Mitochondria-Associated Degradation Pathway (MAD) Function beyond the Outer Membrane.

The mitochondria-associated degradation pathway (MAD) mediates ubiquitination and degradation of mitochondrial outer membrane (MOM) proteins by the proteasome. We find that the MAD, but not other quality-control pathways including macroautophagy, mitophagy, or mitochondrial chaperones and proteases, is critical for yeast cellular fitness under conditions of paraquat (PQ)-induced oxidative stress in mitochondria. Specifically, inhibition of the MAD increases PQ-induced defects in growth and mitochondrial quality and decreases chronological lifespan. We use mass spectrometry analysis to identify possible MAD substrates as mitochondrial proteins that exhibit increased ubiquitination in response to PQ treatment and inhibition of the MAD. We identify candidate substrates in the mitochondrial matrix and inner membrane and confirm that two matrix proteins are MAD substrates. Our studies reveal a broader function for the MAD in mitochondrial protein surveillance beyond the MOM and a major role for the MAD in cellular and mitochondrial fitness in response to chronic, low-level oxidative stress in mitochondria.

ACVR1R206H receptor mutation causes fibrodysplasia ossificans progressiva by imparting responsiveness to activin A.

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder characterized by episodically exuberant heterotopic ossification (HO), whereby skeletal muscle is abnormally converted into misplaced, but histologically normal bone. This HO leads to progressive immobility with catastrophic consequences, including death by asphyxiation. FOP results from mutations in the intracellular domain of the type I BMP (bone morphogenetic protein) receptor ACVR1; the most common mutation alters arginine 206 to histidine (ACVR1(R206H)) and has been thought to drive inappropriate bone formation as a result of receptor hyperactivity. We unexpectedly found that this mutation rendered ACVR1 responsive to the activin family of ligands, which generally antagonize BMP signaling through ACVR1 but cannot normally induce bone formation. To test the implications of this finding in vivo, we engineered mice to carry the Acvr1(R206H) mutation. Because mice that constitutively express Acvr1[R206H] die perinatally, we generated a genetically humanized conditional-on knock-in model for this mutation. When Acvr1[R206H] expression was induced, mice developed HO resembling that of FOP; HO could also be triggered by activin A administration in this mouse model of FOP but not in wild-type controls. Finally, HO was blocked by broad-acting BMP blockers, as well as by a fully human antibody specific to activin A. Our results suggest that ACVR1(R206H) causes FOP by gaining responsiveness to the normally antagonistic ligand activin A, demonstrating that this ligand is necessary and sufficient for driving HO in a genetically accurate model of FOP; hence, our human antibody to activin A represents a potential therapeutic approach for FOP.

Aim44p regulates phosphorylation of Hof1p to promote contractile ring closure during cytokinesis in budding yeast.

Whereas actomyosin and septin ring organization and function in cytokinesis are thoroughly described, little is known regarding the mechanisms by which the actomyosin ring interacts with septins and associated proteins to coordinate cell division. Here we show that the protein product of YPL158C, Aim44p, undergoes septin-dependent recruitment to the site of cell division. Aim44p colocalizes with Myo1p, the type II myosin of the contractile ring, throughout most of the cell cycle. The Aim44p ring does not contract when the actomyosin ring closes. Instead, it forms a double ring that associates with septin rings on mother and daughter cells after cell separation. Deletion of AIM44 results in defects in contractile ring closure. Aim44p coimmunoprecipitates with Hof1p, a conserved F-BAR protein that binds both septins and type II myosins and promotes contractile ring closure. Deletion of AIM44 results in a delay in Hof1p phosphorylation and altered Hof1p localization. Finally, overexpression of Dbf2p, a kinase that phosphorylates Hof1p and is required for relocalization of Hof1p from septin rings to the contractile ring and for Hof1p-triggered contractile ring closure, rescues the cytokinesis defect observed in aim44 cells. Our studies reveal a novel role for Aim44p in regulating contractile ring closure through effects on Hof1p.