A Comparative Proteomic Study of Cervical Cancer Invasion Garay-Baquero D. J1, Vallejo A. F1, Umaña-Pérez A1, Carlos Garcia2, Fábio C.S. Nogueira2, Gilberto B. Domont2 , Cuervo Patricia3, Sánchez -Gómez M1 1 Hormone Laboratory, Chemistry Department, Universidad Nacional de Colombia, Bogotá, Colombia. e–mail:
[email protected] 2 Departamento de Bioquimica, Instituto de Quimica, Universidade Federal de Rio de Janeiro, Brasil. 3 Laboratorio de Leishmaniosis, Fundación Osvaldo Cruz, Fiocruz, Rio de Janeiro, Brasil
EXPERIMENTAL APPROACH
ABSTRACT Worldwide, cervical cancer is the second cause of female cancer mortality, but nearly 80% of all reported cases come from developing countries; remain a public health problem. The leading cause of death from this disease is metastasis. Key molecular elements of the plasma membrane and cytoskeleton are responsible to cell motility and invasion, therefore are usually considered as tumor targets. Association between invasiveness and membrane related protein profile, was determined in two cervical cancer cell lines, HeLa and C33-A, by a GeL-MS/MS analysis. 100 proteins were identified in high molecular weight fractions, 22 proteins expressed in both cell lines, 34 and 22 proteins were expressed only by C33–A and HeLa, respectively. These protein profiles were associated to the cell-phenotypic characteristics, suggesting some particular features involved in cell invasion and metastasis.
INTRODUCTION Membrane proteomic studies constitute an analytical challenge, due to its dynamic physicochemical characteristics, hydrophobicity and heterogeneity. However, about 60% of the therapeutic targets are directed against membrane proteins, representing an interesting field of study. The aim of this work was to establish the relationship between the expression profile of enriched membrane protein fractions and invasive potential of two cervical cancer cell lines, HeLa cells, associated with a highly invasive phenotype and C33-A, a non-invasive and highly proliferative cell line; through a GeL-MS/MS multidimensional approach supported by bioinformatics analysis using Proteome Software Scaffold.
SIGNALLING NETWORKS AND CELL INVASION POTENCIAL
PROTEINS EXCLUSIVELY EXPRESSED BY CERVICAL CANCER CELL LINES HeLa Cells ACCESS NAME SwissProt MYOF_HUMAN
CD44_HUMAN
MOES_HUMAN
C33 C33— —A Cells MAIN FUNCTION ASSOCIATED
LOCATION
Phospholipid-binding protein / calcium. Endocytic recycling
DNJA1_HUMAN FLNB_HUMAN
TBA4A_HUMAN
Interaction with extracellular matrix. Hyaluronic acid receptor, migration, growth and tumor progression
RPN2_HUMAN Plasma Membrane
2AAA_HUMAN
Molecular cell adhesion
Chaperone. Import of proteins into mitochondria
CALR_HUMAN
Membrane-anchoring cytoskeleton, cell differentiation and signal transduction Cytoskeleton
ACTN4_HUMAN
PDIA6_HUMAN
Positive regulation of movement of cellular components
Positive regulation of movement of cellular components Cellular redox homeostasis. Immune evasion of tumor cells
Binding protein. Protein glycosilation Negative regulation of growth via MAPK inactivation
Plasma Membrane
COMMON PROTEINS EXPRESSED BY CERVICAL CANCER CELL LINES ACCESS NAME SwissProt
Chaperone. Potential lipid anchor. Positive DNJA2_HUMAN regulation of cell proliferation
Iron homeostasis
Microtubule-based movement
LOCATION
Positive regulation of TBRG4_HUMAN cell proliferation
PDIA6_HUMAN ACTN4_HUMAN
MAIN FUNCTION ASSOCIATED
AT1A1_HUMAN Ion Transport
NMD3A_HUMAN Calcium transport, response to ethanol TFR1_HUMAN
ACCESS NAME SwissProt
Cytosol, Extracellular Matrix Cytoskeleton Endoplasmic Reticulum/ Golgi/Plasma Membrane
4F2_HUMAN
Endoplasmic Positive regulation of Reticulum/ cell proliferation Extracellular Matrix Cellular redox homeostasis. Immune evasion of tumor cells
KPYM_HUMAN
Cell proliferation
SERA_HUMAN
Amino acid Biosynthesis
Endoplasmic Reticulum/ Golgi/Plasma Membrane
Cytoplasm
Regulation of U2AF2_HUMAN transcription Nucleus RBBP7_HUMAN DNA Replication
MAIN FUNCTION ASSOCIATED
LOCATION
Transport. Cellular Growth
GRP78_HUMAN
Caspases Inhibitor. Response to glucose starvation
RPN2_HUMAN
N- Glycosilation of proteins
ECHA_HUMAN
Fatty acid metabolism
HSP71_HUMAN
Chaperone. Response oxidative stress
Plasma Membrane
Mitochondria
Cytoplasm FLNA_HUMAN
Cytoskeletal Reorganization
PRKDC_HUMAN
DNA repair
Nucleus
CONCLUSIONS Differential or exclusively expressed proteins in HeLa were mainly involved in cytoskeleton remodeling and associated-cell motility, whilst, in C33-A were associated to proliferation according to the results in functional assays. These results suggest a direct relationship of potential invasion of cervical cancer cells and expression profiles of membrane proteins.
acknowledgement The authors thank La Dirección de Investigación, Sede Bogotá, (DIB), de la Universidad Nacional de Colombia and Colciencias (Contract 374/2008) for the financial support for this research work
