Proteomic profiling identifies breast tumor metastasis-associated factors in an isogenic model.
A combination of LC and MS was applied to an isogenic breast tumor metastasis model to identify proteins associated with a cellular phenotype. Chromatofocusing followed by nonporous-RP-HPLC/ESI-TOF MS was applied to cell lysates of a pair of monoclonal cell lines from the human breast carcinoma cell line MDA-MB-435 that have different metastatic phenotypes in immune-compromised mice. This method was developed to separate proteins based on pI and hydrophobicity. The high resolution and mass accuracy of ESI-TOF measurements provided a good correlation of theoretical MW and experimental Mr values of intact proteins measured in mass maps obtained in the pH range 3.8-6.4. The isolated proteins were digested by trypsin and analyzed by MALDI-TOF MS, MALDI-QIT-TOF MS, and monolith-based HPLC/MS/MS. The unique combination of the techniques provided valuable information including quantitation and modification of proteins. We identified 89 selected proteins, of which 43 were confirmed as differentially expressed. Metastasis-associated proteins included galectin-1, whereas annexin I and annexin II were associated with the nonmetastatic phenotype. In this study, we demonstrate that combining a variety of MS tools with a multidimensional liquid-phase separation provides the ability to map cellular protein content, to search for modified proteins, and to correlate protein expression with cellular phenotype.
Kreunin
Paweena
P
Department of Chemistry, University of Michigan, Ann Arbor, MI, USA.
Yoo
Chul
C
Urquidi
Virginia
V
Lubman
David M
DM
Goodison
Steve
S
D014408:Biomarkers, Tumor; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D005260:Female; D020869:Gene Expression Profiling; D006801:Humans; D008954:Models, Biological; D009362:Neoplasm Metastasis / Q000235:genetics / Q000473:pathology*; D009363:Neoplasm Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D040901:Proteomics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins