UC San Diego

Due to the discovery of ribosome heterogeneity, there has opened a huge field of research surrounding the regulatory roles that ribosomal proteins play in cellular functions. Understanding how each one affects cell growth, differentiation, and maturation will build a more robust model of cellular development. This study aimed to analyze the effects of RPL22 and its paralog, RPL22L1, on early cell fate determination in an in vitro embryonic differentiation model. To do this, we generated RPL22 and RPL22L1 knockdown lines using the CRISPRi platform in iPSCs. These cell lines were then used for embryoid body (EB) formation in order to study the effects of RPL22 and RPL22L1 in the development of the three germ layers by accessing the expression of cell lineage markers during differentiation. We found that RPL22L1 can compensate for the loss of RPL22, but the opposite is not true, and that RPL22L1 is essential for cell survival and EB differentiation. Our results also suggest that RPL22 may contribute to definitive endoderm development, affect differentiation of mesoderm-derived cell types, and play a positive role in neuron differentiation. Finally, we adapted a new technology, Surveying Targets by APOBEC Mediated Profiling (STAMP), and successfully synthesized APOBEC-RPL22 and APOBEC-RPL22L1 constructs so that we can identify the translational and RNA binding targets of RPL22 and RPL22L1 to gain insight into the molecular mechanisms they may regulate.