The Assembly Factor Pet117 Couples Heme a Synthase Activity to Cytochrome Oxidase Assembly

Heme a is an essential metalloporphyrin cofactor of the mitochondrial respiratory enzyme cytochrome c oxidase (CcO). Its synthesis from heme b requires several enzymes, including the evolutionarily conserved heme a synthase (Cox15). Oligomerization of Cox15 appears to be important for the process of heme a biosynthesis and transfer to maturing CcO. However, the details of this process remain elusive, and the roles of any additional CcO assembly factors that may be involved remain unclear. Here we report the systematic analysis of one such uncharacterized assembly factor, Pet117, and demonstrate in Saccharomyces cerevisiae that this evolutionarily conserved protein is necessary for Cox15 oligomerization and function. Pet117 is shown to reside in the mitochondrial matrix, where it is associated with the inner membrane. Pet117 functions at the later maturation stages of the core CcO subunit Cox1 that precede Cox1 hemylation. Pet117 also physically interacts with Cox15 and specifically mediates the stability of Cox15 oligomeric complexes. This Cox15-Pet117 interaction observed by co-immunoprecipitation persists in the absence of heme a synthase activity, is dependent upon Cox1 synthesis and early maturation steps, and is further dependent upon the presence of the matrix-exposed, unstructured linker region of Cox15 needed for Cox15 oligomerization, suggesting that this region mediates the interaction or that the interaction is lost when Cox15 is unable to oligomerize. Based on these findings, it was concluded that Pet117 mediates coupling of heme a synthesis to the CcO assembly process in eukaryotes

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

The Role of Conserved Uncharacterized Proteins in Mitochondrial Health and Protein Homeostasis

Establishing and maintaining proper mitochondrial functions are critical for cellular wellbeing. To maintain this homeostasis, several conserved mechanisms are tasked to survey and protect the mitochondria against damage. This system is known as mitochondrial quality control (MQC). Malfunctioning MQC can lead to an accumulation of damaged or misfolded proteins, leading to a multitude of diseases and general degeneration while aging. Although paramount to overall mitochondrial health, few mechanistic details are known about the molecular functions and the proteins involved with the MQC. With a goal in mind to elucidate functions of highly conserved mitochondrial proteins of unknown functions, our lab identified Afg1, a conserved mitochondrial AAA+ ATPase (LACE-1 or AFG1L in higher eukaryotes), in a genetic knockout screen as a potential member of the MQC. Specifically, expression of Afg1 in Saccharomyces cerevisiae model lacking the mitochondrial proteases Oma1 and the m-AAA protease mitigated survival defect of this mutant. By applying mechanistic biochemistry and genetic approaches, we further discovered a novel role of Afg1 in mediating mitochondrial matrix proteostasis, in particular through aging and at times of cellular oxidative stress. Finally, we determined an evolutionarily conserved physiological role of the Afg1 ortholog, LACE-1, in the metazoan model Caenorhabditis elegans. Animals deficient in LACE-1 display a reduced lifespan and impaired oxidative stress tolerance. In particular, we found that loss of LACE-1 lead to impaired mitochondrial proteostasis in motor neuron circuitry and abnormal behavioral plasticity. Taken together, these results demonstrate that Afg1/LACE-1 is a bona fide member of the MQC and establish its conserved role in mediating mitochondrial proteostasis. The molecular function Afg1/LACE-1 performs has a profound effect on both physiological and behavioral aspects in higher eukaryotes

The Role of Conserved Uncharacterized Proteins in Mitochondrial Health and Protein Homeostasis

Establishing and maintaining proper mitochondrial functions are critical for cellular wellbeing. To maintain this homeostasis, several conserved mechanisms are tasked to survey and protect the mitochondria against damage. This system is known as mitochondrial quality control (MQC). Malfunctioning MQC can lead to an accumulation of damaged or misfolded proteins, leading to a multitude of diseases and general degeneration while aging. Although paramount to overall mitochondrial health, few mechanistic details are known about the molecular functions and the proteins involved with the MQC. With a goal in mind to elucidate functions of highly conserved mitochondrial proteins of unknown functions, our lab identified Afg1, a conserved mitochondrial AAA+ ATPase (LACE-1 or AFG1L in higher eukaryotes), in a genetic knockout screen as a potential member of the MQC. Specifically, expression of Afg1 in Saccharomyces cerevisiae model lacking the mitochondrial proteases Oma1 and the m-AAA protease mitigated survival defect of this mutant. By applying mechanistic biochemistry and genetic approaches, we further discovered a novel role of Afg1 in mediating mitochondrial matrix proteostasis, in particular through aging and at times of cellular oxidative stress. Finally, we determined an evolutionarily conserved physiological role of the Afg1 ortholog, LACE-1, in the metazoan model Caenorhabditis elegans. Animals deficient in LACE-1 display a reduced lifespan and impaired oxidative stress tolerance. In particular, we found that loss of LACE-1 lead to impaired mitochondrial proteostasis in motor neuron circuitry and abnormal behavioral plasticity. Taken together, these results demonstrate that Afg1/LACE-1 is a bona fide member of the MQC and establish its conserved role in mediating mitochondrial proteostasis. The molecular function Afg1/LACE-1 performs has a profound effect on both physiological and behavioral aspects in higher eukaryotes

The Assembly Factor Pet117 Couples Heme a Synthase Activity to Cytochrome Oxidase Assembly

Heme a is an essential metalloporphyrin cofactor of the mitochondrial respiratory enzyme cytochrome c oxidase (CcO). Its synthesis from heme b requires several enzymes, including the evolutionarily conserved heme a synthase (Cox15). Oligomerization of Cox15 appears to be important for the process of heme a biosynthesis and transfer to maturing CcO. However, the details of this process remain elusive, and the roles of any additional CcO assembly factors that may be involved remain unclear. Here we report the systematic analysis of one such uncharacterized assembly factor, Pet117, and demonstrate in Saccharomyces cerevisiae that this evolutionarily conserved protein is necessary for Cox15 oligomerization and function. Pet117 is shown to reside in the mitochondrial matrix, where it is associated with the inner membrane. Pet117 functions at the later maturation stages of the core CcO subunit Cox1 that precede Cox1 hemylation. Pet117 also physically interacts with Cox15 and specifically mediates the stability of Cox15 oligomeric complexes. This Cox15-Pet117 interaction observed by co-immunoprecipitation persists in the absence of heme a synthase activity, is dependent upon Cox1 synthesis and early maturation steps, and is further dependent upon the presence of the matrix-exposed, unstructured linker region of Cox15 needed for Cox15 oligomerization, suggesting that this region mediates the interaction or that the interaction is lost when Cox15 is unable to oligomerize. Based on these findings, it was concluded that Pet117 mediates coupling of heme a synthesis to the CcO assembly process in eukaryotes