The Super Elongation Complex Family of RNA Polymerase II Elongation Factors: Gene Target Specificity and Transcriptional Output

Abstract

The elongation stage of transcription is highly regulated in metazoans. We previously purified the AFF1-and AFF4-containing super elongation complex (SEC) as a major regulator of development and cancer pathogenesis. Here, we report the biochemical isolation of SEC-like 2 (SEC-L2) and SEC-like 3 (SEC-L3) containing AFF2 and AFF3 in association with P-TEFb, ENL/MLLT1, and AF9/MLLT3. The SEC family members demonstrate high levels of polymerase II (Pol II) C-terminal domain kinase activity; however, only SEC is required for the proper induction of the HSP70 gene upon stress. Genome-wide mRNA-Seq analyses demonstrated that SEC-L2 and SEC-L3 control the expression of different subsets of genes, while AFF4/SEC plays a more dominant role in rapid transcriptional induction in cells. MYC is one of the direct targets of AFF4/SEC, and SEC recruitment to the MYC gene regulates its expression in different cancer cells, including those in acute myeloid or lymphoid leukemia. These findings suggest that AFF4/SEC could be a potential therapeutic target for the treatment of leukemia or other cancers associated with MYC overexpression.T ranscription by RNA polymerase II (Pol II) is a finely tuned and multistep process (40,42,51). After the synthesis of the first few phosphodiester bonds, RNA Pol II escapes from the promoter and enters the productive elongation stage of transcription, depending on the presence of proper environmental signals (40). For decades, the preinitiation complex (PIC) assembly was thought to be the main target of regulation during the entire transcription process. Recently, however, a large number of studies have demonstrated that in addition to the regulation of PIC, promoter-proximal pausing by Pol II and its controlled release is a major regulatory step, especially on developmentally regulated genes (3,7,27,35,38,42,43,57). Multiple elongation factors regulating the elongation stage of transcription have been identified. These include P-TEFb (positive transcription elongation factor), DSIF (DRB sensitivity-inducing factor), NELF (negative transcription elongation factor), and ELL (eleven-nineteen lysinerich leukemia gene) (21,37,40,42,43). DSIF and NELF coordinately participate in setting up paused Pol II at the promoterproximal region (50, 53). The cyclin-dependent kinase 9 (CDK9) module of the P-TEFb complex phosphorylates serine 2 of the Pol II C-terminal domain (CTD), the SPT5 subunit of DSIF, and the E subunit of NELF, leading to the dissociation of paused Pol II from DSIF and NELF for productive elongation (12,21,29,37).The kinase activity of P-TEFb is tightly regulated in vivo through the formation of different complexes to achieve its regulation of transcription elongation. The inactive form of the P-TEFb complex contains 7SK-RNA, MEPCE, LARP7, and HEXIM1, which sequester PTEFb and inhibit its kinase activity (4, 18). The vast majority of P-TEFb exists in this inactive pool (36, 55). P-TEFb was later found to form a complex with the bromodomain protein BRD4. The BRD4/ P-TEFb complex, which can phos...

HIV-1 Tat recruits transcription elongation factors dispersed along a flexible AFF4 scaffold

“…S1), a positive regulator of P-TEFb (33,34). Because the AF9 and ENL C-terminal domains interact competitively with AFF4 (23,29,30), we analyzed one of the paralogs, ENL. To purify complexes of HIV-1 Tat and P-TEFb, we adapted baculovirus coexpression strategies (35).…”

“…In vivo, AFF4 recruits SEC components through defined regions in the first 900 amino acids (23,29,30). AFF4 associates in vivo with P-TEFb, AFF4 300-600 recruits the C-terminus of ELL2, and AFF4 600-900 binds competitively to the C-terminal domain of homologs ENL and AF9.…”

“…This complex belongs to a family of assemblies that have been named "super elongation complexes" (SECs) (27) to reflect roles in normal transcription and "MLL-fusion complexes" (29) because of the activities of certain subunit chimeras in promoting myeloid leukemias. The SECs form a combinatorial family of related assemblies containing homologous subunits and also normally are recruited to a subset of human genes occupied by paused polymerases (30,31). In embryonic stem cells, several SEC components reside at actively transcribed genes and are required for stimulating transcription during differentiation (31), pointing to a wider, essential role for these factors in regulating transcriptional elongation.…”

“…In embryonic stem cells, several SEC components reside at actively transcribed genes and are required for stimulating transcription during differentiation (31), pointing to a wider, essential role for these factors in regulating transcriptional elongation. mRNA-knockdown experiments suggest some redundancy among members of the AFF and ELL families, but these factors also may have specialized functions (18,30). For example, HIV transcription is stimulated specifically by AFF4 and ELL2 (26,28,32).…”

p53 represses the transcription of snRNA genes by preventing the formation of little elongation complex

“…Importantly, it has been shown that SEC is required for c-Myc expression in several leukemia cell lines, suggesting that SEC is a potential therapeutic target for the treatment of leukemia and other solid cancers associated with overexpression of the c-Myc gene [35]. p53 has also been shown to repress the expression of c-Myc [36]; however, the mechanism by which p53 regulates the expression of c-Myc has not been elucidated.…”