Date Published: April 25, 2019
Publisher: Public Library of Science
Author(s): Ting-Hsuan Wu, Lingfang Shi, Anson W. Lowe, Mark R. Nicolls, Peter N. Kao, Srinivas Mummidi.
Immediate early gene (IEG) transcription is rapidly activated by diverse stimuli. This transcriptional regulation is assumed to involve constitutively expressed nuclear factors that are targets of signaling cascades initiated at the cell membrane. NF45 (encoded by ILF2) and its heterodimeric partner NF90/NF110 (encoded by ILF3) are chromatin-interacting proteins that are constitutively expressed and localized predominantly in the nucleus. Previously, NF90/NF110 chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) in K562 erythroleukemia cells revealed its enriched association with chromatin at active promoters and strong enhancers. NF90/NF110 specifically occupied the promoters of IEGs. Here, ChIP in serum-starved HEK293 cells demonstrated that NF45 and NF90/NF110 pre-exist and specifically occupy the promoters of IEG transcription factors EGR1, FOS and JUN. Cellular stimulation with phorbol myristyl acetate increased NF90/NF110 chromatin association, while decreasing NF45 chromatin association at promoters of EGR1, FOS and JUN. In HEK293 cells stably transfected with doxycycline-inducible shRNA vectors targeting NF90/NF110 or NF45, doxycycline-mediated knockdown of NF90/NF110 or NF45 attenuated the inducible expression of EGR1, FOS, and JUN at the levels of transcription, RNA and protein. Dynamic chromatin association of NF45 and NF90/NF110 at IEG promoters are observed upon stimulation, and NF45 and NF90/NF110 contribute to inducible transcription of IEGs. NF45 and NF90/NF110 operate as chromatin regulators of the immediate early response.
The rapid cellular response that occurs upon recognition of biological or environmental signals is crucial for adaptation and survival of the organism [1–3]. The subset of genes that are rapidly expressed upon induction are termed immediate early genes (IEG) . Inducible expression of IEGs in response to diverse regulatory signals underlies acute inflammation [5–8], neuronal activity , cell proliferation, and differentiation [1, 10, 11]. Aberrant expression of IEGs is involved in malignant cellular transformation  and is a feature of diverse cancers [13, 14].
We demonstrate that NF45 and NF90/NF110 exhibit dynamic chromatin association at the proximal promoters of the ‘forward-driving’ IEG transcription factors, EGR1, FOS and JUN in HEK293 cells. Genetic attenuation of NF45 or NF90/NF110 reduced PMA-stimulated transcription of EGR1, FOS and JUN. Our findings confirm and extend those of Nakadai et al. characterizing NF45 and NF90/NF110 as transcriptional coactivators of FOS. Operating as hierarchical transcriptional regulators of ‘forward-driving’ IE transcription factors, EGR1, FOS and JUN, NF45 and NF90/NF110 represent novel targets for regulation of acute inflammation, neuronal activity, cell proliferation, and differentiation. Increased expression of IEGs in malignant cellular transformation may be a consequence of overexpression of NF45 and/or NF90/NF110.