MicroRNA-mediated regulation of stem cell fate decisions during reprogramming and viral infection
Zika virus (ZIKV) is an emerging arbovirus linked to an increased incidence of microcephaly. To study the potential link between ZIKV and microcephaly, stem cell-based models must be utilized to understand the effect of ZIKV infection. The objectives of this dissertation are to analyze the somatic cell reprogramming to enhance iPSC production, generate human cerebral organoids, and utilize these models to study ZIKV-mediated neurodegeneration in vitro.
To study the stochastic nature of somatic cell reprogramming and enhance reprogramming yield, microRNAs were profiled in mouse embryonic fibroblasts (MEFs) during the early stage of cell fate decisions. miR-135b was highly upregulated and repressed expression of extracellular matrix genes including Wisp1 and Igfbp5. These data reveal a novel link between microRNA-mediated regulation of ECM formation and somatic cell reprogramming.
To investigate the link between ZIKV and microcephaly, human embryonic stem cell-derived cerebral organoids were generated and characterized to recapitulate first trimester fetal brain development. ZIKV infected organoids revealed preferential infection of neural stem cells, attenuated growth and activation of innate immune receptor Toll-Like-Receptor 3 (TLR3). Pathway analysis of differentially expressed genes during TLR3 activation highlighted 41 genes also related to neuronal development.
Lastly, meta-analyses and regulatory interaction networks integrating miRNA and mRNA expression profiling data were used to study the role of miRNA-mediated repression during ZIKV infection. These analyses identified miRNA-mediated repression of cell cycle, metabolism, stem cell maintenance and neurogenesis related genes. Moreover, miR-218 was upregulated during ZIKV infection and directly represses a gene network governing stem cell maintenance.