RNA polymerase II (Pol II) is the main engine that drives transcription of protein-encoding genes in eukaryotes. Despite its intrinsic subunit complexity, Pol II is subject to a host of factors that regulate the multistep transcription process. Indeed, the hallmark of the transcription cycle is the dynamic association of Pol II with initiation, elongation and other factors. In addition, Pol II transcription is regulated by a series of cofactors (coactivators and corepressors). Among these, the Mediator has emerged as one of the key regulatory factors for Pol II. Transcription by Pol II takes place in the context of chromatin, which is subject to numerous epigenetic modifications. This chapter mainly summarizes the various biochemical mechanisms that determine formation and function of a Pol II preinitiation complex (PIC) and those that affect its progress along the gene body (elongation). It further examines the various epigenetic modifications that the Pol II machinery encounters, especially in certain developmental contexts, and highlights newer evidence pointing to a likely close interplay between this machinery and factors responsible for the chromatin modifications.