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Effects of lead on transcriptional activity of the zinc finger Gata‐1 in erythropoiesis
Author(s) -
Oosterhouse Elisabeth,
Aguilar Silvia Zambrano,
Pikaart Michael
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a80-a
Subject(s) - zinc finger , transfection , microbiology and biotechnology , chemistry , transcription factor , gata1 , erythropoiesis , electroporation , blot , dna , dna binding domain , electrophoretic mobility shift assay , plasmid , gata2 , cell , gata transcription factor , biology , gene , biochemistry , gene expression , promoter , anemia , medicine
Gata‐1 is a transcription factor involved in the cell maturation from stem cells to fully functioning erythrocytes. Since Pb is known to cause anemia, and has a high affinity for the cystine residues present on gata‐1 where zinc is bound, its possible role in altering gata‐1 function is being investigated. Previous studies show that binding of the gata‐1 minimal DNA binding domain to DNA is abrogated by Pb in vitro and in yeast cells. Currently, the effects of Pb on the full length gata‐1 are being studied using mouse erytholeukemia (MEL) cells. Gel shift assays have been performed using nuclear cell extracts from MEL cells that have been treated with Pb or without Pb. Results show subtle differences in the DNA binding interaction. Similarly, assays have been performed adding Pb to the extract directly prior to running the gel with analogous results. Determination of total gata‐1 levels by western blotting revealed no detectable difference in nuclear gata‐1 concentration in the presence or absence of Pb. At present, the effects of DNA binding of purified bacterially expressed full‐length gata‐1 are being studied to assess whether the presence of other nuclear proteins in the crude MEL extracts confers a protective effect. In addition, transfection procedures involving electroporation are being optimized where reporter plasmids are taken up into MEL cells thereby detecting gata‐1 activity.