Circular RNA circPLOD2 regulates pericyte function by targeting the transcription factor KLF4.

Circular RNAs are generated by backsplicing and control cellular signaling and phenotypes. Pericytes stabilize capillary structures and play important roles in the formation and maintenance of blood vessels. Here, we characterize hypoxia-regulated circular RNAs (circRNAs) in human pericytes and show that the circular RNA of procollagen-lysine,2-oxoglutarate 5-dioxygenase-2 (circPLOD2) is induced by hypoxia and regulates pericyte functions. Silencing of circPLOD2 affects pericytes and increases proliferation, migration, and secretion of soluble angiogenic proteins, thereby enhancing endothelial migration and network capability. Transcriptional and epigenomic profiling of circPLOD2-depleted cells reveals widespread changes in gene expression and identifies the transcription factor krüppel-like factor 4 (KLF4) as a key effector of the circPLOD2-mediated changes. KLF4 depletion mimics circPLOD2 silencing, whereas KLF4 overexpression reverses the effects of circPLOD2 depletion on proliferation and endothelial-pericyte interactions. Together, these data reveal an important function of circPLOD2 in controlling pericyte proliferation and capillary formation and show that the circPLOD2-mediated regulation of KLF4 significantly contributes to the transcriptional response to hypoxia.

Locus-Conserved Circular RNA cZNF292 Controls Endothelial Cell Flow Responses.

BACKGROUND: Circular RNAs (circRNAs) are generated by back splicing of mostly mRNAs and are gaining increasing attention as a novel class of regulatory RNAs that control various cellular functions. However, their physiological roles and functional conservation in vivo are rarely addressed, given the inherent challenges of their genetic inactivation. Here, we aimed to identify locus conserved circRNAs in mice and humans, which can be genetically deleted due to retained intronic elements not contained in the mRNA host gene to eventually address functional conservation. METHODS AND RESULTS: Combining published endothelial RNA-sequencing data sets with circRNAs of the circATLAS databank, we identified locus-conserved circRNA retaining intronic elements between mice and humans. CRISPR/Cas9 mediated genetic depletion of the top expressed circRNA cZfp292 resulted in an altered endothelial morphology and aberrant flow alignment in the aorta in vivo. Consistently, depletion of cZNF292 in endothelial cells in vitro abolished laminar flow-induced alterations in cell orientation, paxillin localization and focal adhesion organization. Mechanistically, we identified the protein SDOS (syndesmos) to specifically interact with cZNF292 in endothelial cells by RNA-affinity purification and subsequent mass spectrometry analysis. Silencing of SDOS or its protein binding partner Syndecan-4, or mutation of the SDOS-cZNF292 binding site, prevented laminar flow-induced cytoskeletal reorganization thereby recapitulating cZfp292 knockout phenotypes. CONCLUSIONS: Together, our data reveal a hitherto unknown role of cZNF292/cZfp292 in endothelial flow responses, which influences endothelial shape.

The CUE1 domain of the SNF2-like chromatin remodeler SMARCAD1 mediates its association with KRAB-associated protein 1 (KAP1) and KAP1 target genes.

Chromatin in embryonic stem cells (ESCs) differs markedly from that in somatic cells, with ESCs exhibiting a more open chromatin configuration. Accordingly, ATP-dependent chromatin remodeling complexes are important regulators of ESC homeostasis. Depletion of the remodeler SMARCAD1, an ATPase of the SNF2 family, has been shown to affect stem cell state, but the mechanistic explanation for this effect is unknown. Here, we set out to gain further insights into the function of SMARCAD1 in mouse ESCs. We identified KRAB-associated protein 1 (KAP1) as the stoichiometric binding partner of SMARCAD1 in ESCs. We found that this interaction occurs on chromatin and that SMARCAD1 binds to different classes of KAP1 target genes, including zinc finger protein (ZFP) and imprinted genes. We also found that the RING B-box coiled-coil (RBCC) domain in KAP1 and the proximal coupling of ubiquitin conjugation to ER degradation (CUE) domain in SMARCAD1 mediate their direct interaction. Of note, retention of SMARCAD1 in the nucleus depended on KAP1 in both mouse ESCs and human somatic cells. Mutations in the CUE1 domain of SMARCAD1 perturbed the binding to KAP1 in vitro and in vivo Accordingly, an intact CUE1 domain was required for tethering this remodeler to the nucleus. Moreover, mutation of the CUE1 domain compromised SMARCAD1 binding to KAP1 target genes. Taken together, our results reveal a mechanism that localizes SMARCAD1 to genomic sites through the interaction of SMARCAD1's CUE1 motif with KAP1.