PTCD1 Is Required for Mitochondrial Oxidative-Phosphorylation: Possible Genetic Association with Alzheimer's Disease.

In addition to amyloid-ß plaques and tau tangles, mitochondrial dysfunction is implicated in the pathology of Alzheimer's disease (AD). Neurons heavily rely on mitochondrial function, and deficits in brain energy metabolism are detected early in AD; however, direct human genetic evidence for mitochondrial involvement in AD pathogenesis is limited. We analyzed whole-exome sequencing data of 4549 AD cases and 3332 age-matched controls and discovered that rare protein altering variants in the gene pentatricopeptide repeat-containing protein 1 (PTCD1) show a trend for enrichment in cases compared with controls. We show here that PTCD1 is required for normal mitochondrial rRNA levels, proper assembly of the mitochondrial ribosome and hence for mitochondrial translation and assembly of the electron transport chain. Loss of PTCD1 function impairs oxidative phosphorylation and forces cells to rely on glycolysis for energy production. Cells expressing the AD-linked variant of PTCD1 fail to sustain energy production under increased metabolic stress. In neurons, reduced PTCD1 expression leads to lower ATP levels and impacts spontaneous synaptic activity. Thus, our study uncovers a possible link between a protein required for mitochondrial function and energy metabolism and AD risk.SIGNIFICANCE STATEMENT Mitochondria are the main source of cellular energy and mitochondrial dysfunction is implicated in the pathology of Alzheimer's disease (AD) and other neurodegenerative disorders. Here, we identify a variant in the gene PTCD1 that is enriched in AD patients and demonstrate that PTCD1 is required for ATP generation through oxidative phosphorylation. PTCD1 regulates the level of 16S rRNA, the backbone of the mitoribosome, and is essential for mitochondrial translation and assembly of the electron transport chain. Cells expressing the AD-associated variant fail to maintain adequate ATP production during metabolic stress, and reduced PTCD1 activity disrupts neuronal energy homeostasis and dampens spontaneous transmission. Our work provides a mechanistic link between a protein required for mitochondrial function and genetic AD risk.

Proteolytic Processing of Neuregulin 1 Type III by Three Intramembrane-cleaving Proteases.

Numerous membrane-bound proteins undergo regulated intramembrane proteolysis. Regulated intramembrane proteolysis is initiated by shedding, and the remaining stubs are further processed by intramembrane-cleaving proteases (I-CLiPs). Neuregulin 1 type III (NRG1 type III) is a major physiological substrate of ß-secretase (ß-site amyloid precursor protein-cleaving enzyme 1 (BACE1)). BACE1-mediated cleavage is required to allow signaling of NRG1 type III. Because of the hairpin nature of NRG1 type III, two membrane-bound stubs with a type 1 and a type 2 orientation are generated by proteolytic processing. We demonstrate that these stubs are substrates for three I-CLiPs. The type 1-oriented stub is further cleaved by γ-secretase at an ϵ-like site five amino acids N-terminal to the C-terminal membrane anchor and at a γ-like site in the middle of the transmembrane domain. The ϵ-cleavage site is only one amino acid N-terminal to a Val/Leu substitution associated with schizophrenia. The mutation reduces generation of the NRG1 type III ß-peptide as well as reverses signaling. Moreover, it affects the cleavage precision of γ-secretase at the γ-site similar to certain Alzheimer disease-associated mutations within the amyloid precursor protein. The type 2-oriented membrane-retained stub of NRG1 type III is further processed by signal peptide peptidase-like proteases SPPL2a and SPPL2b. Expression of catalytically inactive aspartate mutations as well as treatment with 2,2'-(2-oxo-1,3-propanediyl)bis[(phenylmethoxy)carbonyl]-l-leucyl-l-leucinamide ketone inhibits formation of N-terminal intracellular domains and the corresponding secreted C-peptide. Thus, NRG1 type III is the first protein substrate that is not only cleaved by multiple sheddases but is also processed by three different I-CLiPs.

Dual cleavage of neuregulin 1 type III by BACE1 and ADAM17 liberates its EGF-like domain and allows paracrine signaling.

Proteolytic shedding of cell surface proteins generates paracrine signals involved in numerous signaling pathways. Neuregulin 1 (NRG1) type III is involved in myelination of the peripheral nervous system, for which it requires proteolytic activation by proteases of the ADAM family and BACE1. These proteases are major therapeutic targets for the prevention of Alzheimer's disease because they are also involved in the proteolytic generation of the neurotoxic amyloid ß-peptide. Identification and functional investigation of their physiological substrates is therefore of greatest importance in preventing unwanted side effects. Here we investigated proteolytic processing of NRG1 type III and demonstrate that the ectodomain can be cleaved by three different sheddases, namely ADAM10, ADAM17, and BACE1. Surprisingly, we not only found cleavage by ADAM10, ADAM17, and BACE1 C-terminal to the epidermal growth factor (EGF)-like domain, which is believed to play a pivotal role in signaling, but also additional cleavage sites for ADAM17 and BACE1 N-terminal to that domain. Proteolytic processing at N- and C-terminal sites of the EGF-like domain results in the secretion of this domain from NRG1 type III. The soluble EGF-like domain is functionally active and stimulates ErbB3 signaling in tissue culture assays. Moreover, the soluble EGF-like domain is capable of rescuing hypomyelination in a zebrafish mutant lacking BACE1. Our data suggest that NRG1 type III-dependent myelination is not only controlled by membrane-retained NRG1 type III, but also in a paracrine manner via proteolytic liberation of the EGF-like domain.

