Influence of metal-contamination on distribution in subcellular fractions of the earthworm (Metaphire californica) from Hunan Province, China

Earthworms have the ability to accumulate of heavy metals, however, there was few studies that addressed the metals in earthworm at subcellular levels in fields. The distributions of metals （Cd, Cu, Zn, and Pb） in subcellular fractions （cytosol, debris, and granules） of earthworm Metaphire californica were investigated. The relationship between soil metals and earthworms were analyzed to explain its high plasticity to inhabit in situ contaminated soil of Hunan Province, south China. The concentration of Cd in subcellular compartments showed the same pattern as Cu in the order of cytosol 〉 debris 〉 granules. The distribution of Zn and Pb in earthworms indicated a similar propensity for different subcellular fractions that ranked as granules 〉 debris 〉 cytosol for Zn, and granules 〉 cytosol 〉 debris for Pb. The internal metal concentrations in earthworms increased with the soil metals （p 〈 0.05）. Significant positive correlations were found between soil Cd and Cd concentrations in cytosol and debris （p 〈 0.01）. Moreover, the soil Pb concentration significantly influenced the Pb concentrations in cytosol and debris （p 〈 0.01）, similar to that of Cd. The soil Cu concentrations was only associated with the Cu in granules （p 〈 0.05）. Soil Zn concentrations correlated with the Zn concentrations in each subcellular fraction （p 〈 0.05）. Our results provide insights into the variations of metals partitioning in earthworms at subcellular levels and the relationships of soil metals, which could be one of the detoxiflcation strategies to adapt the long-term contaminated environment.

Temporal and spatial profiling of nuclei-associated proteins upon TNF-α/NF-kB signaling

The tumor necrosis factor （TNF）-α/NF-kB-signaling pathway plays a pivotal role in various processes including apoptosis, cellular differentiation, host defense, inflammation, autoimmunity and organogenesis. The complexity of the TNF-α/NF-kB signaling is in part due to the dynamic protein behaviors of key players in this pathway. In this present work, a dynamic and global view of the signaling components in the nucleus at the early stages of TNF-a/ NF-KB signaling was obtained in HEK293 cells, by a combination of subcellular fractionation and stable isotope la- beling by amino acids in cell culture （SILAC）. The dynamic profile patterns of 547 TNF-α-induced nuclei-associated proteins were quantified in our studies. The functional characters of all the profiles were further analyzed using that Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway annotation. Additionally, many previously unknown effectors of TNF-α/NF-kB signaling were identified, quantified and clustered into differential activation profiles. In- terestingly, levels of Fanconi anemia group D2 protein （FANCD2）, one of the Fanconi anemia family proteins, was found to be increased in the nucleus by SILAC quantitation upon TNF-α stimulation, which was further verified by western blotting and immunofluorescence analysis. This indicates that FANCD2 might be involved in TNF-α/NF-kB signaling through its accumulation in the nucleus. In summary, the combination of subcellular proteomics with quan- titative analysis not only allowed for a dissection of the nuclear TNF-α/NF-kB-signaling pathway, but also provided a systematic strategy for monitoring temporal and spatial changes in cell signaling.