Hypoxia increases m6A in poly(A)+ RNA but not in total or ribosomal RNA. RNA isolated from HEK-293T cells grown in normoxic (Normox) or hypoxic (Hypox) conditions for 24 h. (A) LC-MS/MS of mRNA from HEK-293T cells. Values represent the amount of m6A divided by total adenosine (N of 3). (*) P ≤ 0.05 by paired Student's t-test. Error bars represent standard error of the mean (SEM). (B) LC-MS/MS of total RNA from HEK-293T cells. Values represent the amount of m6A divided by total adenosine (N of 2). (C) LC-MS/MS of 18 and 28S rRNA from HEK-293T cells. Values represent the amount of m6A divided by total adenosine (N of 3). (D) MeRIP of 100 ng of mRNA from HEK-293T cells grown in normoxic or hypoxic conditions for 24 h quantified by qRT-PCR. Fold enrichments calculated from immunoprecipitated mRNA levels normalized to input and bead-only negative control IP and expressed as a ratio of hypoxia/normoxia. (*) P ≤ 0.05 by paired Student's t-test. Error bars represent SEM of five experiments.
