PROTEOMIC EVALUATION OF THE MOUSE UNK CELL PLASMA MEMBRANE

BIZINOTTO M. C.(1), SOUZA G. H. M. F.(2), MARTINS D.(3), OLIVEIRA B. M.(3), EBERLIN M. N.(2), YAMADA A. T.(1)

(1)Department of Histology and Embryology, Institute of Biology; (2)Thomson Mass Spectrometry Laboratory, Institute of Chemistry; (3)Chemistry of Proteins Laboratory Biochemistry, Institute of Biology,

The unique behavior of Natural Killer cell present in the uterine environment (uNK) during pregnancy suggests the expression of specific receptors not shared by circulating NK (cNK) cells. In mice, the uNK cell membrane expresses glycoconjugates with carbohydrate moiety containing N-acetyl D-galactosamine at terminal portion reactive to Dolichos biflorus (DBA) lectin. This lectin has been considered the most useful way to distinguish uNK from cNK. In the present work it was aimed to isolate proteins from mouse uNK cell plasma membrane and performed a proteomic evaluation to identify the proteins reactive to DBA lectin. Mouse uNK cells was isolated from pregnant uterus with lectin-biomagnetic beads and obtained the homogenate of plasma membrane enriched fraction. The plasma membrane protein homogenate was submitted to 2D electrophoresis and blotted to PVDF membrane and reacted with DBA lectin. The DBA positive corresponding spots were excised from the gels and submitted to tryptic digestion and the respective peptide masses were acquired with a MALDI-TOF-MS mass spectrometer. Analyses were performed in reflectron mode with positively charged ions. Database searches were done with MASCOT program (http://www.matrixscience.com) using NCBI and SWISS-PROT databases to achieve a panel of candidate proteins. Among the candidate proteins, it was noticed the HSP-70 and COG4 as potentially committed on uNK cells activities at maternal-fetal interface.

KEY WORDS: Uterine-NK cell, proteome, Reproductive Immunology

FINANCIAL SUPPORT: Graduate Student scholarship recipient from FAPESP.

CORRESPONDENCE TO:

Marcia C Bizinotto, Department of Histology and Embryology, Institute of Biology, UNICAMP, CEP 13083-970, Campinas-SP, Brazil. Email: bizinott@unicamp.br