Evolutionary insights into primate skeletal gene regulation using a comparative cell culture model

Abstract

The evolution of complex skeletal traits in primates was likely influenced by both genetic and environmental factors. Because skeletal tissues are notoriously challenging to study using functional genomic approaches, they remain poorly characterized even in humans, let alone across multiple species. The challenges involved in obtaining functional genomic data from the skeleton, combined with the difficulty of obtaining such tissues from nonhuman apes, motivated us to consider an alternative in vitro system with which to comparatively study gene regulation in skeletal cell types. Specifically, we differentiated six human (Homo sapiens) and six chimpanzee (Pan troglodytes) induced pluripotent stem cell lines (iPSCs) into mesenchymal stem cells (MSCs) and subsequently into osteogenic cells (bone cells). We validated differentiation using standard methods and collected single-cell RNA sequencing data from over 100,000 cells across multiple samples and replicates at each stage of differentiation. While most genes that we examined display conserved patterns of expression across species, hundreds of genes are differentially expressed (DE) between humans and chimpanzees within and across stages of osteogenic differentiation. Some of these interspecific DE genes show functional enrichments relevant in skeletal tissue trait development. Moreover, topic modeling indicates that interspecific gene programs become more pronounced as cells mature. Overall, we propose that this in vitro model can be used to identify interspecific regulatory differences that may have contributed to skeletal trait differences between species.

Characterizing gene expression in an in vitro biomechanical strain model of joint health

“…However, prior work demonstrates that gene expression patterns between separate iPSC-MSC lines differentiated independently from the same individual are highly consistent, and this consistency persists across species and across variations of MSC differentiation media. 78 Therefore, we anticipate that iPSC-MSC differentiation does not contribute significantly to gene expression variation on the individual level.…”

GoM DE: interpreting structure in sequence count data with differential expression analysis allowing for grades of membership

“…Another class of dimensionality reduction approaches that have been used to identify structure from singlecell data are what are sometimes called parts-based representations-these approaches include non-negative matrix factorization (NMF) [30][31][32][33][34][35][36][37][38][39][40][41][42][43][44] and topic modeling [45][46][47][48][49][50][51][52][53][54][55][56], which also have formal connections [48,57,58]. Parts-based representations share some of the features of both a clustering and a dimensionality reduction: on the one hand, they learn a lower dimensional representation of the cells; on the other hand, the individual dimensions (the "parts") of the reduced representation can identify discrete clusters or discrete subpopulations [59,60].…”

Architecture and evolutionary conservation ofXenopus tropicalisosteoblast-specific regulatory regions shed light on bone diseases and early skeletal evolution

“…In Xt, osteogenesis follows a conserved pattern with mammals, as mesenchymal precursors progressively differentiate into osteoblasts, the bone producing cells [29,30]. However, amphibians have never been employed to examine the osteoblastic regulatory landscape, which has so far only been studied in great depth in mammals [31][32][33][34][35]. In mouse, a combination of genome wide approaches, knock-out animals, and reporter experiments, has revealed how signalling pathways and transcription factors cooperate at enhancers to drive ossification in a well-controlled spatio-temporal fashion [35][36][37][38][39][40][41][42][43][44][45][46].…”