Coordinated regulation of synthesis and stability of RNA during the acute TNF-induced proinflammatory response.
Steady-state gene expression is a coordination of synthesis and decay of RNA through epigenetic regulation, transcription factors, micro RNAs (miRNAs), and RNA-binding proteins. Here, we present bromouride labeling and sequencing (Bru-Seq) and bromouridine pulse-chase and sequencing (BruChase-Seq) to assess genome-wide changes to RNA synthesis and stability in human fibroblasts at homeostasis and after exposure to the proinflammatory tumor necrosis factor (TNF). The inflammatory response in human cells involves rapid and dramatic changes in gene expression, and the Bru-Seq and BruChase-Seq techniques revealed a coordinated and complex regulation of gene expression both at the transcriptional and posttranscriptional levels. The combinatory analysis of both RNA synthesis and stability using Bru-Seq and BruChase-Seq allows for a much deeper understanding of mechanisms of gene regulation than afforded by the analysis of steady-state total RNA and should be useful in many biological settings.
Department of Radiation Oncology, University of Michigan Comprehensive Cancer Center and Translational Oncology Program, University of Michigan, Ann Arbor, MI 48109, USA.
Michelle T
MT
Paulsen
Artur
A
Veloso
Jayendra
J
Prasad
Karan
K
Bedi
Emily A
EA
Ljungman
Ya-Chun
YC
Tsan
Ching-Wei
CW
Chang
Brendan
B
Tarrier
Joseph G
JG
Washburn
Robert
R
Lyons
Daniel R
DR
Robinson
Chandan
C
Kumar-Sinha
Thomas E
TE
Wilson
Mats
M
Ljungman
D001973:Bromodeoxyuridine / Q000378:metabolism; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D015894:Genome, Human; D006801:Humans; D007249:Inflammation / Q000209:etiology / Q000235:genetics* / Q000378:metabolism*; D007438:Introns; D012313:RNA / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D020871:RNA Stability; D000077278:RNA, Mitochondrial; D012335:RNA, Ribosomal / Q000235:genetics / Q000378:metabolism; D059467:Transcriptome; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000077278:RNA, Mitochondrial; D012335:RNA, Ribosomal; D014409:Tumor Necrosis Factor-alpha; D012313:RNA; D001973:Bromodeoxyuridine