Multiple myeloma (MM) is an incurable plasma cell malignancy with a terminal phase marked by increased proliferation and resistance to therapy. Arsenic trioxide (ATO), an antitumor agent with a multifaceted mechanism of action, displayed clinical activity in patients with late-stage multiple myeloma. However, the precise mechanism(s) of action of ATO has not been completely elucidated. In the present study, we used proteomics to analyze the ATO-induced protein alterations in MM cell line U266 and then investigated the molecular pathways responsible for the anticancer actions of ATO. Several clusters of proteins altered in expression in U266 cells upon ATO treatment were identified, including down-regulated signal transduction proteins and ubiquitin/proteasome members, and up-regulated immunity and defense proteins. Significantly regulated 14-3-3zeta and heat shock proteins (HSPs) were selected for further functional studies. Overexpression of 14-3-3zeta in MM cells attenuated ATO-induced cell death, whereas RNAi-based 14-3-3zeta knock-down or the inhibition of HSP90 enhanced tumor cell sensitivity to the ATO induction. These observations implicate 14-3-3zeta and HSP90 as potential molecular targets for drug intervention of multiple myeloma and thus improve our understanding on the mechanisms of antitumor activity of ATO.

The aim of this study was to identify novel serological tumor markers for human prostate cancer (PCa). We compared the gene expression profile of PCa tissues to adjacent benign tissues of prostate using gene expression microarray. 1207 genes that were consistently different from adjacent benign tissues of prostate (paired t test, P<0.05) were selected as differentially expressed genes (DEGs). Among them, 652 DEGs were upregulated in PCa, whereas 555 DEGs were downregulated in PCa. In addition, two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) coupled with MS was performed to screen for candidate markers in the proteome of PCa and adjacent benign tissues of prostate. A total of 89 spots were significantly up-regulated (ratio≥2, P<0.01) in PCa samples, whereas 66 spots were down-regulated (ratio≤-2, P<0.01). Sixty gene products were identified among these spots. Moreover, 14 potential candidate markers, which were identified as differentially expressed molecules by both gene expression microarray and 2D-DIGE, were chosen for validation and analysis by ELISA. The serum levels of three proteins correlated well with the 2D-DIGE results. Furthermore, the increased serum level of Inosine monophosphate dehydrogenase II (IMPDH2) was significantly associated with the clinicopathological features of the patients with PCa, suggesting its potential as a serological tumor marker. These results demonstrated that integrative transcriptome and proteome analysis could be a powerful tool for marker discovery in PCa. We suggest IMPDH2 as a novel serological tumor marker for detection of early PCa and evaluation of tumor progression.

Hepatitis B virus (HBV) X protein (HBx), a trans-regulator, is frequently expressed in truncated form without carboxyl-terminus in hepatocellular carcinoma (HCC), but its functional mechanisms are not fully defined. In this report, we investigated frequency of this natural HBx mutant in HCCs and its functional significance. In 102 HBV-infected patients with HCC, C-terminal truncation of HBx, in contrast to full-length HBx, were more prevalent in tumors (70.6%) rather than adjacent non-tumorous tissues (29.4%) (p = 0.0032). Furthermore, two naturally-occurring HBx variants (HBxΔ31), which have 31 amino acids (aa) deleted (codons 123-125/124-126) at C-terminus were identified in tumors and found that the presence of HBxΔ31 significantly correlated with intrahepatic metastasis. We also show that over-expression of HBxΔ31 enhanced hepatoma cell invasion in vitro and metastasis in vivo compared to full-length HBx. Interestingly, HBxΔ31 exerts this function via down-regulating Maspin, RhoGDIα and CAPZB, a set of putative metastasis-suppressors in HCC, in part, by enhancing the binding of transcriptional repressor, myc-associated zinc finger protein (MAZ) to the promoters through physical association with MAZ. Notably, these HBxΔ31-repressed proteins were also significantly lower expression in a subset of HCC tissues with C-terminal HBx truncation than the adjacent non-tumorous tissues, highlighting the clinical significance of this novel HBxΔ31-driven metastatic molecular cascade. Our data suggest that C-terminal truncation of HBx, particularly breakpoints at 124aa, plays a role in enhancing hepatoma cell invasion and metastasis by deregulating a set of metastasis-suppressors partially through MAZ, thus uncovering a novel mechanism for the progression of HBV-associated hepatocarcinogenesis.

