Putative oncogene Brachyury ( T ) is essential to specify cell fate but dispensable for notochord progenitor proliferation and EMT

Abstract

The transcription factor Brachyury (T) gene is expressed throughout primary mesoderm (primitive streak and notochord) during early embryonic development and has been strongly implicated in the genesis of chordoma, a sarcoma of notochord cell origin. Additionally, T expression has been found in and proposed to play a role in promoting epithelial-mesenchymal transition (EMT) in various other types of human tumors. However, the role of T in normal mammalian notochord development and function is still not well-understood. We have generated an inducible knockdown model to efficiently and selectively deplete T from notochord in mouse embryos. In combination with genetic lineage tracing, we show that T function is essential for maintaining notochord cell fate and function. Progenitors adopt predominantly a neural fate in the absence of T, consistent with an origin from a common chordoneural progenitor. However, T function is dispensable for progenitor cell survival, proliferation, and EMT, which has implications for the therapeutic targeting of T in chordoma and other cancers.notochord | Brachyury | cell fate | EMT T he notochord, the defining characteristic of chordates, serves as a signaling center for dorsoventral patterning of both the adjacent neural tube and the somites. Sonic hedgehog (Shh) expressed from the notochord specifies ventral neural fates in spinal cord and ventral somitic fate to form sclerotome that gives rise to the vertebrae of the axial skeleton (1, 2). Genetic lineage tracing in the mouse has revealed that later, during the process of intervertebral disk formation, the notochord fragments into discrete segments and differentiates to form the nucleus pulposus of the intervertebral disks (3, 4). It has been proposed that this process generates the notochord "remnants" that can persist within vertebral bodies in adults (5), which are thought to give rise to chordomas, a rare sarcoma of notochord cell origin. A major advance in understanding the pathogenesis of chordoma has been the discovery that Brachyury (T) gene, which is highly expressed in these tumors, is duplicated in certain familial chordomas (6). T expression, which is pathognomonic for diagnosis of these sarcomas (7), has consequently become a major focus for therapeutic targeting in treatment of these cancers (7,8).T is the founding member of the T-box gene family of transcription factors (9, 10). During embryonic development, T is expressed in the notochord and primitive streak, and is essential for trunk/tail primary mesoderm formation and migration from the primitive streak, which drives axis elongation (11,12). Loss of T function in mouse embryos results in body axis truncation caudal to the forelimb, showing that T is dispensable for anterior-most mesoderm formation (rostral to somite 7). The embryonic role in epithelial-mesenchymal transition (EMT) of cells migrating from primitive streak during mesoderm formation has piqued interest that T, which is variably expressed in a number of common cancers in addition to chordoma (8), may...

Visceral endoderm and the primitive streak interact to build the fetal-placental interface of the mouse gastrula

“…However, in future experiments, it would be informative to carry out AX grafts in the Ptch1 reporter strain, and investigate levels of Ptch1 in these after culture to find out whether it is high in donor AX that is no longer associated with the streak. Finally, while many studies have implicated T in the EMT in various cell lines (e.g., (Hamilton et al, 2012), results of studies in vivo favor an essential role for T in specification of cell fate, and not the EMT or cell proliferation (Zhu et al, 2016). Our data accord with a general role for T in stem cell identity within the AX, where it is found with other stem cell factors.…”

Nonviral Transfection With Brachyury Reprograms Human Intervertebral Disc Cells to a Pro‐Anabolic Anti‐Catabolic/Inflammatory Phenotype: A Proof of Concept Study

“…The transcription factor Brachyury (T) is expressed in the developing notochord and is associated with maintaining a healthy immature NP phenotype and essential for normal spine formation. 28,29 Transfection of transcription factors have widely been established and documented for reprogramming studies, specifically turning somatic cells into induced pluripotent stem cells (iPSCs). 30 Thus, cellular reprogramming in this study refers to the transformation or reversal of a diseased cell back to its healthy (non-degenerate) phenotype.…”

The transcription factors regulating intervertebral disc development

“… 14 Recent studies have shown that T‐Brachyury is expressed and functions in the notochord at E10.5. 15 Shh Cre inducible T‐Brachyury shRNA expressing T‐Brachyury knockdown mice show a phenotype with a costal level of vertebral malformation or loss because of the loss of the notochordal cells, which indicates that T‐Brachyury is essential for normal spine formation 15 . Even after the completion of the NP formation, T‐Brachyury is still strongly expressed in the NP cells, particularly in the notochordal cells.…”

“…T-Brachyury is a transcription factor that plays a critical role in embryonic mesoderm development, particularly for the formation of the notochord ( Figure 2A) (Table 1). 14…”