Regulation of microRNA biogenesis in stem cells. This project aims to understand the mechanisms by which microRNA (miRNA) processing is regulated in embryonic stem cells (ESC). ESCs have the remarkable capacity to differentiate into all cell types and are of potential therapeutic value for numerous degenerative diseases. However, the molecular foundations of ESC biology remain poorly defined. Functioning as negative regulators of gene expression, miRNAs are critical for diverse cell fate decisions during normal development; miRNA dysregulation is linked with several diseases including cancer. It is increasingly well appreciated that posttranscriptional mechanisms play an important role controlling miRNA expression. Lin28a selectively represses let-7 miRNA biogenesis in ESCs. This pathway helps maintain an undifferentiated cell state and is often reactivated in cancer. Although much progress has been made elucidating the mechanism of the Lin28-mediated control of let-7 expression, several outstanding questions remain.
Aim 1 will explore the regulation of the Lin28/let-7 pathway. A combination of biochemical and cell-based assays will explore the molecular determinants for let-7 regulation and will use state-of-the-art genetic engineering to explore the impact of a Lin28/let-7 developmental switch mechanism in ESCs. A variety of approaches will be used to investigate the mechanism by which Lin28b represses let-7 expression and to explore the possible TUTase-independence of this pathway. In vitro binding and cell-based assays will be employed to investigate the regulation of the Lin28/let-7 pathway by an identified lncRNA.
Aim 2 is focused on understanding the posttranscriptional mechanisms controlling miR-17~92 biogenesis. The precise control of miR-17~92 is essential for normal development and overexpression of certain miRNAs from this cluster is oncogenic. New data implicate a novel biogenesis step upstream of Microprocessor that controls miR-17~92 expression in ESCs. A variety of biochemical and cellular assays will be used to test this and will reveal the RNA features and regulatory factors that control miR-17~92 expression in ESCs. Illuminating the molecular details of the Lin28/let-7 pathway and miR-17~92 regulation promises to impact broad areas of biology and disease, reveal novel stem cell regulatory factors, and offer strategies for therapeutic targeting to treat cancer, obesity, and diabetes.

Public Health Relevance

Stem cells hold great promise for the development of new approaches to combat disease. However, the molecular basis for stem cell self-renewal and differentiation remains incompletely understood. The proposed work will provide novel insight into embryonic stem cell gene regulation, and may lead to new therapies to manipulate microRNAs. These studies are relevant to the treatment of cancer, diabetes, developmental disorders and numerous degenerative diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM086386-07
Application #
9102102
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Bender, Michael T
Project Start
2009-07-01
Project End
2019-06-30
Budget Start
2016-07-01
Budget End
2017-06-30
Support Year
7
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Children's Hospital Boston
Department
Type
DUNS #
076593722
City
Boston
State
MA
Country
United States
Zip Code
Lin, Shuibin; Liu, Qi; Lelyveld, Victor S et al. (2018) Mettl1/Wdr4-Mediated m7G tRNA Methylome Is Required for Normal mRNA Translation and Embryonic Stem Cell Self-Renewal and Differentiation. Mol Cell 71:244-255.e5
Pirouz, Mehdi; Ebrahimi, Aref G; Gregory, Richard I (2018) Unraveling 3'-end RNA uridylation at nucleotide resolution. Methods :
Du, Peng; Pirouz, Mehdi; Choi, Jiho et al. (2018) An Intermediate Pluripotent State Controlled by MicroRNAs Is Required for the Naive-to-Primed Stem Cell Transition. Cell Stem Cell 22:851-864.e5
Choe, Junho; Lin, Shuibin; Zhang, Wencai et al. (2018) mRNA circularization by METTL3-eIF3h enhances translation and promotes oncogenesis. Nature 561:556-560
Tsanov, Kaloyan M; Pearson, Daniel S; Wu, Zhaoting et al. (2017) LIN28 phosphorylation by MAPK/ERK couples signalling to the post-transcriptional control of pluripotency. Nat Cell Biol 19:60-67
Lin, Shuibin; Choe, Junho; Du, Peng et al. (2016) The m(6)A Methyltransferase METTL3 Promotes Translation in Human Cancer Cells. Mol Cell 62:335-345
Pirouz, Mehdi; Du, Peng; Munafò, Marzia et al. (2016) Dis3l2-Mediated Decay Is a Quality Control Pathway for Noncoding RNAs. Cell Rep 16:1861-73
Du, Peng; Wang, Longfei; Sliz, Piotr et al. (2015) A Biogenesis Step Upstream of Microprocessor Controls miR-17?92 Expression. Cell 162:885-99
Triboulet, Robinson; Pirouz, Mehdi; Gregory, Richard I (2015) A Single Let-7 MicroRNA Bypasses LIN28-Mediated Repression. Cell Rep 13:260-6
Thornton, James E; Du, Peng; Jing, Lili et al. (2014) Selective microRNA uridylation by Zcchc6 (TUT7) and Zcchc11 (TUT4). Nucleic Acids Res 42:11777-91

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