Fig 2: Myr-Chromosome 9 open reading frame 72 (C9orf72) peptide rescues neuronal loss and motor dysfunction. A The effect of different concentrations of Myr-C9orf72 peptide on the viability of MPP+-treated primary neurons. n = 3. ***p < 0.001, **p < 0.01. B Western blotting of C9orf72, p62, a-synuclein, and LC3. C Statistical analysis of the relative content of C9orf72. D Statistical analysis of the relative content of p62. E Western blotting of a-synuclein. F Statistical analysis of the relative content of LC3. n = 3/group, **p < 0.01, *p < 0.05. G Immunofluorescence staining of TH. H TH+ neuron count and statistical analysis. n = 3/group. ***p < 0.001, *p < 0.05. I Rotarod test data after the experiment. J Open field data after the experiment. n = 12/group, compared with the control group, ***p<0.001, **p<0.01. Data are presented as the mean ± SEM.

Fig 3: Knockdown of Chromosome 9 open reading frame 72 (C9orf72) aggravates autophagy and the overexpression of C9orf72 rescues autophagy. (A) Western blotting of p62, a-synuclein, and LC3 in C9orf72-overexpressing SY5Y cells induced by MPP+. (B) Statistical analysis of the relative content of p62 in C9orf72-overexpressing SY5Y cells induced by MPP+. (C) Statistical analysis of the relative content of a-synuclein in C9orf72-overexpressing SY5Y cells induced by MPP+. (D) Western blotting of LC3 in C9orf72-overexpressing SY5Y cells induced by MPP+. (E) Western blotting of C9orf72, p62, a-synuclein, and LC3 in C9orf72-knockdown mice. (F) Statistical analysis of the relative content of C9orf72 in C9orf72-knockdown mice. (G) Statistical analysis of the relative content of p62 in C9orf72-knockdown mice. (H) Western blotting of a-synuclein in C9orf72-knockdown mice. (I) Statistical analysis of the relative content of LC3 in C9orf72-knockdown mice. n = 3/group, ***p < 0.001, **p < 0.01, *p < 0.05. (J) Immunofluorescence staining of TH enzyme in C9orf72-knockdown mice. (K) TH+ neuron count and statistical analysis in C9orf72-knockdown mice. n = 3/group. ***p < 0.001, **p < 0.01, *p < 0.05. Data are presented as the mean ± SEM.

Fig 4: Cdk5 inhibitors inhibited the phosphorylation degradation of Chromosome 9 open reading frame 72 (C9orf72). (A) Western blotting of Cdk5 inhibitor ROSCOVITINE on the expression of C9orf72, p62, a-synuclein, and LC3 in MPP+-induced neurons. (B) Statistical analysis of the relative content of C9orf72 in MPP+-induced neurons inhibited by ROSCOVITINE. (C) Statistical analysis of the relative content of p62 in MPP+-induced neurons inhibited by ROSCOVITINE. (D) Statistical analysis of the relative content of a-synuclein in MPP+-induced neurons inhibited by ROSCOVITINE. (E) Statistical analysis of the relative content of LC3 in MPP+-induced neurons inhibited by ROSCOVITINE. (F) Western blotting of the Cdk5 inhibitor ROSCOVITINE on the expression of C9orf72, p62, a-synuclein, and LC3 in MPP+-induced SY5Y cells. (G) Statistical analysis of the relative content of C9orf72 in MPP+-induced SY5Y cells inhibited by ROSCOVITINE. (H) Statistical analysis of the relative content of p62 in MPP+-induced SY5Y cells inhibited by ROSCOVITINE. (I) Statistical analysis of the relative content of a-synuclein in MPP+-induced SY5Y cells inhibited by ROSCOVITINE. (J) Statistical analysis of the relative content of LC3 in MPP+-induced SY5Y cells inhibited by ROSCOVITINE. (K) Western blotting of the Cdk5 inhibitor ROSCOVITINE on the expression of C9orf72, p62, a-synuclein, and LC3 in PFF-induced neurons. (L) Statistical analysis of the relative content of C9orf72 in PFF-induced neurons inhibited by ROSCOVITINE. (M) Statistical analysis of the relative content of p62 in PFF-induced neurons inhibited by ROSCOVITINE. (N) Statistical analysis of the relative content of a-synuclein in PFF-induced neurons inhibited by ROSCOVITINE. (O) Statistical analysis of the relative content of LC3 in PFF-induced neurons inhibited by ROSCOVITINE. n = 3/group, **p < 0.01, *p < 0.05. Data are presented as the mean ± SEM.

Fig 5: Chromosome 9 open reading frame 72 (C9orf72) expression was decreased in Parkinson's disease (PD) cellular models. (A) Western blotting of C9orf72, p62, a-synuclein, and LC3 in MPP+-induced neurons. (B) Statistical analysis of the relative content of C9orf72 in MPP+-induced neurons. (C) Statistical analysis of the relative content of p62 in MPP+-induced neurons. (D) Statistical analysis of the relative content of a-synuclein in MPP+-induced neurons. (E) Statistical analysis of the relative content of LC3 in MPP+-induced neurons. (F) Western blotting of C9orf72, p62, a-synuclein, and LC3 in MPP+-induced SH-SY5Y cells. (G) Statistical analysis of the relative content of C9orf72 in MPP+-induced SH-SY5Y cells. (H) Statistical analysis of the relative content of p62 in MPP+-induced SH-SY5Y cells. (I) Statistical analysis of the relative content of a-synuclein in MPP+-induced SH-SY5Y cells. (J) Statistical analysis of the relative content of LC3 in MPP+-induced SH-SY5Y cells. (K) Western blotting of C9orf72, p62, a-synuclein, and LC3 in an A53T-induced SY5Y genetic model. (L) Statistical analysis of the relative content of C9orf72 in an A53T-induced SY5Y genetic model. (M) Statistical analysis of the relative content of p62 in an A53T-induced SY5Y genetic model. (N) Statistical analysis of the relative content of a-synuclein in an A53T-induced SY5Y genetic model. (O) Statistical analysis of the relative content of LC3 in an A53T-induced SY5Y genetic model. (P) Western blotting of C9orf72, p62, a-synuclein, and LC3 in a PFF-induced neurons preclinical model. (Q) Statistical analysis of the relative content of C9orf72 in a PFF-induced neurons preclinical model. (R) Statistical analysis of the relative content of p62 in a PFF-induced neurons preclinical model. (S) Statistical analysis of the relative content of a-synuclein in a PFF-induced neurons preclinical model. (T) Statistical analysis of the relative content of LC3 in a PFF-induced neurons preclinical model. n = 3/group, **p < 0.01, *p < 0.05. Data are presented as the mean ± SEM.