Serine Threonine Kinase Receptor-Associated Protein Deficiency Impairs Mouse Embryonic Stem Cells Lineage Commitment Through CYP26A1-Mediated Retinoic Acid Homeostasis

Abstract

Retinoic acid (RA) signaling is essential for the differentiation of embryonic stem cells (ESCs) and vertebrate development. RA biosynthesis and metabolism are controlled by a series of enzymes, but the molecular regulators of these enzymes remain largely obscure. In this study, we investigated the functional role of the WD-domain protein STRAP (serine threonine kinase receptor-associated protein) in the pluripotency and lineage commitment of murine ESCs. We generated Strap knockout (KO) mouse ESCs and subjected them to spontaneous differentiation. We observed that, despite the unchanged characteristics of ESCs, Strap KO ESCs exhibited defects for lineage differentiation. Signature gene expression analyses revealed that Strap deletion attenuated intracellular RA signaling in embryoid bodies (EBs), and exogenous RA significantly rescued this deficiency. Moreover, loss of Strap selectively induced Cyp26A1 expression in mouse EBs, suggesting a potential role of STRAP in RA signaling. Mechanistically, we identified putative Krüppel-like factor 9 (KLF9) binding motifs to be critical in the enhancement of non-canonical RA-induced transactivation of Cyp26A1. Increased KLF9 expression in the absence of STRAP is partially responsible for Cyp26A1 induction. Interestingly, STRAP knockdown in Xenopus embryos influenced anterior-posterior neural patterning and impaired the body axis and eye development during early Xenopus embryogenesis. Taken together, our study reveals an intrinsic role for STRAP in the regulation of RA signaling and provides new molecular insights for ESC fate determination. Stem Cells 2018;36:1368-1379.

STRAP regulates alternative splicing fidelity during lineage commitment of mouse embryonic stem cells

“…In sum, these studies define a genome-wide AS network that STRAP regulates in a coordinated manner. We reveal the position-dependent and context-dependent effects of STRAP on splicing regulation during neuroectodermal lineage commitment, which adds another layer of complexity to the cross- 18 . The medium was changed daily.…”

“…In our previous study, we found that STRAP affects neural patterning in early Xenopus embryogenesis 18 . Given that STRAP is a conserved WD-40 domain-containing protein across species, we hypothesized that it has a functional role in AS regulation in Xenopus.…”

“…Since the AS program tightly controls the posttranscriptional genes necessary for cell differentiation and development, we reason that these defects are caused, in part, by AS events upon Strap KO. The following evidence supports this concept: (i) Substantial AS patterns, instead of limited transcriptional regulation, are affected upon Strap KO in lineagecommitted murine ESCs; (ii) The involvement of STRAP for the assembly of the 17S U2 snRNP complex suggests its role in AS modification; (iii) STRAP contributes to the selection of important neuronal exons during EB differentiation, as exemplified by Nnat and Mark3; and (iv) Strap-MO in Xenopus embryos influences neural patterning and impaires the body axis 18 .…”

“…STRAP has a regulatory role in mouse early organogenesis. We recently employed zinc finger nuclease (ZFN) technology to generate Strap deficient mice 18 . Although all Strap +/male and female mice did not display any phenotypic abnormalities, their intercrosses yielded no homozygous offspring.…”

“…We next characterized transcripts and splicing variants with genome-wide profiling. Only 154 genes were altered in transcriptome profiling of Strap-KO ESCs relative to that of ESC WT cells (Supplementary Data 4), suggesting that STRAP is dispensable for mouse ESCs viability and identity 18 . Loss of STRAP in 9-day-old EBs resulted in 146 genes upregulated and 261 genes downregulated (Supplementary Data 4).…”