A designed point mutant in Fis1 disrupts dimerization and mitochondrial fission.
Mitochondrial and peroxisomal fission are essential processes with defects resulting in cardiomyopathy and neonatal lethality. Central to organelle fission is Fis1, a monomeric tetratricopeptide repeat (TPR)-like protein whose role in assembly of the fission machinery remains obscure. Two nonfunctional, Saccharomyces cerevisiae Fis1 mutants (L80P or E78D/I85T/Y88H) were previously identified in genetic screens. Here, we find that these two variants in the cytosolic domain of Fis1 (Fis1ΔTM) are unexpectedly dimeric. A truncation variant of Fis1ΔTM that lacks an N-terminal regulatory domain is also found to be dimeric. The ability to dimerize is a property innate to the native Fis1ΔTM amino acid sequence as we find this domain is dimeric after transient exposure to elevated temperature or chemical denaturants and is kinetically trapped at room temperature. This is the first demonstration of a specific self-association in solution for the Fis1 cytoplasmic domain. We propose a three-dimensional domain-swapped model for dimerization that is validated by a designed mutation, A72P, which potently disrupts dimerization of wild-type Fis1. A72P also disrupts dimerization of nonfunctional variants, indicating a common structural basis for dimerization. The obligate monomer variant A72P, like the dimer-promoting variants, is nonfunctional in fission, consistent with a model in which Fis1 activity depends on its ability to interconvert between monomer and dimer species. These studies suggest a new functionally important manner in which TPR-containing proteins may reversibly self-associate.
Department of Biology, Johns Hopkins University, Baltimore, MD 21218, USA.
Jonathan P B
JP
Lees
Cara Marie
CM
Manlandro
Lora K
LK
Picton
Alexandra Z Ebie
AZ
Tan
Salvador
S
Casares
John M
JM
Flanagan
Karen G
KG
Fleming
R Blake
RB
Hill
D001665:Binding Sites; D019281:Dimerization; D008928:Mitochondria / Q000378:metabolism*; D063154:Mitochondrial Dynamics / Q000235:genetics*; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000235:genetics*; D017354:Point Mutation; D011487:Protein Conformation; D055503:Protein Multimerization; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics*
C464859:FIS1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins