Genetic Alterations Causing Structural Changes of the Diaphanous Inhibitory Domain in Inverted Formin 2 May Contribute to the Podocyte Infolding Glomerulopathy

Podocyte infolding glomerulopathy (PIG) was originally reported in 1992 and then proposed as a new disease entity in 2008 (1). There are approximately 160 cases reported globally (2, 3). PIG is characterized by the presence of microtubules and microspheres within the glomerular basement membrane (GBM), podocytic membranous infolding, and foot process effacement. The pathogenesis remains to be illustrated. The largest case series (116 cases) was recently reported by Hong et al. (3). The authors established that a variety of diseases can present with the phenotype of PIG. Additionally, they analyzed alterations in the biochemical constituents of the GBM using laser microdissection and mass spectrometry. Kudose and Stokes (4) reviewed this most recent report and commented that there could be “population-based genetic or epigenetic” alterations that play important roles in the pathogenesis of PIG since most PIG cases are reported from Japan and China. Based on our clinicopathologic observation and currently available literature related to the genetic alteration, we hereby try to understand and explore the possible mechanism that may cause PIG.

The sequence of events leading to podocyte injury begins with the effacement of foot processes, indicating disarray within cytoskeletal structures, including actin filaments and microtubules. This disorganization suggests a potential imbalance in the synthesis and degradation of the cytoskeletal components. Furthermore, a thickened GBM, along with the presence of microtubules and microspheres, as well as the intrusion of the podocyte membrane into the GBM, points to a disruption in the physiologic regulation of substance transport between the podocyte and the GBM. These processes, although seemingly distinct, are actually interrelated steps in a progressive sequence of pathologic changes.

The cytoskeleton dynamics in the podocyte, including the foot process, is intricately regulated by approximately 20 genes, including ARHGDIA, AVIL, CDK20, INF2, FAT1, MAGI2, and PODXL. Genetic alterations causing an even slight impairment of the podocyte cytoskeletal apparatus can result in structural changes in the foot process as well as the slit diaphragm, leading to proteinuria and glomerular disease (5, 6).

The formins family is a group of proteins controlling cellular adhesion, cell shape changes, and morphogenesis by remodeling the actin and microtubule cytoskeletons. As a member, inverted formin 2 (INF2) may play a critical role in the formation of adhesion-associated actin structures adjacent to the extracellular matrix involving cellular protrusions (such as filopodia and lamellipodia), cell migration, and tissue morphogenesis. It also has been reported that INF2 plays roles in actin polymerization and depolymerization as well as microtubule formation (7). Feng et al. (2) reported INF2 genetic alterations (multiple single nucleotide polymorphisms and a nonframeshift deletion) in exon 8 in patients with PIG using whole exome sequencing, although in a very limited number of patients.

INF2 carries the following domains (a simplified scheme from the N to C terminus) and functions: G domain: binding to Rho GTPase; diaphanous inhibitory domain (DID): interacting with the diaphanous autoregulatory domain (DAD); formin homology 1 (FH1) and formin homology 2 (FH2) domains: nucleating G actin and growing the actin filament, also serving the actin filament and promoting formation of stabilized and bundled microtubules; and DAD: binding to the G domain and inhibiting biologic activity of INF2 (Figure, A). In the wild-type, when DID interacts with DAD, this interaction closes the INF2 protein as an inactive status; when Rho GTPase competitively interacts with the G domain, the DID–DAD interaction will be disrupted, opening the INF2 protein in a catalytically active form (Figure, B). When the DID is mutated, it may lose or weaken its capacity to interact with DAD, leaving the mutated INF2 in an active form, letting the FH1 and FH2 to constitutively polymerize actin molecules and stabilize synthesized microtubules (Figure, C) (6).

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Pathogenesis of PIG

Citation: Kidney News 16, 5

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These excessively synthesized actin filaments and microtubules can accumulate inside the podocyte, physically interact with each other, and eventually be mislocalized. (For example, microtubules will migrate into the foot process and coexist with actin filaments in both filopodia and lamellipodia.) These disorganized microtubules–actin filaments in excessive amounts in the foot process can provide force to extend and deform the membrane lipid bilayer toward the GBM, causing membrane invagination, protruding and abandoning actin filaments, microtubules, and other cellular contents into the GBM (Figure, D). It has been reported that the mutations in the DID, originating from exons 2 to 6, lead to focal segmental glomerulosclerosis (8). We are interested in finding the causative effects of genetic alterations in exon 8 or even exon 7 in PIG.

The podocyte is specifically terminally differentiated, and its gene expression profiling is unique. It would be interesting to isolate and purify the podocyte/foot process (9, 10), then check whole exon expressions, and identify all PIG-related variants using whole exome sequencing. In addition to INF2, genetic alterations from other genes (5, 6) that involve in actin polymerization and severance, the microtubule's stabilization, bundling, or severance can also cause the cytoskeleton changes in the podocyte/foot process.

If the aforementioned hypothesis is proven to be true, it could be easier to decide whether or not PIG is a new disease entity. When PIG is induced by the genetic alterations that lead to the destabilization of the podocytic cytoskeleton, it can be counted as the primary PIG and of course a new entity; if PIG is observed in other diseases (pathogenic immune reactions and inflammation) due to the interruption of the podocytic cytoskeleton organization, it could be categorized as acquired PIG.