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Neurodegeneration with brain iron accumulation (NBIA) involves a group of rare neurogenetic disorders often linked with iron overload in the basal nuclei of the brain presenting with spasticity, dystonia, muscle rigidity, neuropsychiatric symptoms, and retinal degeneration. Among NBIA subtypes, beta-propeller-protein-associated neurodegeneration (BPAN) is associated with mutations in the autophagy gene WDR45 (WD repeat domain 45). Previously, we demonstrated that WDR45 mutations in BPAN cellular models impaired autophagy, iron metabolism, and cell bioenergetics. In addition, antioxidant supplementation partially improved cell physiopathology; however, autophagy and cell bioenergetics remained affected. In this work, we explored the possibility of expressing the normal WDR45 allele present in the inactive chromosome X (Xi) of BPAN cells through treatment with epigenetic modulators. The aim of this study was to demonstrate whether biotin, an epigenetic nutrient, was able to restore the expression levels of WDR45 by a mechanism involving Xi reactivation and, consequently, correct BPAN defects. Our study demonstrated that biotin supplementation increases histone biotinylation and allows for the transcription of the WDR45 allele in Xi. Consequently, all physiopathological alterations in BPAN cells were notably corrected. The reactivation of Xi by epigenetic modulators can be a promising approach for the treatment of BPAN and other X-linked diseases. [ABSTRACT FROM AUTHOR]

Abstract Background β-Propeller protein-associated neurodegeneration (BPAN) is a genetic neurodegenerative disease caused by mutations in WDR45. The impairment of autophagy caused by WDR45 deficiency contributes to the pathogenesis of BPAN; however, the pathomechanism of this disease is largely unknown. Lipid dyshomeostasis is involved in neurogenerative diseases, but whether lipid metabolism is affected by Wdr45 deficiency and whether lipid dyshomeostasis contributes to the progression of BPAN are unclear. Methods We generated Wdr45 knockout SN4741 cell lines using CRISPR‒Cas9-mediated genome editing, then lipid droplets (LDs) were stained using BODIPY 493/503. Chaperone-mediated autophagy was determined by RT-qPCR and western blotting. The expression of fatty acid synthase (Fasn) was detected by western blot in the presence or absence of the lysosomal inhibitor NH4Cl and the CMA activator AR7. The interaction between Fasn and HSC70 was analyzed using coimmunoprecipitation (Co-IP) assay. Cell viability was measured by a CCK-8 kit after treatment with the Fasn inhibitor C75 or the CMA activator AR7. Results Deletion of Wdr45 impaired chaperone-mediated autophagy (CMA), thus leading to lipid droplet (LD) accumulation. Moreover, Fasn can be degraded via CMA, and that defective CMA leads to elevated Fasn, which promotes LD formation. LD accumulation is toxic to cells; however, cell viability was not rescued by Fasn inhibition or CMA activation. Inhibition of Fasn with a low concentration of C75 did not affect cell viability but decreases LD density. Conclusions These results suggested that Fasn is essential for cell survival but that excessive Fasn leads to LD accumulation in Wdr45 knockout cells.

