PEG Branched Polymer for Functionalization of Nanomaterials with Ultralong Blood Circulation

Nanomaterials have been actively pursued for biological and medical applications in recent years. Here, we report the synthesis of several new poly(ethylene glycol) grafted branched-polymers for functionalization of various nanomaterials including carbon nanotubes, gold nanoparticles (NP) and gold nanorods (NRs), affording high aqueous solubility and stability for these materials. We synthesize different surfactant polymers based upon poly-(g-glutamic acid) (gPGA) and poly(maleic anhydride-alt-1-octadecene) (PMHC18). We use the abundant free carboxylic acid groups of gPGA for attaching lipophilic species such as pyrene or phospholipid, which bind to nanomaterials via robust physisorption. Additionally, the remaining carboxylic acids on gPGA or the amine-reactive anhydrides of PMHC18 are then PEGylated, providing extended hydrophilic groups, affording polymeric amphiphiles. We show that single-walled carbon nanotubes (SWNTs), Au NPs and NRs functionalized by the polymers exhibit high stability in aqueous solutions at different pHs, at elevated temperatures and in serum. Morever, the polymer-coated SWNTs exhibit remarkably long blood circulation (t1/2 22.1 h) upon intravenous injection into mice, far exceeding the previous record of 5.4 h. The ultra-long blood circulation time suggests greatly delayed clearance of nanomaterials by the reticuloendothelial system (RES) of mice, a highly desired property for in vivo applications of nanomaterials, including imaging and drug delivery

Carbon States in Carbon-Encapsulated Nickel Nanoparticles Studied by Means of X-Ray Absorption, Emission, and Photoelectron Spectroscopies

Electronic structure of nickel nanoparticles encapsulated in carbon was characterized by photoelectron, X-ray absorption, and X-ray emission spectroscopies. Experimental spectra are compared with the density of states calculated in the frame of the density functional theory. The carbon shell of Ni nanoparticles has been found to be multilayer graphene with significant (about 6%) amount of Stone--Wales defects. Results of the experiments evidence protection of the metallic nanoparticles from the environmental degradation by providing a barrier against oxidation at least for two years. Exposure in air for 2 years leads to oxidation only of the carbon shell of Ni@C nanoparticles with coverage of functional groups.Comment: 16 pages, 6 figures, accepted in J. Phys. Chem.

Nanotechnology in Head and Neck Cancer: The Race Is On

Rapid advances in the ability to produce nanoparticles of uniform size, shape, and composition have started a revolution in the sciences. Nano-sized structures herald innovative technology with a wide range of potential therapeutic and diagnostic applications. More than 1000 nanostructures have been reported, many with potential medical applications, such as metallic-, dielectric-, magnetic-, liposomal-, and carbon-based structures. Of these, noble metallic nanoparticles are generating significant interest because of their multifunctional capacity for novel methods of laboratory-based diagnostics, in vivo clinical diagnostic imaging, and therapeutic treatments. This review focuses on recent advances in the applications of nanotechnology in head and neck cancer, with special emphasis on the particularly promising plasmonic gold nanotechnology

Replication fork stalling by bulky DNA damage: localization at active origins and checkpoint modulation

The integrity of the genome is threatened by DNA damage that blocks the progression of replication forks. Little is known about the genomic locations of replication fork stalling, and its determinants and consequences in vivo. Here we show that bulky DNA damaging agents induce localized fork stalling at yeast replication origins, and that localized stalling is dependent on proximal origin activity and is modulated by the intra–S–phase checkpoint. Fork stalling preceded the formation of sister chromatid junctions required for bypassing DNA damage. Despite DNA adduct formation, localized fork stalling was abrogated at an origin inactivated by a point mutation and prominent stalling was not detected at naturally-inactive origins in the replicon. The intra–S–phase checkpoint contributed to the high-level of fork stalling at early origins, while checkpoint inactivation led to initiation, localized stalling and chromatid joining at a late origin. Our results indicate that replication forks initially encountering a bulky DNA adduct exhibit a dual nature of stalling: a checkpoint-independent arrest that triggers sister chromatid junction formation, as well as a checkpoint-enhanced arrest at early origins that accompanies the repression of late origin firing. We propose that the initial checkpoint-enhanced arrest reflects events that facilitate fork resolution at subsequent lesions

In Vitro Structural and Functional Evaluation of Gold Nanoparticles Conjugated Antibiotics

Bactericidal efficacy of gold nanoparticles conjugated with ampicillin, streptomycin and kanamycin were evaluated. Gold nanoparticles (Gnps) were conjugated with the antibiotics during the synthesis of nanoparticles utilizing the combined reducing property of antibiotics and sodium borohydride. The conjugation of nanoparticles was confirmed by dynamic light scattering (DLS) and electron microscopic (EM) studies. Such Gnps conjugated antibiotics showed greater bactericidal activity in standard agar well diffusion assay. The minimal inhibitory concentration (MIC) values of all the three antibiotics along with their Gnps conjugated forms were determined in three bacterial strains,Escherichia coli DH5α,Micrococcus luteusandStaphylococcus aureus. Among them, streptomycin and kanamycin showed significant reduction in MIC values in their Gnps conjugated form whereas; Gnps conjugated ampicillin showed slight decrement in the MIC value compared to its free form. On the other hand, all of them showed more heat stability in their Gnps conjugated forms. Thus, our findings indicated that Gnps conjugated antibiotics are more efficient and might have significant therapeutic implications

