The slit diaphragm

Over the past three decades, more than 50 genetic causes of SRNS have been identified. Because these disease-causing mutations have been identified predominantly in genes expressed by podocytes, these terminally differentiated epithelial cells of the glomerular filtration barrier have emerged as central to disease pathogenesis. Podocytes synthesize various components of the slit diaphragm. Pathogenic mutations in NPHS2, the gene that encodes the slit diaphragm protein Podocin, have been found in 40% of familial (i.e., autosomal recessive) and 6% to 17% of sporadic SRNS. Affected individuals typically manifest with nephrotic-range proteinuria and rapid progression to ESKD. Histologic findings vary on the spectrum of MCN to FSGS (see Table 7.1 ). Podocin is a hairpin-shaped transmembrane protein that is expressed by podocytes. Podocin is a lipid raft- associated protein that localizes to podocyte foot processes and participates in various intracellular signaling processes through recruitment of other slit diaphragm proteins such as nephrin and CD2AP. Disease-causing podocin mutations most commonly manifest in children between 3 months and 5 years of age but can also manifest later in life. The age of disease onset can vary with the specific podocin mutation. Patients who are homozygous for the R138Q mutation or who have a heterozygous truncating mutation manifest disease at ∼2 years of age, whereas patients with two missense podocin mutations will manifest disease at ∼5 years of age. In addition, adult-onset FSGS has been reported in patients who are compound heterozygotes for the R229Q podocin mutation and a second mutant NPHS2 allele.

CD2-associated protein (CD2AP) is a multifunctional scaffolding protein that links membrane proteins to the cytoskeleton. CD2AP is an essential component of the glomerular slit diaphragm that participates in the regulation of actin cytoskeletal dynamics and is involved in various aspects of podocyte motility and survival signaling through interactions with proteins such as nephrin, podocin, anillin, and the p85 subunit of PI-3K. Disease-causing mutations in CD2AP can cause both autosomal dominant and autosomal recessive SRNS but are exceedingly rare (see Table 7.1 ). Pathogenic CD2AP mutations have also been associated with sporadic FSGS. Individuals with autosomal recessive SRNS manifest disease within the first year of life and show FSGS on renal biopsy. In mice, targeted deletion of cd2ap induced congenital nephrotic syndrome (CNS).

Mutations in crumbs homolog 2 (CRB2) have also been found to cause SRNS. CRB2 is a conserved regulator of epithelial cell polarity that is highly expressed in podocytes. CRB2 is a transmembrane protein that is involved in podocyte foot process arborization, slit diaphragm formation, and nephrin trafficking. Individuals with pathogenic CRB2 mutations manifest disease between 3 and 9 years of age and show histology consistent with FSGS (see Table 7.1 ). Targeted deletion of crb2b in zebrafish induced edema, foot process effacement, irregular podocyte morphology, and impaired apical basal patterning in podocytes.

Mutations in membrane-associated guanylate kinase, WW, and PDZ domain-containing 2 (MAGI2) have also been reported to cause familial and sporadic CNS. Patients with pathogenic MAGI2 mutations manifest disease in the first few months of life and show the histologic lesions of late-stage CNS and MCN on renal biopsy (see Table 7.1 ). MAGI2 is a multifunctional adaptor protein that is highly expressed in podocytes. MAGI2 interacts with nephrin in a heteromultimeric protein complex at the slit diaphragm and participates in the regulation of intracellular signaling and actin cytoskeletal dynamics. Patients with disease-causing MAGI2 mutations showed reduced and irregular glomerular nephrin and MAGI2 staining.

