Linking Basement Membrane and Slit Diaphragm in Drosophila Nephrocytes.

KEY POINTS: Drosophila nephrocytes feature a special basement membrane that may serve to model joint function of the glomerular filtration barrier. Silencing of Drosophila laminin and collagen IV genes reduced the density of slit diaphragms in nephrocytes, showing a direct effect of the matrix. Matrix receptor silencing phenocopied basement membrane disruption, indicating that the matrix guides slit diaphragm position through matrix receptors. BACKGROUND: The glomerular basement membrane and the slit diaphragm are essential parts of the filtration barrier. How these layers collaborate remains unclear. The podocyte-like nephrocytes in Drosophila harbor both a slit diaphragm and a basement membrane, serving as a model to address this critical question. METHODS: Basement membrane components and matrix receptors were silenced using RNA interference in nephrocytes. Slit diaphragms were analyzed using immunofluorescence, followed by automated quantification. Tracer endocytosis was applied for functional readouts. RESULTS: Immunofluorescence indicated a significant reduction in slit diaphragm density upon loss of laminin and collagen IV components. This was accompanied by reduced expression of fly nephrin and shallower membrane invaginations. Tracer studies revealed that the basement membrane defines properties of the nephrocyte filtration barrier. Acute enzymatic disruption of the basement membrane via collagenase rapidly caused slit diaphragm mislocalization and disintegration, which was independent of cell death. Loss of matrix-interacting receptors, particularly integrins mys and mew, phenocopied basement membrane disruption. Integrins and nephrin colocalized at the slit diaphragm in nephrocytes in a mutually dependent manner, interacting genetically. Human integrin α3 interacted physically with nephrin. CONCLUSIONS: The glomerular basement membrane model in Drosophila nephrocytes reveals that matrix receptor–mediated cues ensure correct positioning of the slit diaphragm and the overall filtration barrier architecture.