Figure 2.
Figure 2. Membrane association enhances Rab29 activation of pathogenic LRRK2 and Rab phosphorylation. (A) HEK293T cells were transfected with R1441G LRRK2 and Myc-Rab29 or Myc-Rab9 and treated with either DMSO or 10 µM lovastatin to inhibit prenylation. Samples were immunoblotted 24 h later for pS1292 LRRK2, total LRRK2 (UDD3), and Myc. An ∼50-kD nonspecific band is shown at bottom as a loading control. (B) Quantitation of A. Error bars represent SEM from two experiments; P < 0.0001 from two-tailed unpaired t test. (C) HA-tagged Rab8A, Rab10, and Rab29 were coexpressed with FLAG-R1441G LRRK2 (±200 nM MLi-2, 1.5 h) or FLAG-R1441G/D2017A LRRK2 in HEK293T cells. Mutant Rab8A(C204S), Rab10(C199/200S), and Rab29(C202/203S) were used along with the corresponding WT Rabs. Samples were immunoblotted for the respective phosphorylated Rabs, HA, total LRRK2, and tubulin.

Membrane association enhances Rab29 activation of pathogenic LRRK2 and Rab phosphorylation. (A) HEK293T cells were transfected with R1441G LRRK2 and Myc-Rab29 or Myc-Rab9 and treated with either DMSO or 10 µM lovastatin to inhibit prenylation. Samples were immunoblotted 24 h later for pS1292 LRRK2, total LRRK2 (UDD3), and Myc. An ∼50-kD nonspecific band is shown at bottom as a loading control. (B) Quantitation of A. Error bars represent SEM from two experiments; P < 0.0001 from two-tailed unpaired t test. (C) HA-tagged Rab8A, Rab10, and Rab29 were coexpressed with FLAG-R1441G LRRK2 (±200 nM MLi-2, 1.5 h) or FLAG-R1441G/D2017A LRRK2 in HEK293T cells. Mutant Rab8A(C204S), Rab10(C199/200S), and Rab29(C202/203S) were used along with the corresponding WT Rabs. Samples were immunoblotted for the respective phosphorylated Rabs, HA, total LRRK2, and tubulin.

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