The nuclear compartment

Mutations in a variety of genes encoding nuclear pore proteins (NUPs) and karyopherins cause autosomal recessive SRNS. NUPs are a diverse group of almost 30 structural proteins that assemble within the bileaflet nuclear membrane to form channels known as nuclear pore complexes (NPCs). In humans, each NPC has a molecular mass of 120 to 125 MDa and is composed of 500 to 1000 NUPs. These complex macromolecular assemblies negotiate a broad array of functions including nucleocytoplasmic transport, chromatin organization, regulation of gene expression, and DNA repair. Karyopherins, also known as importins or exportins , are a superfamily of nuclear transport receptors that facilitate the translocation of proteins, RNAs, and ribonuclear particles across the NPC in a Ran GTP hydrolase-dependent process. In 2015 pathogenic mutations in NUP107 were reported to cause early-onset SRNS. Affected individuals ranged in age from 2 to 11 years at disease onset and all show findings consistent with FSGS on renal biopsy (see Table 7.1 ). In vivo, the targeted knockdown of NUP107 in zebrafish induced glomerular hypoplasia and irregular podocyte foot process architecture. In vitro, the heterozygous D831A mutation, which was common to all the affected individuals in a small Korean cohort, impaired NUP107 binding to NUP133 and disrupted the localization of NUP107 to the NPC. Subsequently, the frequency of biallelic NUP107 mutations in a small cohort of Korean children with SRNS was determined to be 44.4% in familial cases and 4.3% in sporadic cases.

Further evidence for the role of NUPs in the pathobiology of SRNS was provided in 2016 with the identification of novel disease-causing mutations in NUP93 , NUP205, and exportin 5 ( XP05 ). Affected individuals manifest disease between 1 and 6 years of age and the predominant histologic finding on renal biopsy was FSGS (see Table 7.1 ). NUP93 knockdown attenuated basal podocyte motility and proliferation, increased podocyte apoptosis and impaired NUP205 recruitment to the NPC. Reciprocally, the pathogenic NUP205 mutation abrogated NUP93 binding. Disease-causing NUP93 mutations impaired the assembly of intact NPCs, disrupted the interaction of NUP93 and XP05 with the transcriptional regulator SMAD4 and attenuated SMAD4 transcriptional activity. Disruptions in podocyte SMAD signaling are proposed as a potential mechanism of disease in these patients (see Table 7.1 ).

Wilms’ tumor 1 (WT1) is widely recognized for its multifaceted role in the pathobiology of renal disease. WT1 is a zinc-finger transcription factor that regulates the expression of a variety of genes that regulate podocyte function, viability, and differentiation. Mutations in WT1 can occur de novo or be inherited in an autosomal dominant or autosomal recessive fashion. Pathogenic mutations cause syndromic and nonsyndromic disease. Mutations associated with isolated SRNS most commonly occur in exons 8 and 9, which encode zinc-finger domains 2 and 3, respectively. Affected individuals with isolated autosomal recessive SRNS mutation manifest disease in early in childhood and the predominant histologic finding was DMS; however, FSGS can also be seen (see Table 7.1 ).

The mitochondria

Mutations in genes involved in the coenzyme Q10 (CoQ10) biosynthetic pathway cause syndromic and nonsyndromic forms of autosomal recessive SRNS. CoQ10 is an essential lipophilic antioxidant required for proper functioning of the mitochondrial electron transport chain. CoQ10 deficiency has been demonstrated to cause mitochondrial depolarization and increased apoptosis in podocytes. Pathogenic mutations in the aarF domain-containing kinase 4 ( ADCK4 ), an enzyme involved in CoQ10 biosynthesis, cause nonsyndromic autosomal recessive SRNS. Affected individuals manifest disease between <1 and 21 years of age and FSGS was the predominant histologic lesion on renal biopsy (see Table 7.1 ). ADCK4 is expressed in podocytes and localizes to mitochondria and foot processes. ADCK4 was found to interact with other enzymes of the CoQ10 biosynthetic pathway and individuals with disease-causing mutations were found to have reduced serum levels of CoQ10. Knockdown of adck4 caused nephrosis and podocyte foot process effacement in zebrafish. Further, ADCK4 knockdown in podocytes reduced podocyte motility; however, this effect was reversible with administration of CoQ10.

Pathogenic mutations in the Coenzyme Q2 COQ2 gene have also been identified as causes of syndromic and nonsyndromic SRNS. COQ2 is an enzyme in the CoQ10 biosynthetic pathway that catalyzes prenylation of parahydroxybenzoate, the second step in the final reaction sequence of CoQ biosynthesis. Affected individuals manifest disease in the first few months of life and renal histology can show features of classical FSGS, collapsing glomerulopathy, or crescentic glomerulonephritis (see Table 7.1 ). Electron microscopy can show a “wrinkled and folded” glomerular basement membrane and hyperplastic podocytes populated with numerous dysmorphic mitochondria.

