Fig. 6.

HIGD2A is required for the stability of newly translated COX3. A, The HIGD2A^KO cell line was complemented with a retroviral HIGD2A^FLAG construct and mitochondria analyzed by BN and SDS-PAGE and immunoblotting with the indicated antibodies. B, Affinity-enrichment mass spectrometry (AE-MS) was performed on the HIGD2A^KOHIGD2A^FLAG cell line and significant interaction partners visualized through a volcano plot. The curved line indicates significance determined through an FDR-based approach (FDR<5%, S0 = 1). n = 3. C, Pulse SILAC analysis of newly translated mtDNA encoded OXPHOS subunits in HIGD2A^KO cells. SILAC media was added following overnight treatment with chloramphenicol (CAP) and analysis was performed at 0, 1, 2- and 4-hours post media addition. Log₂ transformed heavy peptide-derived intensities were plotted relative to controls. Data reported as mean ± S.D. n = 3. D, As for C but heavy media exchanged with light media after a 3h pulse and cells cultured for the indicated times. n = 2.