Reactive oxygen species (ROS), at physiological levels, mediate redox control of numerous cellular processes. Excessive ROS levels cause cellular damage and are associated with many human pathophysiologies. Maintenance of redox homeostasis is essential to ensure steady-state equilibrium of ROS production versus ROS elimination. At physiological levels, hydrogen peroxide (H2O2) serves as a second messenger and activates/regulates cellular signals through reversible oxidation of protein cysteine thiolate residues (RS−) to form protein sulfenic acids (protein-SOH). H2O2 activates cell signaling pathways involving kinases that use ATP to phosphorylate protein targets on serine or threonine residues. These signaling pathways include the mitogen-activated protein kinases (MAPK), apoptosis signal-regulating kinase 1 (ASK1) and the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) (PI3K/Akt/mTOR) pathway. ROS regulate several transcription factors including nuclear factor erythroid 2–related factor 2 (Nrf2), hypoxia-inducible factor 1α (HIF1α), p53, nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), activator protein 1 (AP1) and heat shock factor 1 (HSF1). The NAD+-dependent histone deacetylase Sirtuin 1 (SIRT1), which regulates Forkhead box class O (FoxO), is regulated by ROS. Given that H2O2 can regulate gene expression that affects essential cellular processes, improved understanding of mechanisms involved in transcriptional regulation could guide new treatment approaches for redox-associated diseases.