Corticosteroid-resistant nephrotic syndrome is the hallmark renal injury of primary CoQ10 deficiency. It is often the initial manifestation of disease and can present as isolated kidney involvement or in association with other conditions such as encephalopathy, hypertrophic cardiomyopathy, retinopathy, optic atrophy, or sensorineural hearing loss. The genetic diagnosis of primary CoQ10 deficiency is made with identification of biallelic pathogenic variants in one of the nine genes encoding proteins directly involved in the synthesis of coenzyme Q10 (i.e., COQ2 , COQ4 , COQ6 , COQ7 , COQ8A , COQ8B [ADCK4] , COQ9 , PDSS1 , and PDSS2 ). Early diagnosis and treatment with high-dose oral CoQ10 supplementation (ranging from 5 to 50 mg/kg/day) have been shown to limit disease progression and reverse some manifestations; however, established severe kidney and/or neurologic damage cannot be reversed by CoQ10 supplementation. ACE inhibitors may be used in combination with CoQ10 supplementation in persons with proteinuria and kidney transplantation is an option for those with ESKD.

The actin cytoskeleton

The actin cytoskeleton plays a critical role in the regulation of podocyte size, shape, and adhesion. Monogenic causes of SRNS that effect actin-associated proteins or regulators of actin cytoskeletal dynamics are well described and have greatly informed our understanding of the essential role of the actin cytoskeleton in podocyte function. Myosin 1e ( Myo1e ) is a long-tail, class I (nonmuscle) myosin that participates in the organization of actin filaments at lamellipodia and filopodia to facilitate cell motility. Disease-causing mutations in Myo1e cause autosomal recessive SRNS. Affected individuals present in childhood with nephrotic-range proteinuria, microscopic hematuria, and FSGS on kidney biopsy (see Table 7.1 ). Partial remission can be achieved in some patients with cyclosporine treatment; however, some will progress to ESKD. Targeted deletion of Myo1e in mice induces proteinuria, podocyte injury, and CKD. Disease-causing mutations of Myo1e and transient gene knockdown cause mislocalization of the protein from plasma membrane puncta and impair podocyte motility in podocytes.

Members of the Rho family of small guanine nucleotide triphosphate hydrolases (GTPases) are widely recognized as critical regulators of actin cytoskeletal dynamics. In humans, the six known subfamilies of Rho GTP hydrolases (GTPases) are composed of more than 25 proteins encoded by multiple alternatively spliced gene transcripts. Dysregulated activity of the Rho GTPase family members RhoA, Cdc42, and Rac1 is implicated in the pathogenesis of both hereditary and idiopathic forms of NS. Rho GDP dissociation inhibitors (ARHGDIs) are a family of three evolutionarily conserved Rho GTPase interacting proteins that negatively regulate Rho GTPase activity by preventing the transition from the inactive GDP-bound state to the active GTP-bound state. Pathogenic mutations in ARHGDI-α ( ARHGDIA ) cause autosomal recessive CNS and SRNS. Affected individuals manifest disease between 14 days and 2.4 years of age and show DMS on kidney biopsy (see Table 7.1 ). Disease-causing ARHGDIA mutations abrogated interactions with Rho GTPases, increased active GTP-bound RAC1 and CDC42 and enhanced migration of cultured human podocytes. In vivo, knockdown of arhgdia reproduced the nephrotic phenotype.

Kidney ankyrin repeat-containing proteins (KANKs) are another class of Rho GTPase regulatory proteins that have been linked to the pathobiology of SRNS. The KANK family is composed of four evolutionarily conserved proteins (KANK1–4) that are all expressed in the kidney. KANKs negatively regulate the formation of actin stress fibers and cell migration through the inhibition of RhoA activity. Disease-causing mutations in KANK4 cause autosomal recessive SRNS. Affected individuals manifest disease at 2 months of age and show FSGS on kidney biopsy (see Table 7.1 ).

