Dissection of SINEUP ncRNAs, which have a potential to become next hot molecular biotechnology tool.

2017 Fiscal Year Annual Research Report

• Project/Area Number: 17F17775
• Research Institution: Institute of Physical and Chemical Research
• Principal Investigator: Carninci Piero 国立研究開発法人理化学研究所, ライフサイエンス技術基盤研究センター, 副センター長 (10333296)
• Co-Investigator(Kenkyū-buntansha): RAHMAN MOHAMMED 国立研究開発法人理化学研究所, ライフサイエンス技術基盤研究センター, 外国人特別研究員
• Project Period (FY): 2017-11-10 – 2020-03-31
• Keywords: SINEUP / EEA elements / SINEB2 elements

Outline of Annual Research Achievements

Project 1: iPSCs reprogramming using CRISPRa EEA [EGA (embryo genome activation) enriched Alu-motif (EEA-motif)] motifs and SINEUPs - We have effectively designed SINEUPs against Yamanaka factors (OMKSL). We also checked basal expression level of OMKSL in human fibroblasts and iPSC. Moreover, we are optimizing different transfection protocol in human fibroblast and iPSC. Finally, we are designed and cloning necessary constructs.
Project 2: Dissecting physiological role of lncRNAs with SINEB2 elements - We have finalized the list of SINB2 containing lncRNAs (research plan; project2; a) that we want to dissect in details by analyzing different database analysis. We looked for their expression in different tissues and established cell lines and their involvement in different patho-/physiological functions. Moreover, we started with and got some initial data regarding research plan; project2; a) & b).

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

The main reason I feel like I lagged behind a little because of the major revision that we have made to our research proposal in order to accommodate recent advancement in the flied. But, we picked up the pace recently and hopefully we will match our research goal quickly.

Strategy for Future Research Activity

Future plan (project 1):
1) Transfection optimization (both fibroblast and iPSC), 2) Testing SINEUP against Yamanaka factors (OMKSL), 3) Testing CRISPRa against EEA elements, 4) Establish dedifferentiation of fibroblast into IPSC cells (in house)

Future plan (project 2):
1) Defining relationship between IncRNA/SINEB2 and sense protein coding genes by knockdown and overexpression experiments, 2) Optimizing protocols (RNA FISH, ChRIP-Seq/MS etc) to evaluate the mechanism of action of IncRNA/SINEB2.