Shelterin-Like Proteins and Yku Inhibit Nucleolytic Processing of Saccharomyces cerevisiae Telomeres

Eukaryotic cells distinguish their chromosome ends from accidental DNA double-strand breaks (DSBs) by packaging them into protective structures called telomeres that prevent DNA repair/recombination activities. Here we investigate the role of key telomeric proteins in protecting budding yeast telomeres from degradation. We show that the Saccharomyces cerevisiae shelterin-like proteins Rif1, Rif2, and Rap1 inhibit nucleolytic processing at both de novo and native telomeres during G1 and G2 cell cycle phases, with Rif2 and Rap1 showing the strongest effects. Also Yku prevents telomere resection in G1, independently of its role in non-homologous end joining. Yku and the shelterin-like proteins have additive effects in inhibiting DNA degradation at G1 de novo telomeres, where Yku plays the major role in preventing initiation, whereas Rif1, Rif2, and Rap1 act primarily by limiting extensive resection. In fact, exonucleolytic degradation of a de novo telomere is more efficient in yku70Δ than in rif2Δ G1 cells, but generation of ssDNA in Yku-lacking cells is limited to DNA regions close to the telomere tip. This limited processing is due to the inhibitory action of Rap1, Rif1, and Rif2, as their inactivation allows extensive telomere resection not only in wild-type but also in yku70Δ G1 cells. Finally, Rap1 and Rif2 prevent telomere degradation by inhibiting MRX access to telomeres, which are also protected from the Exo1 nuclease by Yku. Thus, chromosome end degradation is controlled by telomeric proteins that specifically inhibit the action of different nucleases

Checkpoint-Dependent and -Independent Roles of Swi3 in Replication Fork Recovery and Sister Chromatid Cohesion in Fission Yeast

Multiple genome maintenance processes are coordinated at the replication fork to preserve genomic integrity. How eukaryotic cells accomplish such a coordination is unknown. Swi1 and Swi3 form the replication fork protection complex and are involved in various processes including stabilization of replication forks, activation of the Cds1 checkpoint kinase and establishment of sister chromatid cohesion in fission yeast. However, the mechanisms by which the Swi1–Swi3 complex achieves and coordinates these tasks are not well understood. Here, we describe the identification of separation-of-function mutants of Swi3, aimed at dissecting the molecular pathways that require Swi1–Swi3. Unlike swi3 deletion mutants, the separation-of-function mutants were not sensitive to agents that stall replication forks. However, they were highly sensitive to camptothecin that induces replication fork breakage. In addition, these mutants were defective in replication fork regeneration and sister chromatid cohesion. Interestingly, unlike swi3-deleted cell, the separation-of-functions mutants were proficient in the activation of the replication checkpoint, but their fork regeneration defects were more severe than those of checkpoint mutants including cds1Δ, chk1Δ and rad3Δ. These results suggest that, while Swi3 mediates full activation of the replication checkpoint in response to stalled replication forks, Swi3 activates a checkpoint-independent pathway to facilitate recovery of collapsed replication forks and the establishment of sister chromatid cohesion. Thus, our separation-of-function alleles provide new insight into understanding the multiple roles of Swi1-Swi3 in fork protection during DNA replication, and into understanding how replication forks are maintained in response to different genotoxic agents

Histone methyltransferase Dot1 and Rad9 inhibit single-stranded DNA accumulation at DSBs and uncapped telomeres

Cells respond to DNA double-strand breaks (DSBs) and uncapped telomeres by recruiting checkpoint and repair factors to the site of lesions. Single-stranded DNA (ssDNA) is an important intermediate in the repair of DSBs and is produced also at uncapped telomeres. Here, we provide evidence that binding of the checkpoint protein Rad9, through its Tudor domain, to methylated histone H3-K79 inhibits resection at DSBs and uncapped telomeres. Loss of DOT1 or mutations in RAD9 influence a Rad50-dependent nuclease, leading to more rapid accumulation of ssDNA, and faster activation of the critical checkpoint kinase, Mec1. Moreover, deletion of RAD9 or DOT1 partially bypasses the requirement for CDK1 in DSB resection. Interestingly, Dot1 contributes to checkpoint activation in response to low levels of telomere uncapping but is not essential with high levels of uncapping. We suggest that both Rad9 and histone H3 methylation allow transmission of the damage signal to checkpoint kinases, and keep resection of damaged DNA under control influencing, both positively and negatively, checkpoint cascades and contributing to a tightly controlled response to DNA damage