Phospholipase Cε1 ( PLCε1 ) is a ubiquitously expressed enzyme that catalyzes the hydrolytic cleavage of phosphoinositide 4,5 bisphosphate into diacylglycerol (DAG) and inositol triphosphate (IP3). Both DAG and IP3 regulate a diversity of intracellular processes, in part, through the activation of Ca ²⁺ -dependent signaling. Disease-causing mutations of PLCε1 cause autosomal recessive SRNS. Affected individuals manifest disease before 4 years of age and DMS is the predominant histologic finding on kidney biopsy; however, FSGS was also observed (see Table 7.1 ). Treatment with corticosteroids or cyclosporine A, an inhibitor of Ca ²⁺ -induced calcineurin/nuclear factor of activated T-cell (Cn/NFAT) signaling, stabilized disease in some cases but most progress to ESKD. PLC ε 1 colocalizes with the IQ motif-containing Rho GTPase activating protein (IQGAP) and they were shown to physically interact by coimmunoprecipitation. This interaction may be integral to the pathobiology of some PLCε1 mutations, because IQGAP has been shown to interact with slit diaphragm and cytoskeletal proteins, it is an activator of Rho GTPase actin cytoskeletal signaling, and it is negatively regulated by its association with calmodulin, a Ca ²⁺ -binding modulatory protein. In vivo, targeted knockdown of plce1 produced nephrosis in zebrafish but plce1 deletion in mice did not induce a kidney disease phenotype.

Glomerular epithelial protein 1 (GLEPP1), also known as Protein Tyrosine Phosphatase Receptor type O (PTPRO), is a receptor membrane protein tyrosine phosphatase located on the apical membrane of podocyte foot processes. Though the molecular functions of GLEPP1 in podocytes are not fully understood, it seems that GLEPP1 may play a role in the regulation of podocyte foot process architecture consistent with its known involvement in actin cytoskeletal and cell motility signaling. Disease-causing mutations in GLEPP1 cause autosomal SRNS. Affected individuals manifest disease between 7 and 14 years of age and kidney histology can show lesions consistent with FSGS or MCN (see Table 7.1 ). Partial remission was achieved with treatment regimens that included immunosuppression and renin-angiotensin system blockade, but some progressed to ESKD. In glepp1- deficient mice, electron microscopy revealed blunting and widening of podocyte foot processes in association with altered distribution of the podocyte intermediate cytoskeletal protein vimentin by electron microscopy. In association with these structural changes, these mice also had a reduced filtration surface area, which was confirmed by the finding of reduced glomerular nephrin content and reduced glomerular filtration rate.

The microtubular network

Tetratricopeptide repeat domain 21B ( TTC21B ) gene encodes an intraflagellar transport component-A, IFT139, which localizes to the base of the primary cilium in developing and undifferentiated cultured podocytes and along the extended microtubular network in mature podocytes. Disease-causing mutations in TTC21B cause autosomal recessive SRNS. Affected individuals manifest disease in the first through the fourth decades of life and can show FSGS or MCN with severe tubulointerstitial lesions on kidney biopsy (see Table 7.1 ). Targeted TTC21B knockdown induced ciliary structural defects, actin cytoskeletal abnormalities, and decreased motility in cultured podocytes. Expression of the disease-causing mutation partially rescued the phenotype suggesting a hypomorphic effect of the mutation.

The glomerular basement membrane

Pathogenic mutations in the complement factor H ( CFH ) gene cause membranoproliferative glomerulonephritis type II, also known as dense deposit disease . Deletion of a single amino acid (lysine 224) within the complement regulatory domain of CFH impaired its ability to bind and inactivate the complement component C3b resulting in complement-mediated destruction of the glomerular basement membrane. Affected individuals manifest disease in childhood through adolescence and about half will progress to ESKD within 10 years of symptom onset. Replacement of functional factor H by administration of fresh frozen plasma has been shown to be of benefit.

The slit diaphragm

In contrast to SRNS, there are fewer known genetic causes of SSNS. Although disease-causing mutations in nephrin are most commonly associated with CNF and SRNS, pathogenic nephrin mutations have been shown to cause childhood-onset SSNS with histologic lesions of DMS and MCN ( Table 7.2 ).

The actin cytoskeleton

As previously noted, some individuals with NS caused by pathogenic PLCε1 mutations received therapeutic benefit from corticosteroid therapy (see Table 7.1 ). In addition, pathogenic mutations in KANKs 1 and 2 cause SSNS (see Table 7.1 ).

