1. Project: Cellular microRNA profilings during the infections of the hepatitis C virus and the influenza A virus Yu Li Postdoc Fellow Katze Lab

2. Projects 1.Cellular microRNA profilings in the Hepatitis C virus infected human liver tissues HCV positive liver biopsies from transplant patients HCV negative liver biopsies from transplant patients 2.Cellular microRNA profilings in the Influenza virus infected human cell line and mouse lung tissues. Viruses: H1N1 (human flu): 1918 (High path) and K173 (Low path) H5N1 (Avian flu): HK483 (High path) and HK486 (Low path) Mock virus In vitro study- Human epithelial cells A549 H1N1, H5N1 influenza flu viruses and mock virus infections In vivo study- Mice lungs from a previously published study H1N1 only- 1918(High path), Texas (Low path) and Mock virus

3. MicroRNA MicroRNA is a group of noncoding small RNA molecules with ~22 nt in length. –847 are registered human microRNA in Sanger database V10 as of Jun 2008 MicroRNAs play a central role in regulating cellular gene expression at posttranscriptional levels. –On average, a microRNA regulates the expression of over 200 cellular genes or microRNA targets –A microRNA targets can be regulated by multiple microRNAs

4. Repression of Gene Expression by MicroRNA Seed region from 2-8 Decreased protein abundance

5. Experimental Design Flu-associated mRNA expression profile (Gene expression arrays) Flu-associated microRNA expression profile (MicroRNA arrays) microRNA target prediction Inverse correlation of flu-associated microRNA and gene expressions A549 cells infected with high-path virus and low-path influenza virus (total RNA)

6. MicroRNA expression profilings: –22 HCV positive liver biopsies (20 finished & 2 will be finished this week) –6 HCV negative liver biopsies (will be finished this week) –All microRNAs arrays are single channel arrays Gene expression profilings: –22 HCV positive liver biopsies. (Finished by NIDA) –All gene expression arrays are duel channel arrays (pooled mock as the reference) MicroRNA target prediction in the human genome –Xinxia used “perfect-match” for predicting microRNA targets that can be repressed by a microRNA on the mRNA level. (Finished) Current status- The HCV-microRNA project

7. MicroRNA expression profilings: –H1N1 and H5N1 infected A549 cells (the majority are finished, 2 more slides to finish ) –H1N1 infected mouse lungs (will be finished in 2 weeks) Gene expression profilings: –H1N1, H5N1 and mock infected A549 cells (finished) –H1N1 and mock infected mouse lung (Kash paper 2006) –We are not only focused on the difference between high path and low path flu virus infections but also focused on the difference between flu and mock. MicroRNA target prediction in the human and mouse genomes –Xinxia used “perfect-match” for predicting microRNA targets that can be repressed by a microRNA on the mRNA level. (Finished) Current status- The Flu-microRNA project

8. Next steps Establish the correlation between the microRNA expression changes and the gene expression changes: –2 independent analyses: From the end of microRNA profilings: –Make the prediction of gene expression changes from microRNA expression changes From the end of gene expression profilings: –Make the prediction of microRNA expression changes from gene expression changes –Both analyses will help to find the pathways that associated with HCV or influenza infections and microRNA expression changes. –The common pathways found by both analyses have higher probability to be real and would be use to make biological stories. Taqman validation of the both microRNA and gene expression changes if the material is still available: Draft a manuscript: