Fig. 4: UBLCP1 ASD mutation results in decreased expression, enhanced proteasomal degradation of ubiquitinated proteins, and altered differential RNA expression of proteasome subunits. | Translational Psychiatry

Fig. 4: UBLCP1 ASD mutation results in decreased expression, enhanced proteasomal degradation of ubiquitinated proteins, and altered differential RNA expression of proteasome subunits.

From: A novel autism-associated UBLCP1 mutation impacts proteasome regulation/activity

Fig. 4

a UBLCP1 mRNA levels in fibroblasts from ASD patient compared to unaffected controls. b UBLCP1 nuclear protein expression in fibroblasts from ASD patient (n = 89) compared to unaffected controls (n = 90). c UBLCP1 cytoplasmic protein expression in fibroblasts from ASD patient (n = 107) compared to unaffected controls (n = 105). d Immunostaining of unaffected control fibroblasts (upper images) and fibroblasts from the ASD patient (lower images). UBLCP1 expression is in green, and the nucleus is counterstained with Hoechst in blue. ****P < 0.0001; Student’s t test; mean ± SEM. Scale bar: 10 µm. e Enhanced proteasome activity in fibroblasts from the proband compared to unaffected controls. Fibroblasts used in these experiments were at the same passage number. Proteasome activity was measured using the fluorogenic substrate Suc-LLVY-AMC. Student’s t test, *P = 0.0498, n = 3 in each group. f Decreased polyubiquitination detected by western blot in fibroblasts from the proband compared to unaffected controls. Ubiquitin and GAPDH were detected by western blot (left panel). Equivalent amounts of total protein were loaded into each lane within the same gel. Quantification is shown on the right. Relative levels of polyubiquitinated proteins (area of interest included 50–250 kDa) to GAPDH were quantified with n = 3 per group. *P = 0.0145; Student’s t test; mean ± SEM. g Increased UBLCP1 mRNA levels in fibroblasts from three independent controls treated with 1uM MG132 for 10 h compared to the same controls treated with DMSO vehicle only. h A general tendency for reduced mRNA expression of proteasome subunits (PSMA1, PSMB5, PSMC2, PSMC4, and PSMD2) from the proband’s cells compared to expression in cells derived from unaffected controls was observed. n = 3 per group. *P < 0.05; Student’s t test; mean ± SEM. i Enhanced expression of UBLCP1 and proteasome subunits (PSMA1, PSMB5, PSMC2, PSMC4, and PSMD2) mRNA extracted from proband cells treated with 1uM MG132 for 10 h compared to expression in proband cells treated only with DMSO vehicle. n = 3 per group. *P < 0.05; **P < 0.01; Student’s t test; mean ± SEM.

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