Membrane trafficking

Pathogenic mutations of epithelial membrane protein 2 ( EMP2 ) have been identified as causes of SSNS. EMP2 is a member of the tetraspanin family of membrane scaffolding proteins that regulate a variety of cellular processes including cell morphology, motility, invasion, membrane fusion, membrane trafficking, and cell signaling. Tetraspanins alter the biologic and functional properties of their membrane protein partners by functioning as organizers of multimolecular membrane complexes. Disease-causing EMP2 mutations caused childhood-onset SSNS by 2.5 years of age. Notably, one case of SRNS was observed in a 3-year-old patient with MCN on kidney biopsy (see Table 7.1 ). In podocytes, EMP2 localized to the cell body and foot processes but did not colocalize with podocyte proteins such as synaptopodin. In addition, targeted knockdown of EMP2 attenuated podocyte proliferation and induced an increase in caveolin-1 expression consistent with the known roles of EMP2 in membrane trafficking and as a negative regulator of caveolin-1 transcription. Expression of caveolin-1 was also increased in podocytes expressing the disease-causing truncating EMP2 mutation. In vivo, emp2 knockdown recapitulated the nephrosis phenotype.

Disorders of lipid metabolism

Lecithin-cholesterol acyltransferase ( LCAT ) deficiency is an autosomal recessive disease characterized by anemia, corneal opacities, low serum HDL, proteinuria, and often NS with progressive kidney failure. Affected individuals often present in adulthood and show irregular thickening of the glomerular capillaries with vacuolization of the capillary basement membrane due to lipid droplet deposition. Electron-dense lamellar aggregates in the mesangium and basement membrane are characteristic diagnostic features of the disease ( Table 7.3 ). Currently, there is no specific therapy for familial LCAT deficiency.

Pathogenic mutations of apolipoprotein E ( APOE ) are associated with lipoprotein glomerulopathy, a disease characterized by type III hyperlipidemia and proteinuria often with a rapid progression to ESKD. Lipoprotein glomerulopathy is generally considered to be a hereditary disease largely because of the early studies of the disease in a number of Japanese kindreds. Affected individuals usually present in adulthood and show characteristic deposition of laminated lipid droplets containing apolipoproteins A, B, and E in the glomerular capillaries. Intensive lipid-lowering therapy has been shown to be effective in reversing glomerular lipid deposition and reducing proteinuria (see Table 7.3 ).

Two independent sequence variants in the apolipoprotein L1 ( APOL1 ) gene have been associated with an increased risk for the development of FSGS and hypertension-attributed ESKD in people of African descent. APOL1 is a serum factor that lyses trypanosomes and the two sequence variants enhance the trypanolytic activity of APOL1 in vitro. Although APOL1 has not been established as a cause of FSGS, investigations into potential pathogenic mechanisms are ongoing.

The slit diaphragm

As previously noted, pathogenic mutations in CD2AP can cause autosomal dominant SRNS. Affected individuals manifest disease in adulthood and show histologic lesions of FSGS ( Table 7.4 ). Pathogenic mutations in anillin ( ANLN ), a critical binding partner of CD2AP at the slit diaphragm, have also been shown to cause autosomal dominant FSGS. Anillin is a ubiquitously expressed nucleocytoplasmic scaffolding protein that regulates various intracellular functions including cytokinesis, cell proliferation, cell motility, F-actin bundling, and cell-survival signaling. Anillin derives its name from the Spanish word for ring, anillo , given its prominent role in the formation of the cytokinetic contractile ring; however, its nonmitotic functions are highly dependent on interactions with a diversity of cytoskeletal proteins and signaling intermediates. In podocytes, anillin localizes to the slit diaphragm where it binds CD2AP at a conserved N-terminal conserved Px(P/A)xxR motif. Anillin was shown to colocalize with CD2AP to peripheral membrane blebs in migratory podocytes and disease-causing mutations in anillin disrupted the anillin–CD2AP interaction. Affected individuals typically manifest disease in the second and third decades of life and show FSGS on kidney biopsy (see Table 7.4 ). Anillin expression is upregulated in the podocytes of patients with collapsing FSGS and in experimental models of HIV-associated nephropathy. In addition, overexpression of anillin induced hypermotility in podocytes. In vivo, targeted knockdown of anln induced nephrosis and irregular foot process architecture in zebrafish.

