DAAM2 Variants Cause Nephrotic Syndrome via Actin Dysregulation

Abstract

The discovery of >60 monogenic causes of nephrotic syndrome (NS) has revealed a central role for the actin regulators RhoA/Rac1/ Cdc42 and their effectors, including the formin INF2. By whole-exome sequencing (WES), we here discovered bi-allelic variants in the formin DAAM2 in four unrelated families with steroid-resistant NS. We show that DAAM2 localizes to the cytoplasm in podocytes and in kidney sections. Further, the variants impair DAAM2-dependent actin remodeling processes: wild-type DAAM2 cDNA, but not cDNA representing missense variants found in individuals with NS, rescued reduced podocyte migration rate (PMR) and restored reduced filopodia formation in shRNA-induced DAAM2-knockdown podocytes. Filopodia restoration was also induced by the formin-activating molecule IMM-01. DAAM2 also co-localizes and co-immunoprecipitates with INF2, which is intriguing since variants in both formins cause NS. Using in vitro bulk and TIRF microscopy assays, we find that DAAM2 variants alter actin assembly activities of the formin. In a Xenopus daam2-CRISPR knockout model, we demonstrate actin dysregulation in vivo and glomerular maldevelopment that is rescued by WT-DAAM2 mRNA. We conclude that DAAM2 variants are a likely cause of monogenic human SRNS due to actin dysregulation in podocytes. Further, we provide evidence that DAAM2-associated SRNS may be amenable to treatment using actin regulating compounds.

Daam2 Regulates Myelin Structure and the Oligodendrocyte Actin Cytoskeleton through Rac1 and Gelsolin

“…This study demonstrates that Daam2 plays a role in regulating the actin-severing protein Gelsolin; however, the direct activity of Daam2 in actin polymerization remains unknown. As part of the formin family of proteins, Daam2 contains and Formin homology 2 (FH2) and FH3 domains (Lee and Deneen, 2012;Schneider et al, 2020); of these domains, FH2 is known to bind to effector proteins and even nucleate actin filaments directly (Liu et al, 2008;Jaiswal et al, 2013). In addition, its closest homologous protein, Daam1, cooperates with Rho GTPases (Habas et al, 2001) and actin-binding proteins such as profilin (Jaiswal et al, 2013) in a variety of contexts.…”

Podocyte Endocytosis in Regulating the Glomerular Filtration Barrier

“…Genetic mutations in actin cytoskeleton-interacting proteins have been shown to cause proteinuric CKD. Mutations such as MYO9A (a Rho-GTPase activating protein myosin) ( 66 ), MYO1E (nonmuscle class I myosin, myosin 1E) ( 67 ), INF2 (a member of the formin family of actin-regulating proteins INF2) ( 68 ), DAMM2 (a member of the formin family of actin-regulating proteins DAAM2) ( 69 ), ACTN4 (alpha-actinin 4) ( 70 ), CD2AP (CD2-associated protein) ( 71 ), and ANLN (filamentous actin [F-actin] binding protein) ( 72 ) have been identified in patients with nephrotic syndrome and histological evidence of focal segmental glomerulosclerosis (FSGS). Recent evidence suggests that the actin cytoskeleton is critical for CME in cells including the podocytes and have been implicated also in CIE ( 35 , 49 ).…”

Formins in Human Disease

“…Recessive mutations of DAAM2 (Figure 2 and Supplementary Table S6), have recently been involved in nephrotic syndrome, type 24 (NPHS24, MIM: 606627) [37]. All the affected individuals presented FSGS with no extra-renal manifestations.…”

“…The mutations at the DID and DAD appear to cause increased autoinhibition and, consequently, loss-of-function of actin polymerization activity. DAAM2, which is expressed by podocytes, colocalizes and associates with INF2 [37], suggesting the existence of crosstalk between the two formins that, given the link between INF2 and renal disease, may explain the renal damage caused by pathogenic DAAM2. Other formins might be involved in other kidney disorders.…”

“…The mutation of some of the formin genes causes monogenic disorders, as is the case of DIAPH1, which was the first formin gene found to be linked to a human Mendelian disorder [6]. Alteration of seven formins genes (Figure 2) have been acknowledged to date by the Online Mendelian Inheritance in Man (OMIM ® , McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University (Baltimore, MD, USA), https://omim.org; accessed on 20 September 2021) as meeting the criteria for consideration as a primary cause of human monogenic disorders [7,8]: DIAPH1-3 [6, (Supplementary Tables S3-S5), DAMM2 [37] (Supplementary Table S6), FORMIN2 [38][39][40][41][42] (FMN2) (Supplementary Table S7), INVERTED FORMIN 2 (INF2) (Supplementary Table S8) and FHOD3 [95][96][97][98][99][100] (Supplementary Table S9). The mutations or dysregulation of the other formins have not been demonstrated to be the primary cause of the phenotype, although they probably contribute to it [101,102].…”