Date Published: June 3, 2019
Publisher: Public Library of Science
Author(s): Noor Kazim, Abhinav Adhikari, Judith Davie, Atsushi Asakura.
The transcription elongation factor TFIIS is encoded by a three member gene family in vertebrates. Here we show that one member of this family, TCEA3, is upregulated during skeletal muscle differentiation and acts to promote gene activation by the myogenic regulatory family of transcription factors, which includes MyoD and myogenin. We show that myogenin is a direct regulator of Tcea3. Myogenin binds to the Tcea3 promoter and is required to recruit RNA polymerase II. TCEA3 can bind to both myogenin and MyoD and is co-recruited with the MRFs to promoters dependent on the MRFs. Depletion of myogenin inhibits the recruitment of TCEA3, suggesting that the interaction of TCEA3 with the MRFs serves to aid in recruitment to target promoters. Like TFIIS, we show that TCEA3 interacts with RNA polymerase II. TCEA3 travels with the elongating RNA polymerase II in the coding region of genes and depletions of TCEA3 inhibit the recruitment of RNA polymerase II to promoters. In proliferating cells, TCEA3 expressed at low levels and is present in both the nucleus and cytoplasm. However, upon differentiation, TCEA3 is upregulated and transported exclusively to the nucleus. Thus, our data show that TCEA3 is a required co-factor for MRF driven gene expression during myogenesis.
TFIIS is a transcription elongation factor conserved through the eukaryotic linage. In vertebrates; including frog, mouse and human, TFIIS is represented by a gene family which includes TCEA1, TCEA2 and TCEA3 . TCEA1 is the gene most closely related to the sole TFIIS present in non-vertebrates and best characterized in the yeast, S. cerevisiae . TCEA1 is ubiquitously expressed and has been shown to regulate the proliferation and differentiation of myeloid cells . Recent work suggest that TCEA1 can also promote proliferation in hepatic carcinoma cells . However, the functions of the other TFIIS isoforms in normal cells or cancer cells are poorly defined. TCEA3 expression is tissue restricted and is known to be expressed in intestine, heart, testis, kidney and skeletal muscle . TCEA3 is also expressed in mouse embryonic stem cells and functions in lineage determination though regulation of the Nodal pathway . TCEA3 has also been shown to regulate the pluripotent differentiation potential of mouse embryonic stem cells . TCEA3 has also been shown to function as a tumor suppressor in ovarian cancer cells  and promotes apoptosis in gastric cancer cells . TCEA3 has recently been shown to promote the differentiation of bovine skeletal muscle cells .
We found that TCEA3 is directly regulated by myogenin, which upregulates TCEA3 upon differentiation. TCEA3 is then recruited to MRF dependent genes through interactions with MYOD1, MYOG and RNAPII and enhances RNAPII recruitment to promoters. TCEA3 then travels with RNAPII along the coding region of the gene. Thus, TCEA3 is a previously undescribed co-factor in MRF dependent gene activation. Our data shows that TCEA3 both contributes to RNAPII recruitment to promoters as well as travels with RNAPII during elongation, suggesting that TCEA3 contributes to enhancing elongation by RNAPII. Recombinant Xenopus proteins representing the three TFIIS family members have been shown to cleave RNA in a stalled RNAP II elongation complex , suggesting that all three family members can function as transcription elongation factors. It is intriguing that skeletal muscle expresses TCEA3, which is the most divergent from the other two TFIIS family members in the linker region that was the region shown to interact with RNAPII . Given this difference, we anticipated that TCEA3 might not interact with RNAPII, but our data clearly show that TCEA3 can interact with RNAPII. It is possible that the longer linker region in TCEA3 allows for additional interactions that promote its activity, such as the interactions with MYOG and MYOD1. TCEA3 has also been shown to be a target of P53, where TCEA3 then contributes to activation of selective P53 targets such Bax, but not Cdkn1a (p21) . These interactions may also function in skeletal muscle, but it is intriguing that our results suggest a similar regulatory circuit, with MYOG activating TCEA3 which then promotes MYOG activation of gene targets.