Disease-causing mutations in the transient receptor potential cation channel, subfamily C, isotype 6 ( TRPC6 ) were first recognized to cause autosomal dominant FSGS in 2005, and since then, more than 20 other pathogenic mutations have been reported. TRPC6 is a receptor operated, nonselective cation channel with a high affinity for Ca ²⁺ . TRPC6 is directly activated by DAG and induces its own gene expression through the activation of Ca ²⁺ -induced Cn/NFAT signaling. TRPC6 localizes to podocyte foot processes where it interacts with proteins such as podocin and nephrin in multimolecular complexes at the slit diaphragm. Dysregulated TRPC6-mediated Ca ²⁺ conductance is thought to induce podocyte injury through alterations in actin cytoskeletal dynamics and derangement of foot process architecture. Most of the disease-causing TRPC6 mutations exert “gain-of-function” effects, inducing an increase in Ca ²⁺ conductance through the channel; however, “loss-of-function” mutations have also been reported. Affected individuals usually manifest in the third and fourth decades of life although some cases of early-onset childhood SRNS have been documented. It is estimated that as many as 60% of affected individuals will progress to ESKD within 10 years of disease onset (see Table 7.4 ).

In addition to its role in familial FSGS, TRPC6 has been implicated in the pathobiology of other forms of acquired proteinuric kidney disease such as MCN, membranous nephropathy, and diabetic kidney disease. Podocyte injury in these diseases is thought to result, in part, from an inappropriate upregulation of TRPC6 expression and activity. Although there are no selective TRPC6 inhibitors currently in clinical use, these findings suggest that therapies targeting reduced TRPC6 expression may be of therapeutic value. This hypothesis is supported by a growing number of animal studies that show amelioration of TRPC6-induced glomerular injury with pharmacologic or molecular genetic inhibition of TRPC6 expression.

The nuclear compartment

In addition to its role in autosomal recessive and sporadic forms of SRNS, pathogenic mutations in WT1 have been found to cause nonsyndromic autosomal dominant FSGS. Affected individuals manifest disease in adolescence through adulthood, and it is thought that the mechanism of disease involves podocyte injury through downregulation of essential slit diaphragm and actin cytoskeletal proteins and increased apoptosis (see Table 7.4 ).

Similar to WT1, Lin-11, Isi-1, and Mec3 (LIM)-homeobox-1β (LMX1β) is another nuclear transcription factor that regulates many facets of kidney development and differentiation. In podocytes, LMX1β plays an essential role in the formation of the actin cytoskeleton, slit diaphragm, and glomerular basement membrane through the transcriptional regulation of genes such as NPHS1 , NPHS2 , CD2AP , GLEPP1 , collagen IV α3 ( COL4A3 ), and collagen IV α4( COL4A4 ). In addition, LMX1β differentially regulates the transcription of WT1 isoforms to selectively modulate WT1-mediated gene regulation. Pathogenic mutations in LMX1β can cause syndromic kidney disease (i.e., Nail Patella syndrome [NPS]) or isolated FSGS without extrarenal manifestations (i.e., Nail Patella–Like Renal disease [NPLRD]). Pathogenical mutations of the LIM homeodomain cause NPLRD. Affected individuals manifest disease in adolescence through adulthood and the kidney histology can vary widely (see Table 7.4 ).

The Paired Box family of nuclear transcription factors is composed of the principal regulators of cellular lineage determination through the maintenance and specification of progenitor cells during embryonic development. PAX2 is essential for the development of the kidney epithelial cells and pathogenic mutations in PAX2 cause kidney coloboma syndrome in childhood. In adults, pathogenic mutations in PAX2 have been found to be associated with isolated autosomal dominant FSGS. Affected individuals manifest disease from late adolescence through adulthood and show histologic lesions consistent with FSGS on kidney biopsy (see Table 7.4 ). Functional analysis of FSGS-associated variants revealed that PAX2 variants perturbed protein function by disrupting its DNA binding and transactivation functions or by altering the interaction of PAX2 with repressor proteins, resulting in enhanced repressor activity.

