A deregulated stress response underlies distinct INF2 associated disease profiles

Abstract

Monogenic diseases provide favorable opportunities to elucidate the molecular mechanisms of disease progression and improve medical diagnostics. However, the complex interplay between genetic and environmental factors in disease etiologies makes it difficult to discern the mechanistic links between different alleles of a single locus and their associated pathophysiologies. Here we systematically characterize a large panel (>50) of autosomal dominant mutants of the actin regulator inverted formin 2 (INF2) that have been reported to cause the podocytic kidney disease focal segmental glomerulosclerosis (FSGS) and the neurological disorder Charcot Marie-Tooth disease (CMT). We found that INF2 mutations lead to deregulated activation of the formin and a constitutive stress response in cultured cells, primary patient cells and Drosophila nephrocytes.Using quantitative live-cell imaging we were able to identify distinct subsets of INF2 variants that exhibit varying degrees of activation. Furthermore, we could clearly distinguish between INF2 mutations that were linked exclusively to FSGS from those that caused a combination of FSGS and CMT. Our results indicate that cellular profiling of disease-associated mutations can contribute substantially to sequence-based phenotype predictions.

Rap1 Activity Is Essential for Focal Adhesion and Slit Diaphragm Integrity

“…To detect integrin β localization upon downregulation of rap1, ROIs were selected at the cell cortex and the cytosol close to the nucleus. The integrin reorganization was then calculated as previously described (Bayraktar et al, 2020): ROI at cell cortex + background area/ROI at cytosol + background area. Slit diaphragms were counted per µm basement membrane for 10 nephrocytes, each of 20-30 animals employing transmission electron microscopy images.…”

“…When imaging the surface of nephrocytes, integrin β presented in a typical fingerprint-like pattern in control nephrocytes (Figure 1B), whereas its localization was severely altered with diffuse targeting of integrin β in rap1 knockdown nephrocytes (Figure 1B). Integrin accumulation at the plasma membrane was quantified in tangential sections (Figure 1C) (Bayraktar et al, 2020). To confirm that integrin β mistargeting is a specific effect of rap1 downregulation, we performed rescue experiments of the rap1 knockdown by expressing human Rap1B, which is not targeted by the dsRNA specific to the Drosophila rap1 gene.…”

PI(4,5)P2 controls slit diaphragm formation and endocytosis in Drosophila nephrocytes

“…PI(4,5)P2 as well as PI(3,4,5)P3 are capable of regulating the actin cytoskeleton by recruiting and activating the small GTPases Rac1 and Cdc42 as well as proteins of the WASP family (Prehoda et al, 2000;Rohatgi et al, 2000;Padrick and Rosen, 2010). Notably, a coordinated actin cytoskeleton remodeling is essential for cortical Nephrin localization and slit diaphragm assembly in Drosophila nephrocytes (Muraleedharan et al, 2018;Bayraktar et al, 2020) as well as in mammalian podocytes (Perico et al, 2016). Vice versa, activated Nephrin recruits PI3K resulting in Rac1 activation, actin branching and lamellipodia formation in cultured rat podocytes (Zhu et al, 2008).…”

INF2 p.Arg214Cys mutation in a Chinese family with rapidly progressive renal failure and follow-up of renal transplantation: case report and literature review

“…INF2 mutations have been identified as being responsible for the development of autosomal dominant (AD) FSGS [3,4,6,7]. These mutations are also associated with Charcot-Marie-Tooth disease (CMT) [2,5,[8][9][10][11], which is characterized by a demyelinating peripheral neuropathy [9]. In CMT patients, an increased prevalence of FSGS has been documented [12].…”