Defining the Molecular Character of the Developing and Adult Kidney Podocyte

Abstract

BackgroundThe podocyte is a remarkable cell type, which encases the capillaries of the kidney glomerulus. Although mesodermal in origin it sends out axonal like projections that wrap around the capillaries. These extend yet finer projections, the foot processes, which interdigitate, leaving between them the slit diaphragms, through which the glomerular filtrate must pass. The podocytes are a subject of keen interest because of their key roles in kidney development and disease.Methodology/Principal FindingsIn this report we identified and characterized a novel transgenic mouse line, MafB-GFP, which specifically marked the kidney podocytes from a very early stage of development. These mice were then used to facilitate the fluorescent activated cell sorting based purification of podocytes from embryos at E13.5 and E15.5, as well as adults. Microarrays were then used to globally define the gene expression states of podocytes at these different developmental stages. A remarkable picture emerged, identifying the multiple sets of genes that establish the neuronal, muscle, and phagocytic properties of podocytes. The complete combinatorial code of transcription factors that create the podocyte was characterized, and the global lists of growth factors and receptors they express were defined.Conclusions/SignificanceThe complete molecular character of the in vivo podocyte is established for the first time. The active molecular functions and biological processes further define their unique combination of features. The results provide a resource atlas of gene expression patterns of developing and adult podocytes that will help to guide further research of these incredible cells.

Defining cell-type specificity at the transcriptional level in human disease

“…Interestingly, neither of these genes is present in the list of 102 genes (Supplemental Table 6) identified as podocyte-specific using the murine ex vivo cell-lineage separation in the GUDMAP data sets. Brunskill et al (2011) recently generated a transcriptional data set (144 genes) regulated during murine podocyte development and enriched in adult podocytes in comparison with the renal cortex. Analysis in HPA of the human orthologs exhibited a similar enrichment to the GUDMAP ex vivo cell-lineage data set used in Figure 3 (two podocyte-specific in kidney [5%] and 13 podocyte-specific in glomeruli [30%]), but the murine data did not reach the specificity of the in silico nanodissection approach (65%).…”

“…Restricted gene expression is defined in this study as podocyte ''specific'' within the renal context if it shows gene expression limited to podocytes within the kidney, and ''podocyte specific within the renal glomerulus'' if it is expressed only in podocytes within the glomeruli but detectable in other extraglomerular cell lineages in the kidney. Previous high-throughput strategies have relied on mouse (Endlich et al 2002;Brunskill et al 2011;Jain et al 2011) or human (Saleem et al 2008) immortalized glomerular visceral epithelial cells, but in vitro culture leads to rapid loss of both lineagespecific phenotypes and lineage-specific gene expression. Wholetissue-based molecular profiles of renal disease are attainable Higgins et al 2004;Schmid et al 2006;Bennett et al 2007;Ju et al 2009;Hodgin et al 2010;Lindenmeyer et al 2010;Woroniecka et al 2011) since human renal tissue is routinely obtained by diagnostic fine needle biopsy, but wholetissue expression profiles have not previously been capable of identifying gene expression at the cell-lineage level.…”

“…Lineage tracing was established by GFP expression using cell-type-specific promoters, followed by FACS and genome-wide expression profiling of GFPpositive single-cell suspension (Brunskill et al 2011). Using the podocyte, mesangial, and endothelial cell-lineage-specific GUDMAP expression data sets (Supplemental Table 5), we subtracted endothelial and mesangial gene expression profiles from the transcriptome obtained from the podocyte preparation and identified 102 podocyte-specific transcripts (Methods; Supplemental Table 6; McMahon et al 2008).…”

“…The identifications of data sets that have been utilized in our study are listed in Supplemental Table 5. The detailed protocol of data generation is described in a recently published paper by Brunskill et al (2011) and can also be found in our Supplemental Methods. We preprocessed and normalized data as described in Microdissected Human Kidney Data.…”

Lmx1b and FoxC Combinatorially Regulate Podocin Expression in Podocytes

“…25 The mammalian FoxC subfamily has two highly related members, FoxC1 and FoxC2, where mouse Foxc2 is expressed in multiple tissues 26 in addition to podocytes. 24,27 Mutations in FOXC2 cause lymphedema-distichiasis syndrome, which is characterized by limb lymphedema and double rows of eyelashes. Lymphedemadistichiasis syndrome with proteinuria has been reported in a family of German-Irish descent, 28 suggesting that certain FOXC2 mutations may cause renal disease.…”