The actin cytoskeleton

α-Actinin-4 ( ACTN4 ) is one member of a family of four actin-bundling proteins (ACTN1–4) that plays an essential role in podocyte foot process architecture and adhesion. α-Actinin-4 is a Ca ²⁺ -responsive, nonmuscle myosin that localizes to F-actin bundles with synaptopodin and MAGI1. Mutations in ACTN4 cause autosomal dominant and sporadic SRNS by a mechanism that involves impaired relaxation of F-actin–ACTN4 cross-bridges, which results in dysregulation of podocyte foot process architecture and function. Interestingly, this cytoskeletal rigidity only manifests in podocyte dysfunction and glomerular sclerosis despite the broad tissue expression of ACTN4. This highlights the essential role of the actin cytoskeletal network for proper podocyte function. Affected individuals manifest disease in adolescence, and some progress to ESKD. Kidney histology shows areas of podocyte foot process effacement and lesions consistent with FSGS (see Table 7.4 ). Similarly, mice expressing a disease-causing ACTN4 mutation developed proteinuria and exhibited histologic features consistent with human FSGS including segmental sclerosis, tuft adhesion, tubular dilation, mesangial matrix expansion, podocyte vacuolization, and foot process fusion.

Inverted formin 2 ( INF2 ), also known as WH-domain containing formin 1 (WHIF1), is an actin-associated protein that binds to actin monomers to accelerate polymerization or depolymerization of F-actin filaments. In podocytes, INF2 negatively regulates lamellipodial actin dynamics and the trafficking of slit diaphragm proteins by opposing Rho/mDia-mediated actin polymerization. Pathogenic mutations in INF2 have been identified as causes of autosomal dominant FSGS. Notably, pathogenic INF2 mutations account for 9% to 16% of familial FSGS making it the most common genetic cause of the disease. Affected individuals can manifest disease in adolescence through adulthood (see Table 7.4 ). Interestingly, mice expressing a disease-causing mutation in INF2 showed no significant kidney pathology or proteinuria at baseline despite diminished INF2 protein levels. However, INF2 mutant podocytes showed impaired reversal of protamine sulfate-induced foot process effacement by heparin sulfate perfusion.

Rho GTPase-activating proteins (ARHGAPs) negatively regulate Rho GTPases by accelerating the hydrolytic conversion of GTP to GDP. Disease-causing mutations in ARHGAP24 cause autosomal dominant FSGS. Affected individuals manifest disease in the second and third decades of life and show FSGS on kidney biopsy (see Table 7.4 ). ARHGAP24 knockdown in podocytes increased Rac1 and CDC42 activity, podocyte motility, and membrane ruffling Similarly, a disease-causing ARHGAP24 mutation demonstrated diminished Rac1-GAP activity in HEK 293 cells.

The glycocalyx

Mature and healthy podocytes are covered by a polyanionic mucinous coating of sulfated molecules and sialated glycoproteins referred to as the glycocalyx . The components of this protective coating are synthesized and secreted by podocytes and are thought to act as an antiadhesin, which maintains the patency of the filtration slits between adjacent podocytes through electrostatic repulsion. Podocalyxin ( PODXL ) is the major sialoglycoprotein of the glycocalyx and disease-causing variants in PODXL are associated with autosomal dominant FSGS. Affected individuals presented in early adolescence though adulthood with proteinuria and FSGS on kidney biopsy (see Table 7.4 ). Notably, an FSGS-associated PODXL variant induced podocalyxin dimerization but did not alter protein stability, extracellular domain glycosylation, cell-surface expression, global subcellular localization, or interaction with its intracellular binding partner, ezrin.

The nuclear compartment

The SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a like 1 ( SMARCAL1 ) is an ATP-dependent annealing helicase that assists in the repair of damaged DNA replication forks to promote genome stability during replicative stress. SMARCAL1 is ubiquitously expressed and localizes to the nuclei of glomerular podocytes, tubular cells, and endothelial cells. Pathogenic mutations in SMARCAL1 cause Schimke immuno-osseous dysplasia (SIOD). SIOD is a rare autosomal recessive multisystem disorder characterized by disproportionate growth failure, impaired T-cell function, and SRNS. Affected individuals manifest in childhood and adolescence and the predominant histologic finding is FSGS ( Table 7.5 ). Although the mechanisms of disease remain unclear, increased DNA fragmentation and dysregulated gene expression have been observed in human tissue and experimental models of SMARCAL1 deficiency.

