The Forkhead Transcription Factor Foxi1 Is a Master Regulator of Vacuolar H+-ATPase Proton Pump Subunits in the Inner Ear, Kidney and Epididymis (original) (raw)
Figure 6
Chromatin Immuoprecipitation (ChIP) analysis.
Chromatin immunoprecipitation confirm presence of Foxi1- ATP6V0A4 promoter interaction at FK1-3. Transfected 3T3-L1 cells were fixed, and chromatin was prepared by sonication. After preclearance with Protein G Sepharose the chromatin was immunoprecipitated with or without an Anti-6X His tag antibody (Foxi1, No Ab). The purified precipitated DNA was used as template for PCR reactions with specific primers covering sequence with Foxi1 binding sites (Fk1-3) or unspecific primers (Random). ChIP analysis reveal a strong interaction of the ATP6V0A4 promoter with wt Foxi1 (Lane 1). The interaction is weakened using promoter with mutated FK1 (Lane2) and even more weakened with Fk2 and 3 (Lanes 3–4). Using the triple mutated promoter Fk1-3 (Lane 5) no interaction is detected.