Developmental Activation of Mitochodnrial OPA1 Processing Is Retinoic Acid-Independent

Mitochondria exist as an organellar network within most eukaryotic organisms, and are critical to biochemical energy production and cellular stress response. Mitochondrial bioenergetic function is directly linked to the complex morphological state of its network, existing in a balance of fusion (interconnected state) or fission (fragmented state). The optic atrophy-1 (OPA1) protein plays a major role in regulating mitochondrial inner membrane fusion, whereas the OMA1 metallopeptidase is highly involved in the degradation of the OPA1 protein, thus contributing to its fragmented state. Cellular stress such as a disruption in mitochondrial membrane potential activates OMA1 cleavage of OPA1. However, our data indicates membrane potential disruption with CCCP after cardiac-like differentiation with Retinoic Acid (RA) in H9c2 cardiomyoblast cell lines activate OMA1, suggesting a developmental switch for OPA1 cleavage. To test whether the activation of OMA1 is RA specific, skeletal muscle-like differentiation (with low serum media lacking RA) upon challenge with CCCP will be used to observe if OPA1 cleavage by OMA1 occurs. We hypothesize that differentiation in H9c2s by FBS will activate OMA1

Mitochondrial OMA1 and OPA1 as Gatekeepers of Organellar Structure/Function and Cellular Stress Response

Mammalian mitochondria are emerging as a critical stress-responsive contributor to cellular life/death and developmental outcomes. Maintained as an organellar network distributed throughout the cell, mitochondria respond to cellular stimuli and stresses through highly sensitive structural dynamics, particularly in energetically demanding cell settings such as cardiac and muscle tissues. Fusion allows individual mitochondria to form an interconnected reticular network, while fission divides the network into a collection of vesicular organelles. Crucially, optic atrophy-1 (OPA1) directly links mitochondrial structure and bioenergetic function: when the transmembrane potential across the inner membrane (ΔΨm) is intact, long L-OPA1 isoforms carry out fusion of the mitochondrial inner membrane. When ΔΨm is lost, L-OPA1 is cleaved to short, fusion-inactive S-OPA1 isoforms by the stress-sensitive OMA1 metalloprotease, causing the mitochondrial network to collapse to a fragmented population of organelles. This proteolytic mechanism provides sensitive regulation of organellar structure/function but also engages directly with apoptotic factors as a major mechanism of mitochondrial participation in cellular stress response. Furthermore, emerging evidence suggests that this proteolytic mechanism may have critical importance for cell developmental programs, particularly in cardiac, neuronal, and stem cell settings. OMA1\u27s role as a key mitochondrial stress-sensitive protease motivates exciting new questions regarding its mechanistic regulation and interactions, as well as its broader importance through involvement in apoptotic, stress response, and developmental pathways

Mitochondrial OPA1 cleavage is reversibly activated by differentiation of H9c2 cardiomyoblasts

Optic atrophy-1 (OPA1) is a dynamin-like GTPase localized to the mitochondrial inner membrane, playing key roles in inner membrane fusion and cristae maintenance. OPA1 is regulated by the mitochondrial transmembrane potential (Δψm): when Δψm is intact, long OPA1 isoforms (L-OPA1) carry out inner membrane fusion. Upon loss of Δψm, L-OPA1 isoforms are proteolytically cleaved to short (S-OPA1) isoforms by the stress-inducible OMA1 metalloprotease, causing collapse of the mitochondrial network and promoting apoptosis. Here, we show that L-OPA1 isoforms of H9c2 cardiomyoblasts are retained under loss of Δψm, despite the presence of OMA1. However, when H9c2s are differentiated to a more cardiac-like phenotype via treatment with retinoic acid (RA) in low serum media, loss of Δ ψm induces robust, and reversible, cleavage of L-OPA1 and subsequent OMA1 degradation. These findings indicate that a potent developmental switch regulates Δ ψm-sensitive OPA1 cleavage, suggesting novel developmental and regulatory mechanisms for OPA1 homeostasis