FIGURE

Figure 3

ID

ZDB-FIG-210512-51

Publication

Sofou et al., 2021 - Bi-allelic VPS16 variants limit HOPS/CORVET levels and cause a mucopolysaccharidosis-like disease

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Figure 3

The c.2272‐18C>A mutation impairs <italic>VPS16</italic> splicing and leads to reduced levels of VPS16 and HOPS/CORVET components

The c.2272‐18C>A variant generates a new splice‐acceptor sequence 16 base pairs upstream of the normal exon 23 border, resulting in a frameshift and premature stop codon.

Levels of VPS16 transcripts in fibroblasts by qPCR. The probe of the exon 22–24 PCR spans the mis‐spliced intron‐exon border. Data were normalized to levels of POLR2A using the ∆∆Ct method.

Quantifications of VPS16 in fibroblast lysates by immunoblotting. Representative immunoblots (bottom) and summary quantifications (top) of the levels of VPS16 in patient A (n = 4) and patient B (n = 3), normalized to levels of actin (ACTB) and expressed as % of controls.

Schematic illustration of HOPS and CORVET complexes showing their subunit compositions. The Rab5‐binding CORVET subunits VPS3 and VPS8 replace the Rab7‐binding VPS39 and VPS41 of HOPS. Models adapted from Bröcker et al (2012).

E–G

Quantifications of HOPS/CORVET subunits in fibroblast lysates (top) and representative immunoblots (bottom), analyzed as in (C). (E) VPS33A, (F) VPS11, (G) VPS18.

Analysis of VPS33A and VPS11 mRNA levels in fibroblasts by qPCR. Data were normalized to levels of POLR2A using the ∆∆Ct method.

Immunoblots (left) and quantifications normalized to control (right) of VPS16, VPS33A, and VPS11 in lysates from fibroblasts transduced with a lentivirus to express VPS16 or control (empty vector).

Data information: All immunoblots were acquired using fluorescent secondaries and pseudocolored to greyscale. Stars denote unspecific bands. All data were shown as mean ±SEM of 3–4 biological replicates. Statistical comparisons were made by two‐tailed unpaired Student’s t‐tests [panels B, C (right), E (right), F, G and I] or one‐way ANOVA with Dunnett's multiple comparisons tests [panels C (left) and E(left)]. *P < 0.05; **P < 0.01; ***P < 0.001.

Expression Data

Expression Detail

Antibody Labeling

Phenotype Data

Phenotype Detail

Acknowledgments

This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ EMBO Mol. Med.