Background: β-Propeller protein-associated neurodegeneration (BPAN) is a genetic neurodegenerative disease caused by mutations in WDR45. The impairment of autophagy caused by WDR45 deficiency contributes to the pathogenesis of BPAN; however, the pathomechanism of this disease is largely unknown. Lipid dyshomeostasis is involved in neurogenerative diseases, but whether lipid metabolism is affected by Wdr45 deficiency and whether lipid dyshomeostasis contributes to the progression of BPAN are unclear. Methods: We generated Wdr45 knockout SN4741 cell lines using CRISPR‒Cas9-mediated genome editing, then lipid droplets (LDs) were stained using BODIPY 493/503. Chaperone-mediated autophagy was determined by RT-qPCR and western blotting. The expression of fatty acid synthase (Fasn) was detected by western blot in the presence or absence of the lysosomal inhibitor NH4Cl and the CMA activator AR7. The interaction between Fasn and HSC70 was analyzed using coimmunoprecipitation (Co-IP) assay. Cell viability was measured by a CCK-8 kit after treatment with the Fasn inhibitor C75 or the CMA activator AR7. Results: Deletion of Wdr45 impaired chaperone-mediated autophagy (CMA), thus leading to lipid droplet (LD) accumulation. Moreover, Fasn can be degraded via CMA, and that defective CMA leads to elevated Fasn, which promotes LD formation. LD accumulation is toxic to cells; however, cell viability was not rescued by Fasn inhibition or CMA activation. Inhibition of Fasn with a low concentration of C75 did not affect cell viability but decreases LD density. Conclusions: These results suggested that Fasn is essential for cell survival but that excessive Fasn leads to LD accumulation in Wdr45 knockout cells. [ABSTRACT FROM AUTHOR]

The four human WIPI β‐propellers, WIPI1 through WIPI4, belong to the ancient PROPPIN family and fulfill scaffold functions in the control of autophagy. In this context, WIPI β‐propellers function as PI3P effectors during autophagosome formation and loss of WIPI function negatively impacts autophagy and contributes to neurodegeneration. Of particular interest are mutations in WDR45, the human gene that encodes WIPI4. Sporadic WDR45 mutations are the cause of a rare human neurodegenerative disease called BPAN, hallmarked by high brain iron accumulation. Here, we discuss the current understanding of the functions of human WIPI β‐propellers and address unanswered questions with a particular focus on the role of WIPI4 in autophagy and BPAN. [ABSTRACT FROM AUTHOR]

Neurodegeneration with brain iron accumulation (NBIA) is a group of rare neurogenetic disorders frequently associated with iron accumulation in the basal nuclei of the brain. Among NBIA subtypes, β-propeller protein-associated neurodegeneration (BPAN) is associated with mutations in the autophagy gene WDR45. The aim of this study was to demonstrate the autophagic defects and secondary pathological consequences in cellular models derived from two patients harboring WDR45 mutations. Both protein and mRNA expression levels of WDR45 were decreased in patient-derived fibroblasts. In addition, the increase of LC3B upon treatments with autophagy inducers or inhibitors was lower in mutant cells compared to control cells, suggesting decreased autophagosome formation and impaired autophagic flux. A transmission electron microscopy (TEM) analysis showed mitochondrial vacuolization associated with the accumulation of lipofuscin-like aggregates containing undegraded material. Autophagy dysregulation was also associated with iron accumulation and lipid peroxidation. In addition, mutant fibroblasts showed altered mitochondrial bioenergetics. Antioxidants such as pantothenate, vitamin E and α-lipoic prevented lipid peroxidation and iron accumulation. However, antioxidants were not able to correct the expression levels of WDR45, neither the autophagy defect nor cell bioenergetics. Our study demonstrated that WDR45 mutations in BPAN cellular models impaired autophagy, iron metabolism and cell bioenergetics. Antioxidants partially improved cell physiopathology; however, autophagy and cell bioenergetics remained affected. [ABSTRACT FROM AUTHOR]

Beta-propeller protein-associated neurodegeneration (BPAN) is a subtype of neurodegeneration with brain iron accumulation (NBIA) that presents with childhood developmental delay (especially speech delay), occasionally associated with epileptic encephalopathy, autism, or Rett-like syndrome. The majority of children described to date have been severely affected, with little to no expressive speech function, severe developmental delay, and cognitive impairment. Herein, five additional patients with BPAN identified in the same center in Canada are described, four with the typical severe phenotype and one with a milder phenotype. Our findings provide further evidence that a spectrum of severity exists for this rare and newly described condition. Challenges in identifying iron accumulation on brain MRI are also addressed. Additionally, the importance of including the WDR45 gene on epilepsy and Rett-like syndrome genetic panels is highlighted.

