Identification and Functional Validation of RAD23B as a Potential Protein in Human Breast Cancer Progression

Abstract

Identification of protein targets that play a role in breast cancer invasion may help to understand the rapid progression of cancer and may lead to the development of new biomarkers for the disease. In this study, we compared two highly invasive and two poorly invasive breast cancer cell lines using comparative label-free LC-MS profiling in order to identify differentially expressed proteins that may be linked to the invasive phenotype in vitro. Forty-five proteins were found to be upregulated, and 34 proteins, downregulated. UV excision repair protein RAD23 homologue B (RAD23B) was found among the downregulated proteins in highly invasive breast cancer cell lines. In poorly invasive breast cancer cell lines, siRNA-mediated downregulation of RAD23B subsequently led to an increase in invasion and adhesion in vitro. Immunohistochemistry analysis of 164 specimens of invasive breast cancer showed that having a high percentage (>80%) of RAD23B positive nuclei was significantly associated with histopathological grades 1 and 2 breast cancer and with low mitotic activity. In addition, a high staining intensity for RAD23B in the cytoplasm was significantly associated with histopathological grade 3 breast cancer. This study suggests a potential role of RAD23B in breast cancer progression and may further imply a tumor suppressor role of nuclear RAD23B in breast cancer.

Proteomic profiling of mdx-4cv serum reveals highly elevated levels of the inflammation-induced plasma marker haptoglobin in muscular dystrophy

“…The exported MS/MS spectra from Progenesis software were used for peptide identification using Proteome Discoverer 2.0 against Mascot (version 2.3) and Sequest HT and searched against the UniProtKB-SwissProt database (taxonomy: Mus musculus ). The following search parameters were used for protein identification: i) peptide mass tolerance set to 10 ppm, ii) MS/MS mass tolerance set to 0.02 Da, iii) up to two missed cleavages were allowed, iv) carbamidomethylation set as a fixed modification and v) methionine oxidation set as a variable modification (54). For re-importation back into Progenesis LC-MS software for further analysis, only peptides classified as high confidence peptides and with ion scores of 40.00 or more (from Mascot) and/or XCorr scores >1.5 for singly charged ions, >2.0 for doubly charged ions and >3.0 for triply charged ions (Sequest HT, Thermo Fisher Scientific) were selected.…”

Concurrent Label-Free Mass Spectrometric Analysis of Dystrophin Isoform Dp427 and the Myofibrosis Marker Collagen in Crude Extracts from mdx-4cv Skeletal Muscles

“…Firstly the data was aligned based on the LC retention time of each sample [65]. This allows for any drift in retention time, thus giving an adjusted time for all runs in the analysis.…”

Structures of Rpn1 T1:Rad23 and hRpn13:hPLIC2 Reveal Distinct Binding Mechanisms between Substrate Receptors and Shuttle Factors of the Proteasome

“…Rad23 proteins are similarly associated with neuroprotection activities (Tsou et al, 2015), but are most well known for their function in nucleotide excision repair (Mueller and Smerdon, 1996; Watkins et al, 1993). hHR23b (a human ortholog of Rad23) is downregulated in highly invasive breast cancer cell lines (Linge et al, 2014), associated with risk of lung, laryngeal and bladder cancer (Abbasi et al, 2009; Chang et al, 2008; Chen et al, 2007), and substitution of A249 for valine in this protein is associated with increased esophageal cancer risk (Pan et al, 2009). …”