The morbidity of NASH has increased, with limited effective treatment options. IL-17 plays a protective role in the gut mucosa in high-fat-diet (HFD)-related metabolic disorders, and HFD-related microbiota dysbiosis is responsible for a decreased number of T helper 17 (T H 17) cells in the lamina propria.

The fruiting body formation mechanisms of Cordyceps sinensis are still unclear. To explore the mechanisms, proteins potentially related to the fruiting body formation, proteins from fruiting bodies, and mycelia of Cordyceps species were assessed by using two-dimensional fluorescence difference gel electrophoresis, and the differential expression proteins were identified by matrix-assisted laser desorption/ionisation tandem time of flight mass spectrometry. The results showed that 198 differential expression proteins (252 protein spots) were identified during the fruiting body formation of Cordyceps species, and 24 of them involved in fruiting body development in both C. sinensis and other microorganisms. Especially, enolase and malate dehydrogenase were first found to play an important role in fruiting body development in macro-fungus. The results implied that cAMP signal pathway involved in fruiting body development of C. sinensis, meanwhile glycometabolism, protein metabolism, energy metabolism, and cell reconstruction were more active during fruiting body development. It has become evident that fruiting body formation of C. sinensis is a highly complex differentiation process and requires precise integration of a number of fundamental biological processes. Although the fruiting body formation mechanisms for all these activities remain to be further elucidated, the possible mechanism provides insights into the culture of C. sinensis.

Abundant proteins of human multiple myeloma (MM) were globally analyzed and identified by using two-dimensional gel electrophoresis (2DE) and MALDI-TOF/TOF mass spectrometry (MS). Spots of 517 corresponding to 268 different proteins were detected on 2DE gels of protein lysate from plasma cells isolated from eight newly diagnosed MM patients. These identified proteins were classified into different categories based on their molecular functions and biological processes. The detailed experimental procedures and MS spectra of all the identified proteins have been deposited in the Proteomics Identifications Database (PRIDE) (http://www.ebi.ac.uk/pride) with Accession No. 8846 & 8847. This 2DE map of MM proteins will be an invaluable resource for further proteomics research that investigates proteomic changes associated with biomarker identification and carcinogenesis analysis of multiple myeloma.

ABSTRACT. The peel of mango (Mangifera indica L.) is a special plant tissue that contains many compounds that interfere with protein extraction. A successful separation with Two-dimensional electrophoresis (2-DE) is the key step for proteomic analysis. To evaluate the efficiencies of mango peel protein extraction for 2-DE, four extraction methods were tested: 1) 2-D clean-up kit, 2) trichloroacetic acid/acetone precipitation, 3) phenol extraction, 4) phenol with methanol/ammonium acetate precipitation. The results showed that the phenol with methanol/ammonium acetate precipitation produced the best quality protein extraction and separation. Proteins were separated in 30-70 and >70 kDa ranges better than with the other methods. Acidic proteins had better resolution with fewer horizontal and vertical streaks. Sixteen proteins were identified by maxtrix-assisted laser desorption/ ionisation time-of-flight tandem mass spectrometry (MALDI-TOF/ TOF-MS/MS). The result demonstrated that each of these four methods can be used to prepare mango peel proteins. The phenol with methanol/ ammonium acetate precipitation was the best choice for proteomic analysis of mango peel.

In this study, a recipient-donor co-culture system was used to research the effect of subinhibitory concentrations of antibiotics on horizontal transmission in bacteria and the influence of antibiotics on protein expression. We employed two-dimensional gel electrophoresis combined with mass spectrometry to compare the protein expression profiles in systems with or without 0.5 × the minimum inhibitory concentration of ampicillin. RT-PCR was used to assess the transcriptional levels of the differentially expressed genes. Fifty-seven different proteins were induced or suppressed. The upregulated proteins were involved in transcription and translation, cell wall synthesis, bacterial SOS response, and detoxifying functions, and the downregulated proteins were involved in metabolism. These results indicated that a global response was induced in the recipient-donor co-culture system by the subinhibitory concentration of ampicillin. Further analysis revealed that a global regulatory network based on key pathways was induced in the system in response to the antibiotic pressure. These findings provide a new, more comprehensive view for research on antibiotic-resistance mechanisms in recipient-donor co-culture.