Ferritin is the main iron storage protein that plays a pivotal role in the regulation of iron homeostasis. Mutations in the autophagy protein WD repeat domain 45 (WDR45) that lead to iron overload is associated with the human β-propeller protein-associated neurodegeneration (BPAN). Previous studies have demonstrated that ferritin was decreased in WDR45 deficient cells, but the mechanism remains unclear. In this study, we have demonstrated that the ferritin heavy chain (FTH) could be degraded via chaperone-mediated autophagy (CMA) in ER stress/p38-dependent pathway. In HeLa cells, inducing the ER stress activated CMA, therefore facilitated the degradation of FTH, and increased the content of Fe2+. However, the increased CMA activity and Fe2+ as well as the decreased FTH by ER stress inducer were restored by pre-treatment with p38 inhibitor. Overexpression of a mutant WDR45 activated CMA thus promoted the degradation of FTH. Furthermore, inhibition of ER stress/p38 pathway resulted in reduced activity of CMA, which consequently elevated the protein level of FTH but reduced the Fe2+ level. Our results revealed that WDR45 mutation dysregulates iron homeostasis by activating CMA, and promotes FTH degradation through ER stress/p38 signaling pathway. [Display omitted] • FTH is a substrate for CMA. • WDR45 mutation activates CMA in an ER-stress/p38 dependent manner. • CMA activation facilitates the degradation of FTH and increased the content of Fe2+. • CMA is a potential therapeutic target for BPAN treatment. [ABSTRACT FROM AUTHOR]

Beta-propeller protein-associated neurodegeneration (BPAN) is a rare neurodegenerative disorder characterized by iron accumulation in the brain with spectrum of neurodevelopmental and movement phenotypes. In anticipation of future clinical trials and to inform clinical care, there is an unmet need to capture the phenotypic diversity of this rare disorder and better define disease subtypes. A total of 27 individuals with BPAN were included in our natural history study, from which traditional outcome measures were obtained in 18 subjects. Demographic and diagnostic information, along with acquisition of basic developmental skills and overall neurologic severity were extracted from the medical records. Functional outcome measures were administered at the time of the evaluation or applied retrospectively at the last clinical encounter for patients who were not able to travel for in person. Based on age and functional level, the following assessments were administered: Leiter-3, Gross Motor Function Measure (GMFM)-66 Item Sets, Vineland-3, and Peabody-2. Overall, cognitive function was more impaired compared to gross motor function. Onset of symptoms of BPAN within the first 6 months of life was associated with decreased gain of ambulation and gain of spoken language (ambulation: log-rank test p = 0.0015; gain of first word: p = 0.0015). There was no difference in age at seizure onset by age at initial symptom onset (p = 0.8823). Collection of prospective outcome measures was limited by attention and behavior in our patient population, reinforcing the complexity of phenotype assessment and inadequacy of available standardized tests. Overall, gross motor and adaptive behavior assessments were better able to capture the dynamic range of function across the BPAN population than the fine motor and non-verbal cognitive tests. Floor effects were noted across outcome measures in a subset of individuals for cognitive and adaptive behavior tests. Our data suggest the distinct phenotypes of BPAN: a severe, early onset form and an attenuated form with higher cognitive capabilities. Early age at onset was a key factor in predicting future neurologic impairment. [ABSTRACT FROM AUTHOR]

