Unique structural features of the mitochondrial AAA+ protease AFG3L2 reveal the molecular basis for activity in health and disease

Abstract

Mitochondrial AAA+ quality control proteases regulate diverse aspects of mitochondrial biology through specialized protein degradation, but the underlying molecular mechanisms that define the diverse activities of these enzymes remain poorly defined. The mitochondrial AAA+ protease AFG3L2 is of particular interest, as genetic mutations localized throughout AFG3L2 are linked to diverse neurodegenerative disorders. However, a lack of structural data has limited our understanding of how mutations impact enzymatic activity. Here, we used cryo-EM to determine a substrate-bound structure of the catalytic core of human AFG3L2. This structure identifies multiple specialized structural features within AFG3L2 that integrate with conserved structural motifs required for hand-over-hand ATP-dependent substrate translocation to engage, unfold and degrade targeted proteins. Mapping disease-relevant AFG3L2 mutations onto our structure demonstrates that many of these mutations localize to these unique structural features of AFG3L2 and distinctly influence its activity and stability. Our results provide a molecular basis for neurological phenotypes associated with different AFG3L2 mutations, and establish a structural framework to understand how different members of the AAA+ superfamily achieve specialized, diverse biological functions.

Expanding the clinical and genetic heterogeneity of SPAX5

“…Cerebellar atrophy seems common, but can be mild or barely detectable 1. Neuroimaging in two patients with p.Val723Met6 showed cerebellar atrophy, whereas the basal ganglia were spared. The new case of SPAX5 described in the present report corroborates the impression that a wide spectrum of features, including iron deposition in the basal ganglia, can be attributed to m‐AAA protease defects.…”

“…The mechanisms by which different mutations in AFG3L2 lead to distinct neurological phenotypes constitute an important, open question in the field. Recently, Puchades and colleagues6 described distinct biochemical impacts of different mutations in the m‐AAA AGF3L2 protease, suggesting that these differences might constitute the molecular basis for the different neurodegenerative conditions associated with mutations in AFG3L2 . We here describe the case of a child with SPAX5 in whom severe MRI features suggesting a mitochondrial disorder are difficult to reconcile with his milder, fluctuating clinical course.…”

“…More than 30 different AFG3L2 disease‐causing mutations have thus far been reported in over 80 SCA28 patients,11,12 whereas few SPAX5 families have been described 1,6,13. Interestingly, previously described patients with bi‐allelic mutations in AFG3L2 (Table 1) have severe phenotypes, including early‐onset spastic ataxia‐neuropathy syndrome or severe developmental delay and microcephaly, and often present progressive myoclonic epilepsy 1,13.…”

“…The relationships found between different AFG3L2 mutations and distinct forms of the disease, different levels of severity and neuropathological correlates in mice indicate that these variants impact differentially on m‐AAA protease structure and activity. However, a lack of structural information has limited our understanding of the molecular mechanisms linking specific mutations to altered enzymatic activity and, ultimately, disease features 6…”

Dissecting the Roles of Mitochondrial Complex I Intermediate Assembly (MCIA) Complex Factors in the Biogenesis of Complex I

“…Assembly factors play a prominent role in the biogenesis of complex I and perform functions that include the post-translational modification of subunits (Rhein et al, 2013;Rhein et al, 2016;Zurita Rendon et al, 2014), delivery of cofactors (Sheftel et al, 2009), insertion of proteins into the inner membrane, and stabilization of partially assembled, intermediate modules . In cases where complex I subunits are damaged or assembly is compromised, modified, unincorporated, labile or misassembled subunits are degraded by mitochondrial proteases to maintain mitochondrial integrity or reduce levels of reactive oxygen species (Pryde et al, 2016;Puchades et al, 2019;Rendón and Shoubridge, 2012;Stiburek et al, 2012).…”