AKAP12 variant 1 knockout enhances vascular endothelial cell motility
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PROVIDER: PRJNA1201108 | ENA |
REPOSITORIES: ENA
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Project description:This study investigates the role of AKAP12 in endothelial cell motility with a specific focus on AKAP12 variants, AKAP12v1 and AKAP12v2. Previous work has shown that AKAP12, a multivalent A-kinase anchoring protein that binds to PKA and several other proteins regulating protein phosphorylation, is expressed at low levels in most endothelia in vivo but is expressed at higher levels in cells in vitro. Here, we found that AKAP12 expression in endothelial cell (HUVEC) cultures was cell density-dependent, with the expression being highest in subconfluent cultures and lowest in confluent cultures. AKAP12 expression was also elevated in cells at the wound edge of wounded endothelial cell monolayers. Knockdown of variants 1 and 2 inhibited cell migration. However, CRISPR/Cas9 knockout of AKAP12v1 enhanced migration, indicating that the absence of this variant and the presence of AKAP12v2 likely alters the signaling events controlling cell motility. Further analysis using bulk RNA sequencing revealed that the loss of AKAP12v1 affected genes associated with cell migration and intercellular junctions. We propose that AKAP12v1 and AKAP12v2 play distinct yet complementary roles in endothelial cell migration and likely work together in controlling the signaling events associated with vascular repair and development.
2025-08-07 | GSE285043 | GEO
Project description:Dysregulation of lipid homeostasis is a feature of alcohol-associated liver disease (ALD). A-kinase anchoring protein 12 (AKAP12) is a scaffolding partner of the cAMP-dependent protein kinase, PKA that controls its spatiotemporal localization. Activation of PKA by cAMP inhibits lipogenesis and facilitates fatty acid oxidation (FAO). We examined how AKAP12’ could regulate alcohol-associated steatosis. Alcohol exposure reduced AKAP12’s interaction with PKA and suppressed PKA activation. Forced expression of AKAP12 overcame the suppression of PKA activity in human hepatocytes and in livers of mice exposed to alcohol. This led to a decrease in hepatic steatosis. Disrupting the scaffold of AKAP12 and PKA by CRISPR editing increased steatosis and inflammation. RNA sequencing analysis of hepatocytes from alchol fed mice and normal mice demonstrated key lipogenic and inflammatory pathways regulated by AKAP12 overexpression. RNA sequencing of total liver from mice with AKAP12-PKA binding site CRISPR editing or AKAP12 overexpression confirmed regulation of lipogenic and inflammatory pathways by AKAP12
2025-03-02 | GSE290266 | GEO
Project description:We compared gene expression in AKAP12 knockdown and control cells in the SW480 background.
2020-09-16 | GSE147739 | GEO
Project description:Neurobiological consequences of traumatic brain injury (TBI) result from a complex interplay of secondary injury responses and sequela that mediates chronic disability. Endothelial cells are important regulators of the cerebrovascular response to TBI. Our work demonstrates that genetic deletion of endothelial cell (EC)-specific EPH receptor A4 (EphA4) using conditional EphA4f/f/Tie2-Cre and EphA4f/f/VE-Cadherin-CreERT2 knockout (KO) mice, promotes blood-brain barrier (BBB) integrity and tissue protection, which correlates with improved motor function and cerebral blood flow recovery following controlled cortical impact (CCI) injury. scRNAseq of capillary-derived KO ECs showed increased differential gene expression of BBB-related junctional and actin cytoskeletal regulators, namely, A-kinase anchor protein 12, Akap12, whose presence at Tie2 clustering domains is enhanced in KO microvessels. Transcript and protein analysis of CCI-injured whole cortical tissue or cortical-derived ECs suggests EphA4 limits the expression of Cldn5, Akt, and Akap12 and promotes Ang2. Blocking the Tie2 receptor using sTie2-Fc, attenuated BBB and tissue protection and reversed Akap12 mRNA and protein levels in KO compared to WT cortical-derived ECs. Conversely, direct stimulation of Tie2 using Vasculotide, an angiopoietin-1 memetic peptide, phenocopied the neuroprotection. Finally, we report a noteworthy rise in soluble Ang2 in the sera of individuals with acute TBI, highlighting its promising role as a vascular biomarker for early detection of BBB disruption. Overall, we describe a novel contribution of the axon guidance molecule, EphA4, in mediating TBI microvascular dysfunction through negative regulation of Tie2/Akap12 signaling.
