Pathogenic variants in BORCS5 cause a spectrum of neurodevelopmental and neurodegenerative disorders with lysosomal dysfunction
Niccolò E. Mencacci, Georgia Minakaki, Reza Maroofian, Raffaella De Pace, Adeline Paimboeuf, Tiago Branco Fonseca, Tatiana Abramova, Patrick Shannon, David Chitayat, Francesca Magrinelli, Wesley J. Peng, Diptaman Chatterjee, Sara H. Eldessouky, Julia Baptista, Tamas Marton, Julie Vogt, Juan Dario Ortigoza-Escobar, Loreto Martorell, Marta Gómez-Chiari, Ingrid M. Wentzensen, Erik-Jan Kamsteeg, Maha S. Zaki, Annarita Scardamaglia, Giovanni Zifarelli, Zuhair Nasser Al-Hassnan, Elka Miller, Shiri Shinar, Lova S. Matsa, Sri Hari Chandan Appikonda, Ghada A. Otaify, Khalid Al-Thihli, Almundher Al-Maawali, Michael Schwake, Mariasavina Severino, Henry Houlden, Shunmoogum A. Patten, Juan S. Bonifacino, Kailash P. Bhatia, Dimitri Krainc
Niccolò E. Mencacci, Georgia Minakaki, Reza Maroofian, Raffaella De Pace, Adeline Paimboeuf, Tiago Branco Fonseca, Tatiana Abramova, Patrick Shannon, David Chitayat, Francesca Magrinelli, Wesley J. Peng, Diptaman Chatterjee, Sara H. Eldessouky, Julia Baptista, Tamas Marton, Julie Vogt, Juan Dario Ortigoza-Escobar, Loreto Martorell, Marta Gómez-Chiari, Ingrid M. Wentzensen, Erik-Jan Kamsteeg, Maha S. Zaki, Annarita Scardamaglia, Giovanni Zifarelli, Zuhair Nasser Al-Hassnan, Elka Miller, Shiri Shinar, Lova S. Matsa, Sri Hari Chandan Appikonda, Ghada A. Otaify, Khalid Al-Thihli, Almundher Al-Maawali, Michael Schwake, Mariasavina Severino, Henry Houlden, Shunmoogum A. Patten, Juan S. Bonifacino, Kailash P. Bhatia, Dimitri Krainc
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Research Article Cell biology Genetics Neuroscience

Pathogenic variants in BORCS5 cause a spectrum of neurodevelopmental and neurodegenerative disorders with lysosomal dysfunction

Abstract

BORCS5 encodes a subunit of the BLOC-One-Related Complex (BORC), which is known to promote anterograde movement and fusion of lysosomes. We identified 16 individuals from 9 families with bi-allelic BORCS5 variants, revealing a spectrum of neurodevelopmental and neurodegenerative phenotypes. Carriers of homozygous protein-truncating variants (PTVs), resulting in complete loss of BORCS5, presented with prenatally lethal arthrogryposis multiplex congenita, brain malformations, and neuropathological evidence of neuroaxonal dystrophy. Individuals with missense or splice-site variants presented differently, with microcephaly, developmental epileptic encephalopathy, optic atrophy, spasticity, and progressive movement disorders. In this group, brain MRI showed diffuse hypomyelination, corpus callosum abnormalities, and progressive global cerebral atrophy, consistent with neurodegeneration. Borcs5 KO in zebrafish resulted in microcephaly, motor deficits, and increased seizure susceptibility, mirroring the patients’ clinical presentation. At the cellular level, only BORCS5 PTVs, but not missense variants, led to perinuclear lysosomal clustering and impaired lysosomal axonal trafficking in induced pluripotent stem cell–derived forebrain neurons. However, PTVs and missense variants were associated with reduced lysosomal proteolysis and activity of lysosomal hydrolases glucocerebrosidase and cathepsin B, indicating lysosomal dysfunction. Our study reveals a role for BORCS5 in modulation of lysosomal function, in addition to its known role in lysosome movement and fusion, possibly underlying the diverse clinical manifestations in individuals with BORCS5-related disorders.

Authors

Niccolò E. Mencacci, Georgia Minakaki, Reza Maroofian, Raffaella De Pace, Adeline Paimboeuf, Tiago Branco Fonseca, Tatiana Abramova, Patrick Shannon, David Chitayat, Francesca Magrinelli, Wesley J. Peng, Diptaman Chatterjee, Sara H. Eldessouky, Julia Baptista, Tamas Marton, Julie Vogt, Juan Dario Ortigoza-Escobar, Loreto Martorell, Marta Gómez-Chiari, Ingrid M. Wentzensen, Erik-Jan Kamsteeg, Maha S. Zaki, Annarita Scardamaglia, Giovanni Zifarelli, Zuhair Nasser Al-Hassnan, Elka Miller, Shiri Shinar, Lova S. Matsa, Sri Hari Chandan Appikonda, Ghada A. Otaify, Khalid Al-Thihli, Almundher Al-Maawali, Michael Schwake, Mariasavina Severino, Henry Houlden, Shunmoogum A. Patten, Juan S. Bonifacino, Kailash P. Bhatia, Dimitri Krainc

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Figure 5

Impact of BORCS5 pathogenic variants on BORCS5 protein expression, BORC assembly, and endolysosome distribution in cell lines.

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Impact of BORCS5 pathogenic variants on BORCS5 protein expression, BORC ...
(A) Western blot analysis of BORCS5 protein in HEK293T cells 24 hours after transfection with constructs expressing GFP (control) and BORCS5 WT or the indicated BORCS5 patient variants. GAPDH was used as a loading control. For WT and R95Q/H99P, the band indicated by * was quantified, whereas the band indicated by an arrow was quantified for A106fs and Y139*. In the case of L128fs, the entire band pattern was measured. The 25 kDa band present in the case of A106fs and Y139* variants correspond to endogenous BORCS5, as shown in GFP-only condition. The graph shows the mean ± SEM of fold change over the mean of the WT variant (n = 4 independent experiments). For BORCS5 levels, 1-way ANOVA F(5,41) = 59.49 (P < 0.0001) with Dunnett’s post hoc test compared with WT. ***P = 0.0001, ****P < 0.0001. (B) GFP trap precipitation of GFP-tagged BORCS5 variants and their interaction (co-IP) with endogenous SNAPIN and BORCS7, with GAPDH used as a specificity control. The graph shows mean ± SEM n of 3 independent experiments. For BORCS7, 1-way ANOVA with Dunnett’s post hoc test F(3,8) = 33.17 (P < 0.0001). **P ≤ 0.003. For SNAPIN, F(3,8) = 26.53 (P = 0.0002). *P < 0.018. (C and D) Immunofluorescence microscopy shows endogenous LAMP1 puncta (white) distribution in untransfected WT and BORCS5 KO HeLa (as control) or BORCS5-KO HeLa cells transiently cotransfected with BORCS5 constructs and GFP (* indicates GFP+ cells). Nuclei were labeled with DAPI (blue), and cell edges were outlined by fluorescent phalloidin (dashed lines). Scale bars: 20 μm. (E and F) Schematic depicts LAMP1+ endolysosomes present in a 2 μm peripheral shell, shown in the graph as mean ± SEM for 3 independent experiments, each dot representing a different cell. For peripheral lysosomes, 1-way ANOVA with Dunnett multiple comparisons test (compared with the WT construct), F(7,289) = 30.96 (P < 0.0001). *P < 0.05, **P = 0.0098, ****P < 0.0001.