A Deregulated Stress Response Underlies Distinct INF2-Associated Disease Profiles

Abstract

BackgroundMonogenic 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. Inverted formin 2 (INF2), an actin regulator, mediates a stress response—calcium mediated actin reset, or CaAR—that reorganizes the actin cytoskeleton of mammalian cells in response to calcium influx. It has been linked to the podocytic kidney disease focal segemental glomerulosclerosis (FSGS), as well as to cases of the neurologic disorder Charcot–Marie–Tooth disease that are accompanied by nephropathy, mostly FSGS.MethodsWe used a combination of quantitative live cell imaging and validation in primary patient cells and Drosophila nephrocytes to systematically characterize a large panel of >50 autosomal dominant INF2 mutants that have been reported to cause either FSGS alone or with Charcot–Marie–Tooth disease.ResultsWe found that INF2 mutations lead to deregulated activation of formin and a constitutive stress response in cultured cells, primary patient cells, and Drosophila nephrocytes. We were able to clearly distinguish between INF2 mutations that were linked exclusively to FSGS from those that caused a combination of FSGS and Charcot–Marie–Tooth disease. Furthermore, we were able to identify distinct subsets of INF2 variants that exhibit varying degrees of activation.ConclusionsOur results suggest that CaAR can be used as a sensitive assay for INF2 function and for robust evaluation of diseased-linked variants of formin. More broadly, these findings indicate that cellular profiling of disease-associated mutations has potential to contribute substantially to sequence-based phenotype predictions.

Impaired Mitochondrial Mobility in Charcot-Marie-Tooth Disease

“…All INF2 CMT mutations are predicted or have been shown to increase actin assembly ( Bayraktar et al, 2020 ). While some actin-binding motor proteins likely facilitate microtubule-independent mitochondrial transport, numerous studies have shown that long-range microtubule-based mobility of mitochondria is antagonized by actin and actin-binding motor proteins ( Chada and Hollenbeck, 2004 ; Quintero et al, 2009 ; Pathak et al, 2010 ; Venkatesh et al, 2019 ; Cardanho-Ramos et al, 2020 ).…”

Formins in Human Disease

“…An in silico analysis of the effect of the pathogenic mutations indicates that they have a destabilizing structural effect in the DID [106]. This destabilization might affect the interaction of the DID with the DAD or with regulatory proteins [110,111], and results in gain-of-function of the actin polymerization activity of INF2. The case of a patient with combined FSGS and CMT has been described, in which a complete duplication of the INF2 gene occurred, which represents further evidence of a gain-of-function phenotype in INF2-linked disease [93].…”

Genetic Neuropathy Due to Impairments in Mitochondrial Dynamics

“…Additionally, mitochondrial motility was also reduced in cells expressing the A149D mutation, which is also likely relevant to the peripheral neuropathy phenotype. Notably, recent work shows that like the A149D mutation, pathogenic variants in INF2 cause aberrant formin activity [ 273 ], suggesting they may have similar impacts on mitochondrial fission. Nonetheless, as INF2 has multiple isoforms and impaired actin function can impact several cellular functions, it is not clear exactly how pathogenic variants in INF2 cause disease.…”