ENCODE tiling array analysis identifies differentially expressed annotated and novel 5' capped RNAs in hepatitis C...

Area of Science:

• Genomics
• Molecular Biology
• Virology

Background:

• Hepatitis C virus (HCV) infection studies using microarrays offer insights into host responses.
• Recent transcriptome data reveal pervasive transcription and short/absent poly(A) tails in RNA transcripts.
• Investigating novel transcripts is crucial for understanding HCV pathogenesis.

Purpose of the Study:

• To identify novel annotated and unannotated genes differentially expressed in the liver during HCV infection.
• To evaluate the utility of 5' cap-purified RNA and ENCODE tiling arrays for transcript discovery.
• To characterize the host response in HCV infection beyond polyadenylated transcripts.

Main Methods:

• Utilized ENCODE tiling arrays to analyze both 5'-capped and poly(A)+ RNA populations.
• Employed affinity purification of Pol II RNAs via their 5' m⁷GpppN cap.
• Performed bioinformatic analysis, PCR, RACE, and qPCR for transcript verification and quantification.

Main Results:

• Identified 64 upregulated and 31 downregulated annotated genes in HCV cirrhotic liver compared to controls.
• Discovered that 42% of upregulated and 50% of downregulated genes were exclusively identified by analyzing 5' capped RNA.
• Revealed upregulated Pol II transcriptionally active regions in unannotated genomic areas, verified by PCR and RACE.
• Confirmed upregulation of novel unannotated transcripts and specific genes (IRF1, TRIM22, MET) in HCV-infected liver biopsies.

Conclusions:

• Analyzing 5' capped RNA alongside ENCODE tiling arrays enhances gene expression information and identifies novel transcripts in HCV infection.
• This approach reveals previously undescribed Pol II transcripts in HCV-infected liver.
• The method is valuable for defining differentially regulated Pol II transcripts in disease states and studying RNA regulation.