to generate new tools for the delivery of fertilizers, herbicides and insecticides 1 , new ways to manipulate plant genomes 2 and new methods to capture and isolate plant natural products. Compared to the thousands of studies describing the uptake and trafficking of nanoparticles (NPs) in biological systems other than plants, less than twenty reports discussed NP uptake by plant species. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16] These studies involved different plant species and different types of NPs which were delivered to intact plants, dissected plant organs or protoplasts using a wide range of application methods. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16] Despite the absence of systematic analyses, it has been determined that plants can take up NPs from the environment and transport them through the vascular system to various shoot organs. 4,9,15 However, little is known about the uptake mechanisms involved or the subcellular localization and distribution of the internalized NPs. 16 Uptake efficiency has also Figure S1), and show additional data describing NC distribution and localization in roots, hypotocyls and cotyledons (Figures S2, S3 and S4). This material is available free of charge via the Internet at http://pubs.acs.org. Here, we report on the uptake and localization of anatase titanium dioxide (TiO 2 ) NPs smaller than 5 nm in the plant model system Arabidopsis thaliana. We chose to study the Col-0 accession because this is the most commonly used ecotype within the Arabidopsis research community. 22 The numerous resources developed for this genetic background 23,24 will not only facilitate future analyses of the molecular mechanisms of uptake, intracellular localization and trafficking of NPs, but will also provide opportunities for NP-mediated manipulations of the Arabidopsis genome. In addition, the well-characterized Arabidopsis null mutants and overexpression lines for enzymes of various biochemical pathways offer the possibility for the targeted in planta chemical modification of NP surface with pathway intermediates. TiO 2 NPs with average diameters of 2.8 ± 1.4 nm and NP dispersity of 43% (see Supporting information) were synthesized by a low-temperature alkaline hydrolysis route as described previously 2...

Characteristic X-ray fluorescence is a technique that can be used to establish elemental concentrations for a large number of different chemical elements simultaneously in different locations in cell and tissue samples. Exposing the samples to an X-ray beam is the basis of X-ray fluorescence microscopy (XFM). This technique provides the excellent trace element sensitivity; and, due to the large penetration depth of hard X-rays, an opportunity to image whole cells and quantify elements on a per cell basis. Moreover, because specimens prepared for XFM do not require sectioning, they can be investigated close to their natural, hydrated state with cryogenic approaches. Until several years ago, XFM was not widely available to bio-medical communities, and rarely offered resolution better then several microns. This has changed drastically with the development of third-generation synchrotrons. Recent examples of elemental imaging of cells and tissues show the maturation of XFM imaging technique into an elegant and informative way to gain insight into cellular processes. Future developments of XFM-building of new XFM facilities with higher resolution, higher sensitivity or higher throughput will further advance studies of native elemental makeup of cells and provide the biological community including the budding area of bionanotechnology with a tool perfectly suited to monitor the distribution of metals including nanovectors and measure the results of interactions between the nanovectors and living cells and tissues. Key words: X-ray fluorescence microscopy; elemental maps; metalome; bionanotechnology The biology of the past decade has significantly changed scope, and large compendia of data have been accumulated to enable scientists to study biological ''meta units'' such as the genome, proteome, and transcriptome. At this time, the term ''metalome'' is still used only rarely, and the majority of scientists interested in establishing metalome databases undertook the daunting task of developing and using different dyes for different elements and their ions in order to determine elemental locations and concentrations in cells and tissues. The presence of metals and trace elements is essential for the existence of life as we know it. In any organism, there are very few intracellular processes that are not dependent on the presence of metals or other trace elements. In fact, it is estimated that one-third of all known proteins contain metal cofactors and the majority of these function as essential metalloenzymes. With current developments in genomics and proteomics, our knowledge of the enormous number of pathways in which metals and trace elements are necessary for life is ever increasing. This knowledge, however, is largely ''static'' as we still do not have appropriate sensitive approaches to follow fluctuations in normal metal homeostasis that accompany processes of development, differentiation, senescence, stress responses, etc. Likewise, our knowledge about the redistribution of metals and trace elements accom...