BME Student Seminar (Zoom): Navied Akhtar and Andrew Phan

Navied Akhtar: Investigating the Role of Cell Cycle Replication as a Source of Heterogeneity in Human Embryonic Stem Cell Populations

Abstract: Human embryonic stem cells (ESCs) have the ability to differentiate into the multitude of specialized cells in the human body. These ESCs are phenotypically similar, but there exists molecular heterogeneity within stem cell populations. Although advances in single-cell technologies have exposed the presence of this heterogeneity at the transcriptional level, a source for this molecular variability remains elusive. The epigenome serves as a regulator of transcription, and new technologies developed in our lab (Repli-Bisulfite Sequencing) identified S-phase replication as a potential window for epigenetic changes to affect cell-fate decision making. We propose that ESCs in a population exist in poised cell states, with a subpopulation of cells primed for self-renewal while other subpopulation exists in states primed for  differentiation. However, we lack the sophisticated technologies necessary to identify epigenetic variability in S-phase and to correlate this variability to changes in transcription and cell-fate changes. As such, development and repurposing of available tools is of paramount importance to elucidate replication-associated epigenetic heterogeneity in human ESCs. Particularly promising tools involve nucleotide analogs that can be integrated into the DNA of cells in S-phase, allowing for cell-cycle separation of ESCs. Additionally, long-read sequencing can provide insights that are otherwise unattainable using conventional sequencing methods. Tools such as these will provide new insights into a novel source of epigenetic heterogeneity.

Andrew Phan: Adhesome Gene Expression Regulates Stemness Acquisition and Cell-Environment Signaling

Abstract: Due to the mechanosensitive nature of stem cells, intense research has focused on the role of the cell microenvironment in directing cell fate decisions and stemness acquisition. Biophysical cues such as micropatterned grooves, decreasing substrate stiffness, lateral confinement and increasing substrate dimensionality can enhance somatic reprogramming. Similarly, many of these findings extend to cancer stem cell (CSC) generation, which suggests the influence of biophysical cues on stemness acquisition is conserved even in oncogenic cells. In summary, several studies suggest that altering states of cell adhesion, typically by weakening adhesion, can enhance the reprogramming of differentiated cells to stem-like states.

Despite this growing evidence, a systematic study on the role of cell adhesion in stemness acquisition has not been performed. Therefore, we investigated the effect of a collection of cell-cell and cell-ECM associated genes, known as the adhesome, on reprogramming of somatic and oncogenic cells. A custom RNAi screen targeting 103 adhesome genes was developed and demonstrated adhesome expression is largely refractory to somatic reprogramming and revealed novel top hits not previously associated with stemness acquisition. To further understand the mechanistic changes caused by modulating cell adhesion gene expression, scRNA-seq was performed on reprogramming fibroblasts and A549 cells with adhesome gene knockdowns. Adhesome expression knockdown in both reprogramming models increased stemness acquisition, created different cell trajectories to stemness, and, using CellChat inferred communication patterns, utilized similar cell-environment signaling pathways during stemness acquisition. Together, our findings support the adhesome as a novel target during somatic and oncogenic dedifferentiation for more effective regenerative therapies and CSC-targeting treatments.