Beta-propeller protein-associated neurodegeneration (BPAN) is a subtype of neurodegeneration with brain iron accumulation (NBIA) caused by loss-of-function variants in WDR45. The underlying mechanism of iron accumulation in WDR45 deficiency remains elusive. We established a primary skin fibroblast culture of a new BPAN patient with a missense variant p.(Asn61Lys) in WDR45 (NM_007075.3: c.183C>A). The female patient has generalized dystonia, anarthria, parkinsonism, spasticity, stereotypies, and a distinctive cranial MRI with generalized brain atrophy, predominantly of the cerebellum. For the functional characterization of this variant and to provide a molecular link of WDR45 and iron accumulation, we looked for disease- and variant-related changes in the patient's fibroblasts by qPCR, immunoblotting and immunofluorescence comparing to three controls and a previously reported WDR45 patient. We demonstrated molecular changes in mutant cells comprising an impaired mitochondrial network, decreased levels of lysosomal proteins and enzymes, and altered autophagy, confirming the pathogenicity of the variant. Compared to increased levels of the ferritinophagy marker Nuclear Coactivator 4 (NCOA4) in control cells upon iron treatment, patients' cells revealed unchanged NCOA4 protein levels, indicating disturbed ferritinophagy. Additionally, we observed abnormal protein levels of markers of the iron-dependent cell death ferroptosis in patients' cells. Altogether, our data suggests that WDR45 deficiency affects ferritinophagy and ferroptosis, consequentially disturbing iron recycling. [ABSTRACT FROM AUTHOR]

WDR45 is an autophagy-related protein that involves in the formation of autophagosome. Mutations in WDR45 lead to the impairment of autophagy which is associated with the human β-propeller protein-associated neurodegeneration (BPAN). However, the relationship between autophagy and brain iron accumulation in patients with BPAN remains unclear. Here, we demonstrated that transferrin receptor (TfRC) which is critical for the iron import of cells was degraded via autophagy. TfRC was accumulated after the inhibition of autophagy by treatment with autophagic inhibitor chloroquine or knockdown of ATG2A. The intracellular iron content was increased in cells overexpressing TfRC or mutant WDR45, however, ferritin H (FTH) chain was decreased. Increased TfRC and simultaneously decreased FTH consequently resulted in an elevated level of ferrous iron (Fe2+) which further promoted cell ferroptosis, demonstrated by the increased lipid peroxidation and reactive oxygen species (ROS) and the decreased glutathione peroxidase 4 (GPX4) and cell viability. Taken together, these findings provide a piece of important evidence that WDR45 deficiency impairs autophagic degradation of TfRC, therefore leading to iron accumulation, and the elevated iron promotes ferroptosis which may contribute to the progression of BPAN.

Abstract Background Beta‐propeller protein‐associated neurodegeneration (BPAN, OMIM 300894) is an X‐linked neurodegenerative disorder caused by mutations in WDR45. WDR45 is required for autophagy, defect in WDR45 impaired autophagy which contributes for the pathogenesis of BPAN. Previously, we reported a novel de novo mutation (c.1040_1041del, p.Glu347GlyfsTer7) in WDR45 (NM_007075) in a 3‐year‐old Chinese girl with BPAN. Methods The protein structure was constructed using SWISS‐MODEL and the isoelectric point (pI) was predicted by the online pI/Mw tool at ExPASy. The functional effects of this mutation were predicted by two online software programs: PROVEN and MutationTaster. Stable overexpression of Flag‐tagged wild‐type or mutant WDR45 in HeLa cells was constructed. Protein levels of LC3 and p62 were analyzed by western blot upon treatment with/without autophagy inhibitor Bafilomycin A1, the formation of LC3 puncta were analyzed in HeLa cells transfected with mCherry‐LC3 by confocal microscopy. Results The mutation resulted in a shift of pI from 6.74 to 8.84 and was predicted to be pathogenic. The protein levels of LC3‐II and p62 were increased in cells overexpression of wild‐type and mutant WDR45 while the protein levels were not increased in cells overexpression of mutant WDR45 upon treatment with autophagy inhibitor Bafilomycin A1. Results from confocal microscopy revealed that LC3‐positive puncta were increased in cells expressing both wild‐type and mutant WDR45 while the number of LC3‐positive puncta was not increased in cells expressing mutant WDR45 upon treatment with Bafilomycin A1. Conclusion Our study evidenced that this novel mutation in WDR45 impaired autophagy in cells thus this mutation is the cause for BPAN in this patient.