TABLE 7.5

Syndromic Proteinuric Kidney Diseases (Autosomal Recessive)

Gene	Protein	Affected Podocyte Compartment or Function	Clinical Presentation	Histologic Features	Mechanism of Disease
SMARCAL1	SMARCAL1	Nucleus	Schimke immuno-osseous dysplasia (SIOD), disease manifests in childhood through adolescence, disproportionate growth failure, impaired T-cell function, SRNS	FSGS	Chromatin instability and dysregulated gene expression
WT1	WT1	Nucleus	DDS and Frasier syndromes, congenital nephrotic syndrome, DDS typically progresses to ESKD within 3 years, Frasier syndrome manifests slowly over a decade.	DMS with DDS, FSGS with Frasier syndrome	Impaired transactivation of various developmental genes of the kidney and urogenital tract
DGKE	DGKε	Actin cytoskeleton	aHUS, SRNS	Foot process effacement (rare)	Dysregulated Ca 2+ -dependent actin cytoskeletal dynamics
SGLP1	SGLP1	Actin cytoskeleton	Adrenal insufficiency, congenital nephrotic syndrome, SRNS, disease manifests in the early postnatal period	FSGS	Dysregulated actin cytoskeletal dynamics
WDR73	WDR73	Microtubules	Galloway–Mowat syndrome, disease manifests in childhood	FSGS	Dysregulated microtubular dynamics
FAT1	FAT1	Slit diaphragm	SRNS with tubular ectasia, hematuria, and facultative neurologic involvement, disease manifests in early childhood through late adolescence	Glomerulotubular nephropathy	Disruption of the podocyte slit diaphragm
EXT1	Exostosin-1	Glomerular basement membrane	Glomerular sclerosis with multiple exostoses, disease manifests in childhood through adulthood	Glomerular sclerosis with abundant fibrillary collagen deposits	Disruption of glomerular basement membrane composition and function
ITGA3 and ITGB4	ITGα3 and ITGβ4	Adhesion to the glomerular basement membrane	Disease manifests in the first year of life	FSGS	Impaired adhesion to the glomerular basement membrane
CD151	CD151	Glomerular basement membrane	Disease manifests in adulthood	Classical changes of Alport syndrome without COL4A5 mutations	Impaired formation of the glomerular and tubular basement membranes
LAMB2	LAMB2	Glomerular basement membrane	Pierson syndrome, disease manifests in the early postnatal period	Glomerular sclerosis, DMS	Impaired formation of the glomerular basement membrane
ZMPSTE24	ZMPSTE24	Glomerular basement membrane	Disease manifests in adulthood	FSGS	Impaired formation of the glomerular basement membranes
COQ6	COQ6	Mitochondria	Disease manifests in first decade of life, SRNS and sensorineural hearing loss; responsive to CoQ10 supplementation	FSGS	Impaired CoQ10 biosynthesis
PDSS2	PDSS2	Mitochondria	Disease manifests in the early postnatal period, Leigh syndrome and SRNS	Renal tubulopathy or diffuse glomerulocystic kidney damage	Impaired CoQ10 biosynthesis
MTTL1	MTTL1	Mitochondria	Disease manifests in childhood through adulthood, MELAS syndrome with nephropathy, A3243G mutation particularly associated with adult-onset FSGS	FSGS	Impaired mitochondrial function
SCARB2	SCARB2	Membrane trafficking	Disease manifests in the second decade of life, action myoclonus-renal failure syndrome, substrate removal therapy targeting reduction of glucocerebrosidase accumulation found to be effective in one patient	C1q nephropathy	Impaired lysosomal formation and function
CUBN	CUBN	Membrane trafficking	Disease manifest in the first decade of life; intermittent proteinuria and megaloblastic anemia secondary to vitamin B 12 deficiency	Unknown	Impaired protein reuptake from glomerular filtrate
ALG1	ALG1	Glycosylation	Disease manifests in the early postnatal period, congenital nephrotic syndrome with congenital disorder of glycosylation	FSGS	Impaired N-glycosylation and targeting of slit diaphragm proteins
PMM2	PMM2	Glycosylation	Disease manifests in the early postnatal period, Jaeken syndrome with congenital nephrotic syndrome	DMS with focal tubular dilation	Impaired N-glycosylation and targeting of slit diaphragm proteins

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