Figure 7. TMEM175 knockout (KO) mitigates 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐induced Parkinson's disease (PD) symptoms in mice.

• A
  Representative immunoblots for TMEM175 expression levels in the midbrains of mice of different ages (W: weeks; M: months).
• B
  Statistics of the relative TMEM175 protein levels normalized to GAPDH (glyceraldehyde‐3‐phosphate dehydrogenase) in (A).
• C
  Schematic diagram showing the KO strategy of TMEM175 in mice. Introns 2–10 were removed from the TMEM175 locus.
• D
  TMEM175 messenger RNA (mRNA) levels in the midbrains of wild‐type (WT) or TMEM175 KO mice were detected by reverse transcription–quantitative polymerase chain reaction (RT‐qPCR) using SYBR Green PCR Master Mix.
• E
  The knockout of TMEM175 in mouse was verified by Western blot of brain tissues.
• F
  Representative TMEM175 basal lysosomal currents recorded in macrophages of WT or TMEM175 KO mice. Ψ is the lysosomal membrane potential (defined as V_cytosol – V_lumen). The currents were elicited with ramp voltage protocols (−100 to +100 mV in 1 s, Vh = 0 mV).
• G
  Statistics of the current amplitudes of (F).
• H, I
  Quantification of rears in the cylinder (H) and the latency to fall from a wire hang apparatus (I) pre‐ and post‐MPTP treatment.
• J
  Swing duration coefficient of variation (CV) (CV%) (left panel) and Ataxia coefficient (right panel) in Gait tests 7 days post‐MPTP.
• K
  Latency to fall in rotarod tests 7 days post‐MPTP.
• L
  Immunoblots of the tyrosine hydroxylase (TH) levels post‐MPTP treatment in WT and TMEM175 KO mouse midbrains.
• M
  Quantification of the relative TH levels normalized to GAPDH post‐MPTP.
• N
  TH immunofluorescent labeling on substantia nigra slices of WT or TMEM175 KO mice post‐MPTP treatment. Scale bars = 100 μm.
• O
  Statistics of the TH‐positive neurons in (N).
• P
  An overview model illustrating the presumed roles of TMEM175 in apoptosis and PD. Antiapoptotic mitochondrial protein Bcl‐2 binds directly to and regulates lysosomal potassium channel TMEM175. Bcl‐2 inhibitors and mutations of the binding sites release the inhibition and thus activate TMEM175. Activation of TMEM175 inhibits mitophagy, impairs mitochondrial function, and consequently increases reactive oxygen species (ROS) production. ROS activates TMEM175 through a positive feedback way to augment apoptosis. TMEM175‐mediated apoptosis promotes the death of DA neurons in PD.

Data information: The data are presented as the mean ± SEM. Statistical significance was analyzed with two‐sided Student's t‐tests, and is indicated with NS for not significant (P > 0.05), * for P < 0.05, ** for P < 0.01, and *** for P < 0.001. n value means the number of biological replicates made for each data point.

Source data are available online for this figure.