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Transcriptome data of wt, Rbx1- or Ube2m- deficient Treg cells from mice

ABSTRACT: Cullin-RING Ligases (CRLs), upon activation by neddylation, play the key roles in regulating many biological processes. However, how CRL-neddylation regulates the function of Treg cells remains elusive. Here we show that mice with Treg-specific deletion of Rbx1, a RING component of CRLs required for its activity, developed an early-onset fetal inflammatory disorders and death at day ~25 after birth with disrupted homeostasis and impaired suppressive functions of Treg cells. Specifically, Rbx1 is essential for maintenance of the effector subpopulations in Treg cells, and regulates several inflammatory pathways. Similar phenotypes were seen in mice with deletion of Ube2m, a neddylation E2, in Treg cells, but with much lesser severity. Interestingly, Treg-specific deletion of Rbx2/Sag or Ube2f, the family member of Rbx1 or Ube2m, respectively, had no obvious phenotype. Thus, the Ube2m-Rbx1 axis is required for the maintenance of homeostasis and functions of Treg cells; and Rbx1 has Ube2m-independent roles in the fitness of Treg cells, suggesting a layer of complexity in neddylation activation of CRLs. We use the tranpritome dara to show the mechanism of the function of Rbx1 and Ube2m in Treg cells, also research the relationship of Rbx1 and Ube2m in Treg cells.

ORGANISM(S): Mus musculus

PROVIDER: GSE192664 | GEO | 2021/12/28

REPOSITORIES: GEO

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Project description:Clearance of damaged mitochondria via mitophagy is crucial for cellular homeostasis. While the role of ubiquitin (Ub) ligase PARKIN in mitophagy has been extensively studied, increasing evidence suggests the existence of PARKIN-independent mitophagy in highly metabolically active organs such as the heart. Here, we identify a crucial role for Cullin-RING Ub ligase 5 (CRL5) in basal mitochondrial turnover in cardiomyocytes. CRL5 is a multi-subunit Ub ligase comprised by the catalytic RING box protein RBX2 (also known as SAG), scaffold protein Cullin 5 (CUL5), and a substrate-recognizing receptor. Analysis of the mitochondrial outer membrane-interacting proteome uncovered a robust association of CRLs with mitochondria. Subcellular fractionation, immunostaining, and immunogold electron microscopy established that RBX2 and Cul5, two core components of CRL5, localizes to mitochondria. Depletion of RBX2 inhibited mitochondrial ubiquitination and turnover, impaired mitochondrial membrane potential and respiration, and increased cell death in cardiomyocytes. In vivo, deletion of the Rbx2 gene in adult mouse hearts suppressed mitophagic activity, provoked accumulation of damaged mitochondria in the myocardium, and disrupted myocardial metabolism, leading to rapid development of dilated cardiomyopathy and heart failure. Similarly, ablation of RBX2 in the developing heart resulted in dilated cardiomyopathy and heart failure. Notably, the action of RBX2 in mitochondria is not dependent on PARKIN, and PARKIN gene deletion had no impact on the onset and progression of cardiomyopathy in RBX2-deficient hearts. Furthermore, RBX2 controls the stability of PINK1 in mitochondria. Proteomics and biochemical analyses further revealed a global impact of RBX2 deficiency on the mitochondrial proteome and identified several mitochondrial proteins as its putative substrates. These findings identify RBX2-CRL5 as a mitochondrial Ub ligase that controls mitophagy under physiological conditions in a PARKIN-independent, PINK1-dependent manner, thereby regulating cardiac homeostasis.

Project description:The COP9 signalosome (CSN) is a conserved protein complex occurring in all eukaryotes. The mammalian CSN consists of eight subunits (CSN1 – CSN8) and possesses an intrinsic metalloprotease JAMM motif localised to CSN5. In addition, it is associated with kinases such as protein kinase CK2 and D and the ubiquitin (Ub) specific protease 15. It regulates cullin-RING Ub ligases (CRLs) that label proteins for proteolysis with poly-Ub chains in the Ub proteasome system (UPS). CRLs contain one of the scaffolding cullins 1 – 7, a RING domain protein e. g. Rbx1, and one of hundreds of substrate binding units such as F-box or BTB proteins that specifically target proteins for ubiquitination. The CSN forms functional supercomplexes with CRLs. It removes the Ub-like protein Nedd8 from cullins by its metalloprotease activity and thereby inactivates CRLs. On the other hand, it is essential for CRL function by protecting components and allowing reassembly of the complexes. As a regulator of the UPS the CSN is involved in cell cycle10, DNA repair and development. Overexpression of CSN subunits has been reported in tumour cells. Up to date nothing is known about CSN biogenesis and its regulation. Ten years ago we have observed that overexpression of selected CSN subunits caused a coordinated increase of all CSN subunits and a de novo biosynthesis of the entire complex. Here we show that overexpression of CSN1 mediates an increase of c-Myc which coordinates CSN subunit expression via microRNAs (miRNAs). miRNAs are evolutionarily conserved, endogenous, small, noncoding RNA molecules of about 22 nucleotides that function as posttranscriptional gene regulators. We found that inhibitors of let-7 family miRNAs or upregulation of c-Myc, which represses let-7 miRNAs, resulted in a coordinated increase of CSN subunits and de novo CSN biogenesis. Keywords: Gene expression analysis of human RNA samples using topic-defined PIQOR™ Ubiquitin-PS Microarrays. Two different platforms were used (these are two different microarray batches which are identical in terms of the spotted cDNAs, but differ in the position of some cDNAs) Gene expression analysis of Hela cells overexpressing either CSN2 or CSN1 was performed to study CSN biogenesis and its regulation. Curcumin, Nr. 8 (Curcumin analogon) and Piceatannol were used for the treatment of Hela cells since they inhibit the CSN associated kinases. The aim was to investigate the effect on the signalosome. For each treatment 50 µM Curcumin, Nr. 8 and Piceatannol were used.

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Transcriptome data of wt, Rbx1- or Ube2m- deficient Treg cells from mice

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