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Screening for micropeptides in HUVEC with tryptic digestion, part 2

ABSTRACT: Long non-coding RNAs (lncRNA) are transcribed but not translated ribonucleic acids with various functions. We analyzed a so far unreported lncRNAn 342419, which we named MANTIS. A search for micropeptides after overexpression of MANTIS in Human Umbilical Vein Endothelial Cells (HUVEC) with subsequent LC-MS/MS with trypsination showed that none of the micropeptides found had any similarity to potential MANTIS ORFs.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Ilka Wittig

LAB HEAD: Ralf P. Brandes

PROVIDER: PXD005957 | Pride | 2017-03-27

REPOSITORIES: Pride

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Long noncoding RNA LISPR1 is required for S1P signaling and endothelial cell function.

Josipovic Ivana I Pflüger Beatrice B Fork Christian C Vasconez Andrea E AE Oo James A JA Hitzel Juliane J Seredinski Sandra S Gamen Elisabetta E Heringdorf Dagmar Meyer Zu DMZ Chen Wei W Looso Mario M Pullamsetti Soni Savai SS Brandes Ralf P RP Leisegang Matthias S MS

Journal of molecular and cellular cardiology 20180203

Sphingosine-1-Phosphate (S1P) is a potent signaling lipid. The effects of S1P are mediated by the five S1P receptors (S1PR). In the endothelium S1PR1 is the predominant receptor and thus S1PR1 abundance limits S1P signaling. Recently, lncRNAs were identified as a novel class of molecules regulating gene expression. Interestingly, the lncRNA NONHSAT004848 (LISPR1, Long intergenic noncoding RNA antisense to S1PR1), is closely positioned to the S1P1 receptors gene and in part shares its promoter re ...[more]

PMID: 29408197

Similar Datasets

Screening for micropeptides in HUVEC without tryptic digestion, part 1

Project description:Long non-coding RNAs (lncRNA) are transcribed but not translated ribonucleic acids with various functions. We analyzed a so far unreported lncRNA n342419, which we named MANTIS. A search for micropeptides after overexpression of MANTIS in Human Umbilical Vein Endothelial Cells (HUVEC) with subsequent LC-MS/MS without trypsination showed that none of the micropeptides found had any similarity to potential MANTIS ORFs, whereas the positive control yielded micropeptides encoded from GFP plasmid.

2017-03-27 | PXD005951 | Pride

Identification of interacting proteins of lncRNA HIF1α-AS1

Project description:LncRNA Hypoxia-inducible factor 1α-antisense 1 (HIF1α-AS1) is located on the antisense strand of the important Hypoxia-inducible factor 1α (HIF1α) gene, but being transcribed in antisense direction along the HIF1α promoter. Here we used the 3’end biotinylated HIF1a-AS1 RNA and a control RNA for RNA Pulldown and searched for interacting proteins in nuclear extracts of human umbilical vein endothelial cells (HUVEC).

2022-09-23 | PXD023512 | Pride

VE-cadherin associated proteins in the absence of lncRNA LASSIE

Project description:The shear stress-regulated lncRNA LASSIE interacts with junctional proteins (e.g. PECAM-1, which interacts with VE-cadherin) and influences endothelial barrier function. Here we characterize the remodeling of the VE-Cadherin complex by the lncRNA LASSIE. LASSIE silenced HUVECs were subjected to co-immunoprecipitation using an anti-VE-cadherin antibody. Differentially associated proteins were identified by Mass spectrometry. This analysis revealed a significantly decreased association of cytoskeleton-linked proteins with VE-cadherin after silencing of LASSIE. Functional assays confirmed this result and characterized LASSIE as a stabilizer of junctional complexes in endothelial cells, important for normal shear stress sensing and barrier function.

2021-09-09 | PXD018725 | Pride

Identification of lncRNA CALA-dependent protein interaction partners of G3BP1 in the cytoplasm

Project description:We previously identified the stimuli-responsive lncRNA CALA to impact endothelial cell function and identified G3BP1 as a main interaction partner of CALA in the cytoplasm of human umblical vein endothelial cells (HUVECs). To uncover the role of the lncRNA CALA in G3BP1-RNPs in the cytoplasm, we purified the G3BP1 interactome in control and CALA-silenced HUVECs via anti-G3BP1 immunoprecipitation followed by mass spectrometry. We thereby uncover the basal G3BP1 protein interactome in HUVECs and additionally identify lncRNA-dependent protein interactions of G3BP1 in the cytoplasm of HUVEC.

