The Core MICOS Complex Subunit mic60 has Been Substituted by Two Cryptic Mitofilin-containing Proteins in Euglenozoa.

Area of Science:

• Cell Biology
• Mitochondrial Biology
• Evolutionary Biology

Background:

• Cristae are mitochondrial subcompartments essential for cellular respiration, formed with the help of the Mitochondrial contact site and Cristae Organizing System (MICOS) complex.
• The core MICOS protein, Mic60, is conserved from alphaproteobacterial ancestors and is vital for crista formation in most eukaryotes.
• Euglenozoa, including trypanosomes, lack Mic60, suggesting an alternative mechanism for MICOS complex assembly and crista maintenance.

Purpose of the Study:

• To investigate the replacement of Mic60 in Euglenozoa and identify the proteins responsible for MICOS complex function.
• To elucidate the evolutionary divergence and functional roles of novel MICOS subunits in Euglenozoa.
• To understand the impact of these divergent proteins on mitochondrial structure and biogenesis.

Main Methods:

• Comparative genomics to identify Mic60 homologs in Euglenozoa.
• Reverse genetics and protein interaction studies to characterize Mic34 and Mic40.
• In vitro phospholipid binding assays.
• Heterologous expression in bacteria and yeast to assess membrane remodeling.
• Analysis of mitochondrial morphology in Trypanosoma brucei under varying conditions.

Main Results:

• Mic60 is replaced by two distinct, divergent MICOS subunits, Mic34 and Mic40, in Euglenozoa.
• Mic34 and Mic40 are not integral membrane proteins but are involved in the oxidative protein folding pathway essential for mitochondrial and crista biogenesis.
• Mic34 binds phospholipid bilayers and, upon overexpression, elaborates the mitochondrion in Trypanosoma brucei, indicating a role in membrane remodeling.
• Mutations in Mic34's mitofilin domain abolish its function, highlighting conserved functional motifs.

Conclusions:

• The mitofilin protein family exhibits greater diversity than previously known, with Mic34 and Mic40 representing highly divergent members.
• Euglenozoan MICOS complex assembly relies on these novel subunits, which have adapted to maintain mitochondrial function and crista structure.
• This study reveals an alternative evolutionary trajectory for mitochondrial membrane organization and the MICOS complex.