Direct targets of pStat5 signalling in erythropoiesis

Kevin R. Gillinder, Hugh Tuckey, Charles C. Bell, Graham W. Magor, Stephen Huang, Melissa D. Ilsley, Andrew C. Perkins

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Erythropoietin (EPO) acts through the dimeric erythropoietin receptor to stimulate proliferation, survival, differentiation and enucleation of erythroid progenitor cells. We undertook two complimentary approaches to find EPO-dependent pStat5 target genes in murine erythroid cells: RNA-seq of newly transcribed (4sU-labelled) RNA, and ChIP-seq for pStat5 30 minutes after EPO stimulation. We found 302 pStat5-occupied sites: ~15% of these reside in promoters while the rest reside within intronic enhancers or intergenic regions, some >100kb from the nearest TSS. The majority of pStat5 peaks contain a central palindromic GAS element, TTCYXRGAA. There was significant enrichment for GATA motifs and CACCC-box motifs within the neighbourhood of pStat5-bound peaks, and GATA1 and/or KLF1 co-occupancy at many sites. Using 4sU-RNA-seq we determined the EPO-induced transcriptome and validated differentially expressed genes using dynamic CAGE data and qRT-PCR. We identified known direct pStat5 target genes such as Bcl2l1, Pim1 and Cish, and many new targets likely to be involved in driving erythroid cell differentiation including those involved in mRNA splicing (Rbm25), epigenetic regulation (Suv420h2), and EpoR turnover (Clint1/EpsinR). Some of these new EpoR-JAK2-pStat5 target genes could be used as biomarkers for monitoring disease activity in polycythaemia vera, and for monitoring responses to JAK inhibitors.

Original languageEnglish
Article numbere0180922
Number of pages20
JournalPLoS ONE
Volume12
Issue number7
DOIs
Publication statusPublished - 1 Jul 2017
Externally publishedYes

Keywords

  • STAT5
  • pSTAT5
  • ChIP-seq
  • RNA-seq
  • 4sU-RNA-seq
  • MPN
  • EPO

Cite this

Gillinder, K. R., Tuckey, H., Bell, C. C., Magor, G. W., Huang, S., Ilsley, M. D., & Perkins, A. C. (2017). Direct targets of pStat5 signalling in erythropoiesis. PLoS ONE, 12(7), [e0180922]. https://doi.org/10.1371/journal.pone.0180922
Gillinder, Kevin R. ; Tuckey, Hugh ; Bell, Charles C. ; Magor, Graham W. ; Huang, Stephen ; Ilsley, Melissa D. ; Perkins, Andrew C. / Direct targets of pStat5 signalling in erythropoiesis. In: PLoS ONE. 2017 ; Vol. 12, No. 7.
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Gillinder, KR, Tuckey, H, Bell, CC, Magor, GW, Huang, S, Ilsley, MD & Perkins, AC 2017, 'Direct targets of pStat5 signalling in erythropoiesis' PLoS ONE, vol. 12, no. 7, e0180922. https://doi.org/10.1371/journal.pone.0180922

Direct targets of pStat5 signalling in erythropoiesis. / Gillinder, Kevin R.; Tuckey, Hugh; Bell, Charles C.; Magor, Graham W.; Huang, Stephen; Ilsley, Melissa D.; Perkins, Andrew C.

In: PLoS ONE, Vol. 12, No. 7, e0180922, 01.07.2017.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Gillinder, Kevin R.

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Gillinder KR, Tuckey H, Bell CC, Magor GW, Huang S, Ilsley MD et al. Direct targets of pStat5 signalling in erythropoiesis. PLoS ONE. 2017 Jul 1;12(7). e0180922. https://doi.org/10.1371/journal.pone.0180922