Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Gene expression profiles of wild type and Actinin 4 point mutant knock-in adult podocyte cells at 2, 5 and 44 weeks following TRAP purification. (GUDMAP Series ID: 49)

ABSTRACT: The long term goal is to create an encyclopaedia of podocyte gene expression both in homeostasis, development and disease. Podocytes are identified by their expression of an eGFPL10a fusion protein whose expression is driven by a Col1a1 promoter fragment. Podocyte-specific RNA is generated by eGFP-affinity purification of polysomes and then subject to microarray analysis. Expression profiling of wild type and Actinin 4 point mutant knock-in kidney podocytes by TRAP. Col1a1-eGFPL10a mice were crossed against Actn4+/tm1.1Plk (KI) mice. Wild-type or KI mice were subject to TRAP at the following time points: 2, 5 and 44 weeks.

ORGANISM(S): Mus musculus

SUBMITTER: GUDMAP Developers

PROVIDER: E-GEOD-53156 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Glomerular podocytes are highly differentiated cells that are key components of the kidney filtration units. The podocyte cytoskeleton builds the basis for the dynamic podocyte cytoarchitecture and plays a central role for proper podocyte function. Recent studies implicate that immunosuppressive agents including the mTOR-inhibitor everolimus have a protective role directly on the stability of the podocyte cytoskeleton. To elucidate mechanisms underlying mTOR-inhibitor mediated cytoskeletal rearrangements, we carried out microarray gene expression studies to identify target genes and corresponding pathways in response to everolimus. We analyzed the effect of everolimus in a puromycin aminonucleoside experimental in vitro model of podocyte injury. Upon treatment with puromycin aminonucleoside, microarray analysis revealed gene clusters involving cytoskeletal-associated pathways, adhesion, migration and extracellular matrix composition to be affected. Everolimus is capable of protecting podocytes from injury, both on the transcriptome and protein level. Rescued genes included TUBB2B and DCDC2, both involved in microtubule structure formation in neuronal cells but not identified in podocytes so far. Confirming gene expression data, Western-blot analysis in cultured podocytes showed an increase of TUBB2B and DCDC2 protein after everolimus treatment, and immunohistochemistry in healthy control kidneys confirmed a podocyte-specific expression. Microtubule-inhibitor experiments led to a maldistribution of TUBB2B and DCDC2 as well as an aberrant reorganization of the actin cytoskeleton. Tubb2bbrdp/brdp mice showed a delay in glomerular podocyte and capillary development. Taken together, our study suggests that off-target, non-immune mediated effects of the mTOR-inhibitor everolimus on the podocyte cytoskeleton might involve regulation of microtubules, revealing a potential novel role of TUBB2B and DCDC2 in glomerular podocyte development We analyzed the effect of everolimus on gene expression in a puromycin aminonucleoside experimental in vitro model of podocyte injury using microarrays

Dataset Information

Gene expression profiles of wild type and Actinin 4 point mutant knock-in adult podocyte cells at 2, 5 and 44 weeks following TRAP purification. (GUDMAP Series ID: 49)

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