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ORIGINAL ARTICLE: Suppression of Mamu‐AG by RNA Interference
Author(s) -
Drenzek Jessica G.,
Vidiguriene Jolanta,
Vidiguris Geminis,
Grendell Richard L.,
Dambaeva Svetlana V.,
Durning Maureen,
Golos Thaddeus G.
Publication year - 2009
Publication title -
american journal of reproductive immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.071
H-Index - 97
eISSN - 1600-0897
pISSN - 1046-7408
DOI - 10.1111/j.1600-0897.2009.00706.x
Subject(s) - gene knockdown , rna interference , small hairpin rna , small interfering rna , biology , transfection , cytotoxicity , major histocompatibility complex , microbiology and biotechnology , flow cytometry , cytolysis , cell , human leukocyte antigen , rna , cell culture , antigen , gene , immunology , in vitro , genetics
Problem The role of placental major histocompatibility complex (MHC) class I molecules in pregnancy is not well understood. Mamu‐AG, the rhesus monkey homology of human leukocyte antigen (HLA)‐G expressed in the human placenta, was targeted for degradation by RNA interference (RNAi), a powerful tool to aid in determining gene function, to determine the effect that this knockdown has on NK cell function. Method of study A series of potential target short hairpin RNA (shRNA) sequences to suppress Mamu‐AG expression was screened, which identified an optimal sequence to use in transfection experiments. Knockdown in two different Mamu‐AG‐expressing cell lines was measured by flow cytometry. Cytotoxicity assays were performed to correlate Mamu‐AG expression with NK cell cytotoxicity. Results Decreased expression of Mamu‐AG by short interfering RNA (siRNA) (70–80%) in cell types tested was associated with increased lysis of Mamu‐AG target cells. Conclusion Target sequences have been identified that knocked down Mamu‐AG expression by RNAi and increased lysis by NK cells. This supports the concept that NK cell receptors recognize this placental non‐classical MHC class I molecule.