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The DNA damage response activates HPV16 late gene expression at the level of RNA processing
Kersti Nilsson1, Chengjun Wu1, Naoko Kajitani1
1Department of Laboratory Medicine, Lund University, BMC-B13, 221 84 Lund, Sweden.
Abstract:
We show that the alkylating cancer drug melphalan activated the DNA damage response and induced human papillomavirus type 16 (HPV16) late gene expression in an ATM- and Chk1/2-dependent manner. Activation of HPV16 late gene expression included inhibition of the HPV16 early polyadenylation signal that resulted in read-through into the late region of HPV16. This was followed by activation of the exclusively late, HPV16 splice sites SD3632 and SA5639 and production of spliced late L1 mRNAs. Altered HPV16 mRNA processing was paralleled by increased association of phosphorylated BRCA1, BARD1, BCLAF1 and TRAP150 with HPV16 DNA, and increased association of RNA processing factors U2AF65 and hnRNP C with HPV16 mRNAs. These RNA processing factors inhibited HPV16 early polyadenylation and enhanced HPV16 late mRNA splicing, thereby activating HPV16 late gene expression.
Insights
Melphalan, a cancer drug, triggers DNA damage response and activates human papillomavirus type 16 (HPV16) late gene expression. This involves altered mRNA processing, leading to HPV16 L1 mRNA production.
Area of Science:
- Molecular Biology
- Virology
- Oncology
Background:
- Human papillomavirus type 16 (HPV16) is a major cause of cervical cancer.
- Understanding HPV16 gene regulation is crucial for developing targeted therapies.
Purpose of the Study:
- To investigate the effect of the alkylating cancer drug melphalan on HPV16 gene expression.
- To elucidate the molecular mechanisms underlying melphalan-induced HPV16 late gene activation.
Main Methods:
- Treatment of cells with melphalan.
- Analysis of HPV16 gene expression and mRNA processing.
- Assessment of protein-DNA and protein-RNA interactions.
- Investigating the roles of ATM and Chk1/2 kinases.
Main Results:
- Melphalan activated the DNA damage response and HPV16 late gene expression in an ATM- and Chk1/2-dependent manner.
- Melphalan inhibited HPV16 early polyadenylation, leading to read-through transcription into the late region.
- Activation of late splice sites (SD3632, SA5639) and production of spliced L1 mRNAs were observed.
- Increased association of phosphorylated BRCA1, BARD1, BCLAF1, TRAP150 with HPV16 DNA and U2AF65, hnRNP C with HPV16 mRNAs occurred.
- These factors inhibited early polyadenylation and enhanced late splicing.
Conclusions:
- Melphalan activates HPV16 late gene expression through modulation of mRNA processing.
- The DNA damage response pathway plays a key role in regulating HPV16 late gene expression.
- Targeting these RNA processing mechanisms could offer novel therapeutic strategies for HPV16-associated cancers.
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