Introduction Beta-propeller protein-associated neurodegeneration (BPAN) is a very rare, X-linked dominant (XLD) inherited member of the neurodegeneration with brain iron accumulation (NBIA) disease family. Case report We present a female case of BPAN with infantile spasms in the first year, Rett-like symptomatology, focal epilepsy, and loss of motor skills in childhood. Menarche occurred at the age of 9, after precocious pubarche and puberty. Dystonia-parkinsonism as extrapyramidal sign at the age of 10 years resulted in radiological and genetic work-up. Results Burke–Fahn–Marsden Dystonia Rating Scale (BFMDRS) measured 66/120 points in body part-related dystonia symptoms. Cerebrospinal fluid examination showed dopamine depletion. T2 and B0 sequences of the diffusion-weighted magnetic resonance imaging showed susceptibility artifacts with NBIA-typical hypointense globus pallidus (GP) and substantia nigra (SN). Next-generation sequencing revealed a BPAN-causing pathogenic variant in WDR45 (WD repeat-containing protein 45) gene (c.830 + 1G > A, XLD, heterozygous, de novo). Skewed X-inactivation was measured (2:98). Conclusions Autophagy-related X-linked BPAN disease might still be underdiagnosed in female cases of infantile spasms. Skewed X-inactivation will have mainly influenced the uncommon, very early childhood neurodegenerative symptomatology in the present BPAN case. Oral levodopa substitution led to improvement in sleep disorder, hypersalivation, and swallowing. Reduced white matter and hypointense signals in SN and GP on susceptibility sequences in magnetic resonance imaging are characteristic radiological findings of advanced disease in NBIA. No BPAN-typical halo sign in T1-weighted scan at midbrain level was seen at the age of 11 years. NBIA panel is recommended for early diagnosis. [ABSTRACT FROM AUTHOR]

Beta-propeller protein-associated neurodegeneration (BPAN) is an X-linked rare dominant disorder of autophagy. The role of WDR45 has been implicated in BPAN almost exclusively in females possibly due to male lethality. Characterization of distinctive clinical manifestations and potentially the complex genetic determinants in rare male patients remain crucial for deciphering BPAN and other X-linked dominant diseases. We performed whole exome sequencing (WES) followed by segregation analysis and identified a novel nonsense and mosaic variant in WDR45, namely NM_007075.3:c.873C>G; p.(Tyr291*) in an affected male at the age of 34. His biphasic medical history was compatible with BPAN, which was characterized by delayed psychomotor development, intellectual disability, and progression into dystonia parkinsonism in his twenties. The variant had an apparently mosaic pattern both in whole exome and Sanger sequencing findings. In order to figure out if mosaicism was restricted to this variant or related to a chromosomal level mosaicism, we used our in-house WES data from 129 unrelated individuals to calculate the threshold values of male and female X chromosome heterozygosity (XcHet) in WES data for our pipeline. A background level of heterozygous variants on X chromosome excluding the pseudoautosomal loci is an observed phenomenon in WES analysis and this level has been used as a quality measure. Herein, we suggest utilization of this measure for detection of digital anomalies of the X chromosome in males by potentially observing a higher XcHet value than the threshold value. This approach has revealed a variant level mosaicism in the affected male, which was further supported with cytogenetic analyses. [ABSTRACT FROM AUTHOR]

Mutations in the WDR45 gene have been identified as causative for the only X-linked type of neurodegeneration with brain iron accumulation (NBIA), clinically characterized by global developmental delay in childhood, followed by a secondary neurological decline with parkinsonism and/or dementia in adolescence or early adulthood. Recent reports suggest that WDR45 mutations are associated with a broader phenotypic spectrum. We identified a novel splice site mutation (c.440-2 A > G) in a 5-year-old Argentinian patient with Rett-like syndrome, exhibiting developmental delay, microcephaly, seizures and stereotypic hand movements, and discuss this finding, together with a review of the literature. Additional patients with a clinical diagnosis of Rett (-like) syndrome were also found to carry WDR45 mutations before (or without) clinical decline or signs of iron accumulation by magnetic resonance imaging (MRI). This information indicates that WDR45 mutations should be added to the growing list of genetic alterations linked to Rett-like syndrome. Further, clinical symptoms associated with WDR45 mutations ranged from early-onset epileptic encephalopathy in a male patient with a deletion of WDR45 to only mild cognitive delay in a female patient, suggesting that analysis of this gene should be considered more often in patients with developmental delay, regardless of severity. The increasing use of next generation sequencing technologies as well as longitudinal follow-up of patients with an early diagnosis will help to gain additional insight into the phenotypic spectrum associated with WDR45 mutations. [ABSTRACT FROM AUTHOR]