2022-07-04 | PXD033517 | Pride

Identification of VE-cadherin associated proteins regulated through the lncRNA LASSIE

Project description:The shear stress-regulated lncRNA LASSIE interacts with cytoskeletal proteins as shown by RNA-antisense purification and subsequent mass spectrometry analysis using an anti-LASSIE and a non-targeting 3’ Desthiobiotin-TEG labeled 2’ O-Me-RNA. As this analysis resulted in high protein binding to the non-targeting control oligonucleotide, the RNA antisense purification was repeated in HUVECs treated with control and anti-LASSIE siRNA. Mass spectrometry analysis confirmed binding of LASSIE to the Intermediate filament protein nestin, as nestin enrichment was decreased in the absence of LASSIE. Additional functional analyses demonstrate a role of LASSIE in shear stress sensing and barrier function in endothelial cells, by stabilizing endothelial cell junctions through connection to the cytoskeleton.

2021-09-09 | PXD018734 | Pride

Identification of protein interaction partners of the lncRNA ADPGK-AS1

Project description:M2-like macrophage upregulated lncRNA ADPGK-AS1 regulates macrophage phenotype and modulates the influence of macrophages on lung tumor apoptosis and migration. Here we identified proteins that interact with ADPGK-AS1 that might give insight into how this regulation this is regulated on molecular level. We were able to identify interaction with mitochondrial ribosomal proteins, leading to regulation of mitochondrial oxidative phosphorylation.

2023-08-13 | PXD030605 | Pride

Identification of protein interaction partners of the lncRNA Lassie

Project description:The shear stress-regulated lncRNA LASSIE interferes with endothelial cell function as knockdown of Lassie affects apoptosis, proliferation and angiogenic sprouting in vitro. RNA-antisense purification and subsequent mass spectrometry analysis identifed junctional proteins (such as PECAM-1) as interactors of LASSIE . Additional functional analyses demonstrate a role of LASSIE in shear stress sensing and barrier function in endothelial cells, by stabilizing endothelial cell junctions through connection to the cytoskeleton.

2021-07-22 | PXD018724 | Pride

Comprehensive computational identification and characterization of conserved small ORFs in animals

Project description:There is increasing evidence that transcripts or transcript regions annotated as non-coding can harbor functional short open reading frames (sORFs). Loss-of-function experiments have identified essential developmental or physiological roles for a few of the encoded peptides (micropeptides), but genome-wide experimental or computational identification of functional sORFs remains challenging. In this project we expand our previously developed method and present results of an integrated computational pipeline for the identification of conserved sORFs in human, mouse, zebrafish, fruit fly and the nematode C. elegans. This dataset represents the peaklist of identified novel and annotated mouse micropeptides by mass spectrometry.

2015-07-28 | PXD002583 | Pride

SILAC-based quantitative Nox4-Co-immunoprecipitation

Project description:Within the family of NADPH oxidases, Nox4 is unique as it is predominantly localized in the endoplasmic reticulum, has constitutive activity and generates H2O2. We hypothesize that these features are consequences of a so far unidentified Nox4-interacting protein. Interacting proteins were screened by quantitative SILAC-Co-immunoprecipitation in HEK293 cells stably overexpressing Nox4. By this technique, several interacting proteins were identified with calnexin showing the most robust interaction.

2016-02-17 | PXD003514 | Pride

Endothelial protein interaction partners of the lncRNA PCAT19

Project description:PCAT19 is an endothelial-enriched lncRNA that contributes to the proliferation-quiescence switch in a cell density-dependent manner. Here we performed an antisense-oligonucleotide pulldown of the PCAT19 lncRNA followed by mass spectrometry to identify protein interaction partners of PCAT19. Interaction partners were further analysed for their potential role with PCAT19.

2022-09-29 | PXD032669 | Pride

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