For all the transgene constructs (Trpc6-wild type, Trpc6-HA P111Q and Trpc6-HA E896K) the CMV promoter was replaced with the human podocin promoter (NPHS2) [20] (kindly donated by Dr. Holzman) utilizing the same two step strategy.
All utilize the county jail.
The Glu-Cre transgenic mice were generated utilizing the pBKCMV/Glu-CreGH plasmid kindly provided by Dr. M. Magnuson.
For enzymatic activity analysis, the mouse PLD2 expression vector pcGN-mPLD2 [14], which was kindly provided by Dr. M. Frohman, was utilized.
The rabbit anti-human VASH2 monoclonal antibody utilized in the western blot analyses was kindly provided by Professor Sato.
The Escherichia coli cysteine auxotroph, BL21- λDE3) selB::kan cys51E(49) (kindly provided by Dr. Paul Ortiz de Montellano, University of California, San Francisco), henceforth called cysE, was utilized in these preparations.
pGEM4Z/GFP/A64 plasmid containing the poly(A) template (kindly provided by Dr. E. Gilboa, University of Miami, FA, USA) was utilized for in vitro transcription of EGFP mRNA.
Samples were kindly generated (Acknowledgements) for the purpose of developing an observed reference peptide database for each organism, utilizing the high-throughput proteomic capabilities present at Pacific Northwest National Laboratory, Richland WA.
Antigen-specific stimulation utilized 2 µg/ml of overlapping peptide pools from either a consensus gp120 of HIV clade C (kindly provided by Dr. Christian Brander), or from SIVmac239 Gag (AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH, 15mers).
We also utilized the expO datasets (1185 tumors on the Affymetrix U133plus2 mRNA expression platform) available online as part of the Gene Expression Omnibus (GEO/NCBI) and kindly provided by the International Genomics Consortium.
