Transmembrane BAX Inhibitor-1 Motif Containing Protein 5 (TMBIM5) Sustains Mitochondrial Structure, Shape, and Function by Impacting the Mitochondrial Protein Synthesis Machinery

Abstract

The Transmembrane Bax Inhibitor-1 motif (TMBIM)-containing protein family is evolutionarily conserved and has been implicated in cell death susceptibility. The only member with a mitochondrial localization is TMBIM5 (also known as GHITM or MICS1), which affects cristae organization and associates with the Parkinson’s disease-associated protein CHCHD2 in the inner mitochondrial membrane. We here used CRISPR-Cas9-mediated knockout HAP1 cells to shed further light on the function of TMBIM5 in physiology and cell death susceptibility. We found that compared to wild type, TMBIM5-knockout cells were smaller and had a slower proliferation rate. In these cells, mitochondria were more fragmented with a vacuolar cristae structure. In addition, the mitochondrial membrane potential was reduced and respiration was attenuated, leading to a reduced mitochondrial ATP generation. TMBIM5 did not associate with Mic10 and Mic60, which are proteins of the mitochondrial contact site and cristae organizing system (MICOS), nor did TMBIM5 knockout affect their expression levels. TMBIM5-knockout cells were more sensitive to apoptosis elicited by staurosporine and BH3 mimetic inhibitors of Bcl-2 and Bcl-XL. An unbiased proteomic comparison identified a dramatic downregulation of proteins involved in the mitochondrial protein synthesis machinery in TMBIM5-knockout cells. We conclude that TMBIM5 is important to maintain the mitochondrial structure and function possibly through the control of mitochondrial biogenesis.

The Evolutionary Conserved Transmembrane BAX Inhibitor Motif (TMBIM) Containing Protein Family Members 5 and 6 Are Essential for the Development and Survival of Drosophila melanogaster

“…Reduces ATP and Results in a More Pro-apoptotic State dTmbim5 contains a mitochondrial targeting signal and human TMBIM5 localizes to the inner mitochondrial membrane where it controls cristae structure (Oka et al, 2008) and mitochondrial function including mitochondrial respiration and ATP production (Seitaj et al, 2020). To clarify how loss of dTmbim5 causes its detrimental effect, we thus focused on mitochondria.…”

“…We next quantified the expression levels of genes involved in apoptosis because KO of human TMBIM5 was shown to make cells more susceptible to apoptosis of the intrinsic pathway involving cytochrome c release (Oka et al, 2008;Seitaj et al, 2020). Similar to vertebrates, apoptosis in flies is also regulated by the correct balance between pro-apoptotic and anti-apoptotic proteins (Figure 5C; Steller, 2008).…”

“…KD of dTmbim5 in the nervous system induced p53A expression (Figure 5E). The Bcl-2 family of anti-apoptotic proteins in particular regulates apoptosis by affecting mitochondrial permeability to Cytochrome c [reviewed by Singh et al (2019)] which is facilitated by TMBIM5 KO (Oka et al, 2008;Seitaj et al, 2020). In flies, there are two Bcl-2 members: the anti-apoptotic ER protein Buffy (Quinn et al, 2003;Doumanis et al, 2007) and the pro-apoptotic mitochondrial protein Debcl (Colussi et al, 2000;Doumanis et al, 2007).…”

“…TMBIM4 over-expression prevents apoptosis induced by different kinds of stimuli -the proapoptotic Bax, staurosporine, doxorubicin, C2ceramide, as well as TNFα and Fas ligand while its KD results in spontaneous apoptosis (Gubser et al, 2007). TMBIM5, a family member targeted to the mitochondrial inner membrane protein, maintains mitochondrial morphology and decreases the release of cytochrome c from mitochondria which indirectly links it to the induction of apoptosis (Oka et al, 2008) possibly by interfering with the mitochondrial protein synthesis machinery (Seitaj et al, 2020). Overexpression of TMBIM6 inhibits cell death caused by BAX, etoposide, staurosporine and growth factor withdrawal, possibly by forming a complex with the anti-apoptotic proteins BCL-2 and BCL-X L (Xu and Reed, 1998).…”

The Evolutionary Conserved Transmembrane BAX Inhibitor Motif (TMBIM) Containing Protein Family is Essential for the Development and Survival of Drosophila Melanogaster

“…TMBIM4 over-expression prevents apoptosis induced by different kinds of stimuli -the proapoptotic Bax, staurosporine, doxorubicin, C2-ceramide, as well as TNFa and Fas ligand while its knockdown results in spontaneous apoptosis 5 . TMBIM5, a family member targeted to the mitochondrial inner membrane protein, maintains mitochondrial morphology and decreases the release of cytochrome c from mitochondria which indirectly links it to the induction of apoptosis 6 possibly by interfering with the mitochondrial protein synthesis machinery 20 .…”