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Capturing the dynamic nascent transcriptome during acute cellular responses: The serum response.

Kirkconnell KS, Paulsen MT, Magnuson B, Bedi K, Ljungman M - Biol Open (2016)

Bottom Line: Surprisingly, transcription of important DNA damage response genes and histone genes were rapidly repressed.We also show that RNA polymerase II accelerates as it transcribes large genes and this was independent of whether the gene was induced or not.These results provide a unique genome-wide depiction of dynamic patterns of transcription of serum response genes and demonstrate the utility of Bru-seq to comprehensively capture rapid and dynamic changes of the nascent transcriptome.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, University of Michigan Comprehensive Cancer Center, and Translational Oncology Program, University of Michigan, Ann Arbor, MI 48109, USA Department of Human Genetics, University of Michigan Medical School, Ann Arbor, MI 48109, USA.

No MeSH data available.


Related in: MedlinePlus

Immediate serum-response genes. Bru-seq traces for TPM1 (A), FERMT2 (B), APCDD1 (C) and RUNX2 (D) during starved conditions and after different periods of serum stimulation. Annotated genes are shown at the top in either red or green. The positive y-axis represents plus-strand signal of transcription moving from left to right and the negative y-axis represents minus-strand signal of transcription moving from right to left. Transcription induction is indicated by a green arrow and transcription repression is indicated by a red T. The graphs at the bottom depict the log2-fold change values calculated within the first 30 kb of the genes for each labeling period.
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BIO019323F2: Immediate serum-response genes. Bru-seq traces for TPM1 (A), FERMT2 (B), APCDD1 (C) and RUNX2 (D) during starved conditions and after different periods of serum stimulation. Annotated genes are shown at the top in either red or green. The positive y-axis represents plus-strand signal of transcription moving from left to right and the negative y-axis represents minus-strand signal of transcription moving from right to left. Transcription induction is indicated by a green arrow and transcription repression is indicated by a red T. The graphs at the bottom depict the log2-fold change values calculated within the first 30 kb of the genes for each labeling period.

Mentions: We observed that some genes, such as TPM1 encoding an actin skeleton protein, were immediately upregulated following serum addition, visible through an increase in transcription reads across the entire gene (Fig. 2A). This increase was observed during each labeling period. The FERMT2 gene encoding a scaffolding protein behaves in a very similar manner, but because this gene is considerably longer we only observe transcription reads at the 5′ end of the gene during the first labeling period (Fig. 2B); however, during the subsequent labeling period (30-60 min), the transcription wave had reached the 3′ end of the gene. One advantage of nascent RNA Bru-seq over traditional RNA-seq, which measures steady-state RNA, is that it can capture rapid inhibition of transcription very efficiently since it does not rely on the degradation of pre-existing RNA for detection of inhibition. For example, the APCDD1 gene coding for a protein that inhibits WNT signaling was actively transcribed during serum starvation but was rapidly repressed upon serum addition (Fig. 2C). RUNX2, encoding a transcription factor, was also repressed by serum and due to its considerable length (>100 kb) transcription at the end of the gene was unaffected until 60-90 min after serum addition (Fig. 2D). Thus, even though genes can be induced or repressed immediately after serum addition, there is a time delay before the generation or loss of full-length products that is proportional to their gene lengths.Fig. 2.


Capturing the dynamic nascent transcriptome during acute cellular responses: The serum response.

Kirkconnell KS, Paulsen MT, Magnuson B, Bedi K, Ljungman M - Biol Open (2016)

Immediate serum-response genes. Bru-seq traces for TPM1 (A), FERMT2 (B), APCDD1 (C) and RUNX2 (D) during starved conditions and after different periods of serum stimulation. Annotated genes are shown at the top in either red or green. The positive y-axis represents plus-strand signal of transcription moving from left to right and the negative y-axis represents minus-strand signal of transcription moving from right to left. Transcription induction is indicated by a green arrow and transcription repression is indicated by a red T. The graphs at the bottom depict the log2-fold change values calculated within the first 30 kb of the genes for each labeling period.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4920201&req=5

BIO019323F2: Immediate serum-response genes. Bru-seq traces for TPM1 (A), FERMT2 (B), APCDD1 (C) and RUNX2 (D) during starved conditions and after different periods of serum stimulation. Annotated genes are shown at the top in either red or green. The positive y-axis represents plus-strand signal of transcription moving from left to right and the negative y-axis represents minus-strand signal of transcription moving from right to left. Transcription induction is indicated by a green arrow and transcription repression is indicated by a red T. The graphs at the bottom depict the log2-fold change values calculated within the first 30 kb of the genes for each labeling period.
Mentions: We observed that some genes, such as TPM1 encoding an actin skeleton protein, were immediately upregulated following serum addition, visible through an increase in transcription reads across the entire gene (Fig. 2A). This increase was observed during each labeling period. The FERMT2 gene encoding a scaffolding protein behaves in a very similar manner, but because this gene is considerably longer we only observe transcription reads at the 5′ end of the gene during the first labeling period (Fig. 2B); however, during the subsequent labeling period (30-60 min), the transcription wave had reached the 3′ end of the gene. One advantage of nascent RNA Bru-seq over traditional RNA-seq, which measures steady-state RNA, is that it can capture rapid inhibition of transcription very efficiently since it does not rely on the degradation of pre-existing RNA for detection of inhibition. For example, the APCDD1 gene coding for a protein that inhibits WNT signaling was actively transcribed during serum starvation but was rapidly repressed upon serum addition (Fig. 2C). RUNX2, encoding a transcription factor, was also repressed by serum and due to its considerable length (>100 kb) transcription at the end of the gene was unaffected until 60-90 min after serum addition (Fig. 2D). Thus, even though genes can be induced or repressed immediately after serum addition, there is a time delay before the generation or loss of full-length products that is proportional to their gene lengths.Fig. 2.

Bottom Line: Surprisingly, transcription of important DNA damage response genes and histone genes were rapidly repressed.We also show that RNA polymerase II accelerates as it transcribes large genes and this was independent of whether the gene was induced or not.These results provide a unique genome-wide depiction of dynamic patterns of transcription of serum response genes and demonstrate the utility of Bru-seq to comprehensively capture rapid and dynamic changes of the nascent transcriptome.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, University of Michigan Comprehensive Cancer Center, and Translational Oncology Program, University of Michigan, Ann Arbor, MI 48109, USA Department of Human Genetics, University of Michigan Medical School, Ann Arbor, MI 48109, USA.

No MeSH data available.


Related in: MedlinePlus