β-Actin regulates interleukin 6-induced p21 transcription by interacting with the Rpb5 and Rpb7 subunits of RNA polymerase II

Abstract

In pre-initiation complexes, RNA helicase A interacts with β-actin and acts as a bridging factor linking nuclear actin with RNA polymerase II (Pol II). In addition, β-actin participates in Pol IIdependent transcription elongation by interacting with the positive transcription elongation factor Cdk9. However, many relationships between β-actin and Pol II remain to be identified. In an interleukin 6 (IL-6)-induced p21 expression model, we demonstrated that β-actin knockdown reduced p21 expression. Immunofluorescence analysis showed that the colocalization of β-actin and Pol II increased significantly in cells treated with IL-6. It is known that the Rpb5, Rpb6 and Rpb7 subunits are located at the surface of the enzyme. We next constructed recombinant pcDNA-HA-Rpb5, pcDNA-HA-Rpb6 and pcDNA-HA-Rpb7 plasmids and expressed the three polymerase II subunits in HepG2 cells. We found that β-actin could be immunoprecipitated with HA-Rpb5 and HA-Rpb7. A Glutathione-S-transferase pull-down assay revealed that β-actin was associated with Rpb5 and Rpb7 in vitro. Furthermore, overexpression of Rpb5 and Rpb7 in cells reduced p21 expression significantly, suggesting that Rpb5 and Rpb7 competitively interact with β-actin. This study shows that β-actin associates with Pol II subunits through direct proteinprotein interactions and provides fundamental insight into Pol II transcriptional regulation.

Nuclear actin dynamics in gene expression and genome organization

“…In addition to regulating transcription initiation through transcription factors, actin has also been directly linked to RNA polymerase (Pol) function. Actin co-purifies with all three RNA polymerases [2,[70][71][72] and has been shown to interact with Pol II subunits Rpb5 and Rpb7 [73], as well as Pol III subunits Rpc3, Rpb6 and Rpb8 [72] in co-immunoprecipitation experiments. However, these interactions are poorly understood at the molecular level.…”

Mechanotransduction and epigenetic modulations of chromatin: Role of mechanical signals in gene regulation

“…217 Along with the transcriptional regulation by interacting with other transcription factors, nuclear actin also directly binds to RNA polymerases and regulates their function. It has been reported that actin, specifically β-actin, interacts with Rbp5 and Rbp7 subunits of RNA polymerase II (RNA Pol II) 250 and also Rpb6, Rpb8, and Rpc3 subunits of RNA Pol III. 251 Studies highlighting the function of actin in transcriptional regulation and chromatin dynamics are reviewed in the following references 211,251 ; however, the underlying molecular mechanism is still enigmatic.…”

Actin filaments accumulate in the nucleus during interphase and remain in the vicinity of condensing chromosomes during prophase to metaphase in zebrafish early embryos

“…In the cytoplasm, actin serves a role in cell migration, stress fiber formation, and cell division by forming a contractile ring during cytokinesis. Actin is also present in nuclei, where it plays a crucial role in transcriptional regulation (Vartiainen et al ., 2007; Baarlink et al ., 2013; Tian et al ., 2016; Sokolova et al ., 2018) and in DNA damage repair processes (Chiolo et al ., 2011; Belin et al ., 2015; Caridi et al ., 2019). Actin nuclear localization is mediated through the interplay of nuclear import regulated by importin 9 with cofilin (Dopie et al ., 2012) and nuclear export regulated by exportin 6 (Exp6), which transports profilin-actin complexes (Stüven et al ., 2003).…”