Integrator orchestrates RAS/ERK1/2 signaling transcriptional programs.

Jingyin Yue,Ramin Shiekhattar,Felipe Beckedorff,Anda Zhang,Chiara Pastori,Fan Lai

doi:10.1101/gad.301697.117

Abstract

Activating mutations in the mitogen-activated protein kinase (MAPK) cascade, also known as the RAS-MEK-extracellular signal-related kinase (ERK1/2) pathway, are an underlying cause of >70% of human cancers. While great strides have been made toward elucidating the cytoplasmic components of MAPK signaling, the key downstream coactivators that coordinate the transcriptional response have not been fully illustrated. Here, we demonstrate that the MAPK transcriptional response in human cells is funneled through Integrator, an RNA polymerase II-associated complex. Integrator depletion diminishes ERK1/2 transcriptional responsiveness and cellular growth in human cancers harboring activating mutations in MAPK signaling. Pharmacological inhibition of the MAPK pathway abrogates the stimulus-dependent recruitment of Integrator at immediate early genes and their enhancers. Following epidermal growth factor (EGF) stimulation, activated ERK1/2 is recruited to immediate early genes and phosphorylates INTS11, the catalytic subunit of Integrator. Importantly, in contrast to the broad effects of Integrator knockdown on MAPK responsiveness, depletion of a number of critical subunits of the coactivator complex Mediator alters only a few MAPK-responsive genes. Finally, human cancers with activating mutations in the MAPK cascade, rendered resistant to targeted therapies, display diminished growth following depletion of Integrator. We propose Integrator as a crucial transcriptional coactivator in MAPK signaling, which could serve as a downstream therapeutic target for cancer treatment.

Highlights

The canonical mitogen-activated protein kinase (MAPK) or extracellular signal-related kinase (ERK1/2) cascade is one of the key signaling pathways that transmits growth signals to the nucleus (Chen et al 1992; Gonzalez et al 1993; Karin and Hunter 1995)
To dissect the signaling pathway that mediates the epidermal growth factor (EGF) transcriptional response of immediate early genes (IEGs), we treated HeLa cells with an ERK1/2 (SCH772984) or MEK (PD0325901) inhibitor prior to EGF stimulation and analyzed EGF-responsive gene expression using chromatin RNA sequencing (ChromRNA-seq), which provides for an enriched fraction of nascent RNAs
While Integrator knockdown (INTS11 knockdown using an inducible shRNA) did not affect ERK1/2 activation (Supplemental Fig. S2B), it mimicked the pharmacological inhibition of MAPK inhibitors, resulting in the loss of EGF responsiveness in the same group of genes

Summary

Introduction

The canonical mitogen-activated protein kinase (MAPK) or extracellular signal-related kinase (ERK1/2) cascade is one of the key signaling pathways that transmits growth signals to the nucleus (Chen et al 1992; Gonzalez et al 1993; Karin and Hunter 1995). Two-thirds of human cancers, including skin, colon, lung, and pancreas; multiple myeloma; and hairy cell leukemia, have aberrations in the ERK1/2 cascade, largely due to activating mutations in signaling intermediates such as EGFR, KRAS, or BRAF (Davies et al 2002; Garnett and Marais 2004; Dhillon et al 2007; Bryant et al 2014) This understanding led to the development of targeted inhibitors against kinase components of the MAPK pathway that could be used for cancer therapy (Roberts and Der 2007; Santarpia et al 2012). Depletion of INTS11 diminishes cellular proliferation in A375 cells rendered resistant to MAPK inhibitors, highlighting a possible avenue to overcome drug resistance by targeting INTS11 in cancer cells

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Conclusion