Figure 3.

mcircRasGEF1B is predominantly located in cytoplasm and is not translated. (A) RAW264.7 cells were induced with and without LPS for 2 hours. Whole cell lysates were fractionated into cytoplasmic and nuclear fractions. The levels of L32, U6, mlinRasGEF1B and mcircRasGEF1B in these fractions were measured by qRT-PCR. All experiments were carried out in duplicates. (*, p < 0.05; **, p < 0.01). (B) RAW264.7 cells were induced with for 2 hours. Cytoplasmic supernatants were separated by sucrose gradient centrifugation. The levels of linear transcripts (mlinRasGEF1B, A20, TNFα, IP10, IκBα, ICAM-1 and GAPDH), and circular transcript mcircRasGEF1B, in free mRNA and polysome-bound fractions were measured by qRT-PCR (n = 2).