Analysis of mutations in Neurospora crassa ERMES components reveals specific functions related to β-barrel protein assembly and maintenance of mitochondrial morphology.

Abstract

The endoplasmic reticulum mitochondria encounter structure (ERMES) tethers the ER to mitochondria and contains four structural components: Mmm1, Mdm12, Mdm10, and Mmm2 (Mdm34). The Gem1 protein may play a role in regulating ERMES function. Saccharomyces cerevisiae and Neurospora crassa strains lacking any of Mmm1, Mdm12, or Mdm10 are known to show a variety of phenotypic defects including altered mitochondrial morphology and defects in the assembly of β-barrel proteins into the mitochondrial outer membrane. Here we examine ERMES complex components in N. crassa and show that Mmm1 is an ER membrane protein containing a Cys residue near its N-terminus that is conserved in the class Sordariomycetes. The residue occurs in the ER-lumen domain of the protein and is involved in the formation of disulphide bonds that give rise to Mmm1 dimers. Dimer formation is required for efficient assembly of Tom40 into the TOM complex. However, no effects are seen on porin assembly or mitochondrial morphology. This demonstrates a specificity of function and suggests a direct role for Mmm1 in Tom40 assembly. Mutation of a highly conserved region in the cytosolic domain of Mmm1 results in moderate defects in Tom40 and porin assembly, as well as a slight morphological phenotype. Previous reports have not examined the role of Mmm2 with respect to mitochondrial protein import and assembly. Here we show that absence of Mmm2 affects assembly of β-barrel proteins and that lack of any ERMES structural component results in defects in Tom22 assembly. Loss of N. crassa Gem1 has no effect on the assembly of these proteins but does affect mitochondrial morphology.