Introduction: Beta-propeller protein associated neurodegeneration (BPAN) is associated with mutations in the WD repeat domain 45 (WDR45) gene on chromosome Xp11 resulting in reduced autophagic flux. This study describes the clinical and neuropathological features of a female 51 year old BPAN case. The clinical history includes learning disability and progressive gait abnormalities since childhood followed by progressive dystonic features in young adulthood. Brain imaging revealed generalised brain atrophy and bilateral mineralisation of the globus pallidus and substantia nigra. Results: The major pathological findings were observed in the substantia nigra with excess iron deposition, gliosis, axonal swellings and severe neuronal loss. Iron deposition was also observed in the globus pallidus. There was extensive hyperphosphorylated-tau deposition in the form of neurofibrillary tangles, pre-tangles and neuropil threads. Furthermore, histological studies and immunoblotting confirmed a mixed Alzheimer type 3-and 4-repeat tau pathology. Microtubule-associated protein 1A/1B-light chain 3 (LC3) immunoblotting of brain homogenates indicated autophagic activity and may support the role of WDR45 in autophagy. Conclusions: The widespread Alzheimer-type tau pathology in this disease indicates that this should be considered as a tauopathy and adds further support to the proposal that impaired autophagy may have a role in tauopathies. [ABSTRACT FROM AUTHOR]

Background Neurodegeneration with brain iron accumulation (NBIA) comprises a clinically and genetically heterogeneous group of diseases presenting with movement disorders and brain iron deposits. In addition to NBIA subtypes caused by mutations in PANK2 and PLA2G6 , mutations in the C19orf12 gene were recently described as the third frequent cause of NBIA (called mitochondrial membrane protein-associated neurodegeneration, MPAN). Additionally, the X-linked gene WDR45 was found causative for a special subtype named static encephalopathy in childhood with neurodegeneration in adulthood (also called BPAN); however, analysis of this gene in a broader spectrum of NBIA has not been reported yet. Methods In a heterogeneous cohort of 69 patients with suspected NBIA that did not carry mutations in PANK2 and PLA2G6 , the coding region of C19orf12 was evaluated by Sanger sequencing. The WDR45 gene was analyzed via high resolution melting and subsequent sequence analysis. Results Previously described homozygous C19orf12 mutations were found in 3/69 NBIA patients (4.3%). Analysis of the WDR45 gene revealed a novel heterozygous missense mutation in one female NBIA patient showing psychomotor retardation with secondary decline. Conclusions C19orf12 mutations were confirmed in our heterogeneous NBIA cohort, while WDR45 mutations appear to be restricted to the subtype showing encephalopathy in childhood with neurodegeneration in adulthood. [ABSTRACT FROM AUTHOR]

The article presents a case study of a young woman who showed signs of Rett syndrome. It notes that the patient suffered deterioration in her communication and motor skills. A historical background of the disease and the present condition of the patient is discussed. It is concluded that there is a hard time in predicting the disease course and provide accurate genetic counseling to the family.

