Microarray Studies on the STAT5A MUTANT.

Stat5+/- mice were bred into the C57BL/6 background. Stat5+/- mice were intercrossed and mouse embryonic fibroblasts (MEFs) were isolated from 12.5-13.5-day WT or Stat5-/- fetuses. The retroviral-expression vector carrying a DNA binding domain mutant Stat5A (E437/E438?AA) or tyrosine-phosphorylated Stat5A Y694F mutation based on an MSCV-IRES-GFP backbone (gift from Richard Moriggl, Ludwig-Boltzmann Institute, Vienna, Austria) was infected into Stat5-/- MEFs. FACS was used to select GFP+ cells. After 5 hours starvation in serum free medium with 0.1% of BSA, MEFs were treated with growth hormone for 2 hours. Total cellular RNA from each group of the MEFs was extracted with TRIzol reagent (Invitrogen) according to the manufacturer's instructions. Microarray analyses were performed using Affymetrix Mouse Genome 430 2.0 GeneChips (Affymetrix, Santa Clara, CA) (four groups, biological replicates for each group). Expression values were determined with GeneChip Operating Software (GCOS) v1.1.1 software. RMA signals were summarized using GeneSpring GX 10.0.1 (Agilent) and normalized by quantile normalization. All data analysis was performed with GeneSpring software GX 10.01.

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