The faithful and efficient transcription of genomic DNA into mRNA is crucial for cell survival under DNA damage caused by UV irradiation, oxidative stress or chemical DNA modifications. Similar to arrest of Pol II after making transcription error, DNA lesions can interfere with replication, potentially causing mutations in DNA, and also hinder transcription, affecting genome stability and regulation of gene expression. To maintain genomic integrity, cells have evolved separate cellular strategies involving multiple DNA damage repair and DNA damage tolerance mechanisms. Non-bulky DNA lesions are preferentially repaired by the base excision repair (BER) pathway while damages that cause large DNA distortion, such as UV light-induced cyclobutane pyrimidine dimers (CPDs)/cisplatin adducts and oxidative cyclopurines are primarily subject to the nucleotide excision repair (NER) pathway. Despite ongoing repair, some lesions escape detection, presenting the cell with a challenge for continued DNA and RNA synthesis. During replication, the deleterious effect of DNA lesions can be alleviated by translesion DNA synthesis [TLS]. During TLS, the high-fidelity replicative DNA polymerases are switched transiently to specialized translesion DNA polymerases that can accommodate bulky lesions within a more spacious active site, thus enabling their bypass. Recently, we demonstrated that yeast Pol II employs a distinct mechanism for CPD bypass (called A-rule) involving a conformational flexibility of its active center (the mobile trigger loop domain in Rpb1 subunit) facilitating accommodation of bulky lesions. We recently expanded this project to Pol II bypass of bulky cyclopurine (CyPn) oxidative damages in transcribed genes and to analysis of RTLS (RNA polymerase translesion synthesis) through the bulky lesions by mammalian (calf thymus, CT) Pol II. We also plan to identify and test in vitro protein factors involved in RTLS and in initiation of TCR in yeast and mammalian cells. We have already found that mammalian TFIIF significantly promotes RTLS by CT Pol II in vitro.