Mutations in WDR45 and WDR45B cause the human neurological diseases β-propeller protein-associated neurodegeneration (BPAN) and intellectual disability (ID), respectively. WDR45 and WDR45B, along with WIPI1 and WIPI2, belong to a WD40 repeat-containing phosphatidylinositol-3-phosphate (PI(3)P)-binding protein family. Their yeast homolog Atg18 forms a complex with Atg2 and is required for autophagosome formation in part by tethering isolation membranes (IMs) (autophagosome precursor) to the endoplasmic reticulum (ER) to supply lipid for IM expansion in the autophagy pathway. The exact functions of WDR45/45B are unclear. We show here that WDR45/45B are specifically required for neural autophagy. In Wdr45/45b -depleted cells, the size of autophagosomes is decreased, and this is rescued by overexpression of ATG2A, providing in vivo evidence for the lipid transfer activity of ATG2-WIPI complexes. WDR45/45B are dispensable for the closure of autophagosomes but essential for the progression of autophagosomes into autolysosomes. WDR45/45B interact with the tether protein EPG5 and target it to late endosomes/lysosomes to promote autophagosome maturation. In the absence of Wdr45/45b , formation of the fusion machinery, consisting of SNARE proteins and EPG5, is dampened. BPAN- and ID-related mutations of WDR45/45B fail to rescue the autophagy defects in Wdr45/45b -deficient cells, possibly due to their impaired binding to EPG5. Promoting autophagosome maturation by inhibiting O -GlcNAcylation increases SNARE complex formation and facilitates the fusion of autophagosomes with late endosomes/lysosomes in Wdr45/45b double knockout (DKO) cells. Thus, our results uncover a novel function of WDR45/45B in autophagosome-lysosome fusion and provide molecular insights into the development of WDR45/WDR45B mutation-associated diseases. • β-propeller proteins WDR45/45B regulate autophagosome maturation in neural cells • WDR45/45B bind to the tether protein EPG5 and assist its late endosome localization • Disease-related mutations of WDR45/45B impair their function in autophagy • Inhibition of O -GlcNAcylation suppresses the defects in Wdr45/45b -deficient cells Ji et al. demonstrate that the β-propeller proteins WDR45 and WDR45B modulate autophagosome-lysosome fusion in neural cells. WDR45/45B interact with the tether protein EPG5 for its targeting to late endosomes/lysosomes and therefore promote the formation of SNARE-tether fusion complexes during autophagosome maturation into autolysosomes. [ABSTRACT FROM AUTHOR]

Background: Beta‐propeller protein‐associated neurodegeneration (BPAN, OMIM 300894) is an X‐linked neurodegenerative disorder caused by mutations in WDR45. WDR45 is required for autophagy, defect in WDR45 impaired autophagy which contributes for the pathogenesis of BPAN. Previously, we reported a novel de novo mutation (c.1040_1041del, p.Glu347GlyfsTer7) in WDR45 (NM_007075) in a 3‐year‐old Chinese girl with BPAN. Methods: The protein structure was constructed using SWISS‐MODEL and the isoelectric point (pI) was predicted by the online pI/Mw tool at ExPASy. The functional effects of this mutation were predicted by two online software programs: PROVEN and MutationTaster. Stable overexpression of Flag‐tagged wild‐type or mutant WDR45 in HeLa cells was constructed. Protein levels of LC3 and p62 were analyzed by western blot upon treatment with/without autophagy inhibitor Bafilomycin A1, the formation of LC3 puncta were analyzed in HeLa cells transfected with mCherry‐LC3 by confocal microscopy. Results: The mutation resulted in a shift of pI from 6.74 to 8.84 and was predicted to be pathogenic. The protein levels of LC3‐II and p62 were increased in cells overexpression of wild‐type and mutant WDR45 while the protein levels were not increased in cells overexpression of mutant WDR45 upon treatment with autophagy inhibitor Bafilomycin A1. Results from confocal microscopy revealed that LC3‐positive puncta were increased in cells expressing both wild‐type and mutant WDR45 while the number of LC3‐positive puncta was not increased in cells expressing mutant WDR45 upon treatment with Bafilomycin A1. Conclusion: Our study evidenced that this novel mutation in WDR45 impaired autophagy in cells thus this mutation is the cause for BPAN in this patient. [ABSTRACT FROM AUTHOR]