2023-09-24 | GSE240430 | GEO
Project description:Regulation of subcellular mRNA localisation is a fundamental biological mechanism, which adds a spatial dimension to the diverse layers of post-transcriptional control of gene expression. Insights into this phenomenon have been described in a multitude of cell types and across taxonomic kingdoms, highlighting its biological significance. The cellular compartment in which mRNAs are located may define distinct aspects of the encoded proteins, ranging from production rate and complex formation to localised activity. Ultimately, mRNA localisation supports compartmentalised mechanistic outputs that can respond to local stimuli, orient migration, or even shape cells. We used an unbiased method to profile the RNA-bound proteome in migrating endothelial cells and found that the plasma membrane (PM)-associated A-kinase anchor protein 12 (AKAP12) interacts with various mRNAs, including mRNAs encoding kinases with Actin remodelling activity. To identify transcripts bound by AKAP12, we carried out UV crosslinking experiments followed by RNA immunoprecipitation and high throughput sequencing. Our data offer novel insights in complex mechanisms of spatial control of gene expression.
2024-03-28 | GSE247682 | GEO
Project description:AKAP12 KO mice showed reduced fibrosis resolution in DDC-induced hepatic fibrosis and resolution model. To compare gene expression profile between genotypes, transcriptome was analyzed by RNA sequencing.
2018-09-01 | GSE104030 | GEO
Project description:Total RNA sequencing of AKAP12 knockdown cells
| PRJNA616288 | ENA
Project description:We applied single-cell RNA-Seq to analyze human diseased arteries, and identified histone variant H2A.Z as a key histone signature to maintain vascular smooth muscle cell (VSMC) identity. We show that H2A.Z occupies genomic regions near VSMC marker genes and its occupancy is decreased in VSMC undergoing dedifferentiation. Mechanistically, H2A.Z occupancy preferentially promotes nucleosome turnover, facilitates the recruitment of Smad3 and Med1 to VSMC marker genes, thereby activating gene expression. In human diseased vascular tissue, H2A.Z expression dramatically decreased. Notably, in vivo overexpression of H2A.Z rescued injury-induced loss of VSMC identity and neointima formation. Together, our data introduce dynamic occupancy of histone variant as a novel regulatory basis contributing to cell fate decisions, and imply H2A.Z as a potential intervention node for vascular diseases.
2018-06-08 | GSE112417 | GEO
Project description:The integrity of blood vessels controls vascular permeability and extravasation of blood cells, across the endothelium. Thus, the impairment of endothelial integrity leads to hemorrhage, edema, and inflammatory infiltration. However, the molecular mechanism underlying vascular integrity has not been fully understood. Here, we demonstrate an essential role for A-kinase anchoring protein 12 (AKAP12) in the maintenance of endothelial integrity during vascular development. Zebrafish embryos depleted of akap12 (akap12 morphants) exhibited severe hemorrhages. In vivo time-lapse analyses suggested that disorganized interendothelial cell-cell adhesions in akap12 morphants might be the cause of hemorrhage. To clarify the molecular mechanism by which the cell-cell adhesions are impaired, we examined the cell-cell adhesion molecules and their regulators using cultured endothelial cells. The expression of PAK2, an actin cytoskeletal regulator, and AF6, a connector of intercellular adhesion molecules and actin cytoskeleton, was reduced in AKAP12-depleted cells. Depletion of either PAK2 or AF6 phenocopied AKAP12-depleted cells, suggesting the reduction of PAK2 and AF6 results in the loosening of intercellular junctions. Consistent with this, overexpression of PAK2 and AF6 rescued the abnormal hemorrhage in akap12 morphants. We conclude that AKAP12 is essential for integrity of endothelium by maintaining the expression of PAK2 and AF6 during vascular development.
| S-EPMC3317486 | biostudies-literature
Project description:The flagellar motor is a powerful macromolecular machine used to propel bacteria through various environments. Flagellar motility of the alpha-proteobacterium Sinorhizobium meliloti is nearly abolished in the absence of the transcriptional regulator LdtR, which is involved in peptidoglycan remodeling. We report that LdtR does not regulate motility gene transcription. Remarkably, the motility defects of the DldtR mutant can be restored by secondary mutations in the motility gene motA or a previously uncharacterized gene in the flagellar regulon, which we named motS. MotS is not essential for S. meliloti motility and may serve an accessory role in flagellar motor function. Structural modeling predicts that MotS is comprised of an N-terminal transmembrane segment, a long-disordered region, and a conserved β-sandwich domain. The C-terminus of MotS is localized in the periplasm. Genetics-based substitution of MotA with a MotAG12S variant protein also restored the ΔldtR motility defect. The MotAG12S variant causes a local polarity shift at the periphery of the MotAB stator units. We propose that MotS may be required for optimal alignment of stators in wild-type flagellar motors but becomes detrimental in cells with altered peptidoglycan. Similarly, the polarity shift in the stator units composed of MotB/MotAG12S might stabilize its interaction with altered peptidoglycan.
2024-06-18 | GSE251837 | GEO