Two-dimensional separations of the genome and proteome of neuroblastoma cells.
Two-dimensional (2-D) electrophoretic methods have been available that allow separation of the protein constituents of a cell population. It has also become feasible to electrophoretically separate in two dimensions and to display DNA fragments derived from genomic digests. Through the appropriate choice of restriction enzymes, the functional component of the genome that encompasses CpG islands can be preferentially visualized in 2-D gels. The same computerized approach for the analysis of 2-D patterns can be applied to investigations at either the protein or DNA levels. Our group has utilized 2-D electrophoresis to investigate both protein and DNA changes in cancer. The emphasis to date has been on the identification of proteins, the abundance of which is related to specific biological features of the tumors analyzed and of DNA fragments encompassed in genomic amplifications, as the latter commonly contain growth-related genes. Findings derived from our analysis of neuroblastoma tumors and cell lines using 2-D approaches are reviewed. Data for four proteins observed in 2-D gels are presented because of our demonstrated association of these proteins with differentiation and proliferation properties of neuroblastoma. At the genomic level, the detection of amplifications using 2-D gels has necessitated an understanding of the variability displayed by multi-copy genomic fragments, which we have accomplished to a large part and which we present. An important benefit of 2-D approaches is the efficiency of scale and the ease with which abundant proteins or multicopy genomic fragments can be detected, identified and quantitatively analyzed.
Wimmer
K
K
University of Michigan School of Medicine, Department of Pediatric Hematology/Oncology, Ann Arbor, USA.
Kuick
R
R
Thoraval
D
D
Hanash
S M
SM
D002454:Cell Differentiation / Q000187:drug effects; D002458:Cell Fractionation / Q000379:methods*; D003001:Cloning, Molecular; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D016193:G1 Phase; D005784:Gene Amplification; D015972:Gene Expression Regulation, Neoplastic; D016259:Genes, myc; D016678:Genome; D006360:Heat-Shock Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D008868:Microtubule Proteins; D020559:Monomeric GTP-Binding Proteins; D054778:NM23 Nucleoside Diphosphate Kinases; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D009367:Neoplasm Staging; D009447:Neuroblastoma / Q000737:chemistry* / Q000235:genetics; D009701:Nucleoside-Diphosphate Kinase; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D018809:Proliferating Cell Nuclear Antigen / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D050777:Stathmin; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D006360:Heat-Shock Proteins; D008868:Microtubule Proteins; D054778:NM23 Nucleoside Diphosphate Kinases; D009363:Neoplasm Proteins; D010750:Phosphoproteins; D018809:Proliferating Cell Nuclear Antigen; D050777:Stathmin; D014157:Transcription Factors; D014212:Tretinoin; D009701:Nucleoside-Diphosphate Kinase; D020559:Monomeric GTP-Binding Proteins