Characterization of novel genes of importance for renal glomerular function and disease

Glomerular kidney diseases are a major health care burden. The glomerular filtration barrier consists of three layers: the slit diaphragm that bridges the interlocking foot pro- cesses of the podocytes, the glomerular basement membrane and fenestrated endothelial cells. The filtration barrier is permselective to plasma macromolecules based on size, shape, and charge. The molecular makeup of the filtration barrier determines its permselectivity. Knowledge about the molecular mechanisms of the glomerular filtration barrier has been gained with the study of genes mutated in humans and animal models of glomerular kidney disease.

In the thesis work, we performed a proteome analysis of healthy glomeruli in mice using two-dimensional gel electrophoresis coupled to mass spectrometry. A total of 232 unique proteins were identified from 414 gel spots. This study provided a snapshot of the glomerular proteome that can serve as reference for future glomerular protein biomarker studies.

We describe the expression and physiological function of the gene Glcci1 in zebrafish. His- tological analysis of Glcci1 showed expression in podocytes and mesangial cells. In vivo and in vitro studies demonstrated that Glcci1 expression is induced by glucocorticoids. Depletion of Glcci1 by morpholino knockdown resulted in the development of pericardial edema and defects in glomerular filtration. Our results suggest a role for Glcci1 in glomerular injury and proteinuria.

Knockdown experiments of the paralogs Plekhh1 and Plekhh2 in zebrafish resulted in gross morphological changes in the glomerulus, including thickening of the glomerular basement membrane and disorganization of the podocyte foot processes associated with a defective filtration barrier. These results suggest a role for Plekhh1 and Plekhh2 in regulating podocyte foot process morphology in zebrafish. We further characterized Plekhh1 and Plekhh2 in knockout mouse models. Single knockouts of Plekhh1 and Plekhh2 do not develop any apparent phenotype. Plekhh1 and Plekhh2 deficient mice were intercrossed to produce mice lacking both genes. This yielded fewer than expected number of double knockout offspring, suggesting functional redundancy. Ultrastructural analysis of surviving double knockout mice did not reveal changes in glomerular morphology suggesting that Plekhh1 and Plekhh2 are largely redundant for kidney function in mice.

These results give insight into glomerular biology and pathomechanisms of kidney disease that might provide a basis for translational research in the future.

List of scientific papers

I. Sam Tryggvason, Masatoshi Nukui, Asmundur Oddsson, Karl Tryggvason, and Hans Jörnvall. Glomerulus proteome analysis with two-dimensional gel electrophoresis and mass spectrometry. Cell Mol Life Sci. 64.24 (Dec. 2007), 3317-35.
https://doi.org/10.1007/s00018-007-7463-6

II. Yukino Nishibori, Kan Katayama, Mataleena Parikka, Asmundur Oddsson, Masatoshi Nukui, Kjell Hultenby, Annika Wernerson, Bing He, Lwaki Ebarasi, Elisabeth Raschperger, Jenny Norlin, Mathias Uhlén, Jaakko Patrakka, Christer Betsholtz, and Karl Tryggvason. Glcci1 deficiency leads to proteinuria. Journal of the American Society of Nephrology. JASN. 22.11 (Nov. 2011), 2037-46.
https://doi.org/10.1681/ASN.2010111147

III. Asmundur Oddsson, Mataleena Parikka, Lwaki Ebarasi, Patricia Rodriguez, Yukino Nishibori, Kjell Hultenby, Annika Wernerson, Christer Betsholtz, Jaakko Patrakka, and Karl Tryggvason. Zebrafish Plekhh1 and Plekhh2 are involved in organization of foot processes and normal kidney function. [Manuscript]

IV. Asmundur Oddsson, Mark Lal, Kjell Hultenby, Annika Wernerson Annika, Jaakko Patrakka, and Karl Tryggvason. Analysis of Plekhh1 and Plekhh2 knockout mice reveal redundancy of the paralogs in kidney function. [Manuscript]