Estimation of Health and Economic Benefits of a Small Automatic External Defibrillator for Rapid Treatment of Sudden Cardiac Arrest (SMART): A Cost-Effectiveness Analysis.

Background: Sudden cardiac arrest (SCA) occurs in 0.4% of the general population and up to 6% or more of at-risk groups each year. Early CPR and defibrillation improves SCA outcomes but access to automatic external defibrillators (AEDs) remains limited. Methods: Markov models were used to evaluate the cost-effectiveness of a portable SMART (SMall AED for Rapid Treatment of SCA) approach to early SCA management over a life-time horizon in at-risk and not at-risk populations. Simulated patients (n = 600,000) who had not received an implantable cardioverter defibrillator (ICD) were randomized to a SMART device with CPR prompts or non-SMART approaches. Annual SCA risk was varied from 0.2 to 3.5%. Analysis was performed in a US economy from both societal (SP) and healthcare (HP) perspectives to evaluate the number of SCA fatalities prevented by SMART, and SMART cost-effectiveness at a threshold of $100,000/Quality Adjusted Life Year (QALY). Results: A SMART approach was cost-effective when annual SCA risk exceeded 1.51% (SP) and 1.62% (HP). The incremental cost-effectiveness ratios (ICER) were $95,251/QALY (SP) and $100,797/QALY (HP) at a 1.60% SCA annual risk. At a 3.5% annual SCA risk, SMART was highly cost-effective from both SP and HP [ICER: $53,925/QALY (SP), $59,672/QALY (HP)]. In microsimulation, SMART prevented 1,762 fatalities across risk strata (1.59% fatality relative risk reduction across groups). From a population perspective, SMART could prevent at least 109,839 SCA deaths in persons 45 years and older in the United States. Conclusions and Relevance: A SMART approach to SCA prophylaxis prevents fatalities and is cost-effective in patients at elevated SCA risk. The availability of a smart-phone enabled pocket-sized AED with CPR prompts has the potential to greatly improve population health and economic outcomes.

Discovery of Potent and Selective Dual Leucine Zipper Kinase/Leucine Zipper-Bearing Kinase Inhibitors with Neuroprotective Properties in In Vitro and In Vivo Models of Amyotrophic Lateral Sclerosis.

Dual leucine zipper kinase (DLK) and leucine zipper-bearing kinase (LZK) are regulators of neuronal degeneration and axon growth. Therefore, there is a considerable interest in developing DLK/LZK inhibitors for neurodegenerative diseases. Herein, we use ligand- and structure-based drug design approaches for identifying novel amino-pyrazine inhibitors of DLK/LZK. DN-1289 (14), a potent and selective dual DLK/LZK inhibitor, demonstrated excellent in vivo plasma half-life across species and is anticipated to freely penetrate the central nervous system with no brain impairment based on in vivo rodent pharmacokinetic studies and human in vitro transporter data. Proximal target engagement and disease relevant pathway biomarkers were also favorably regulated in an in vivo model of amyotrophic lateral sclerosis.

Antibody-mediated stabilization of NRG1 induces behavioral and electrophysiological alterations in adult mice.

Neuregulin 1 (NRG1) is required for development of the central and peripheral nervous system and regulates neurotransmission in the adult. NRG1 and the gene encoding its receptor, ERBB4, are risk genes for schizophrenia, although how alterations in these genes disrupt their function has not been fully established. Studies of knockout and transgenic mice have yielded conflicting results, with both gain and loss of function resulting in similar behavioral and electrophysiological phenotypes. Here, we used high affinity antibodies to NRG1 and ErbB4 to perturb the function of the endogenous proteins in adult mice. Treatment with NRG1 antibodies that block receptor binding caused behavioral alterations associated with schizophrenia, including, hyper-locomotion and impaired pre-pulse inhibition of startle (PPI). Electrophysiological analysis of brain slices from anti-NRG1 treated mice revealed reduced synaptic transmission and enhanced paired-pulse facilitation. In contrast, mice treated with more potent ErbB4 function blocking antibodies did not display behavioral alterations, suggesting a receptor independent mechanism of the anti-NRG1-induced phenotypes. We demonstrate that anti-NRG1 causes accumulation of the full-length transmembrane protein and increases phospho-cofilin levels, which has previously been linked to impaired synaptic transmission, indicating enhancement of non-canonical NRG1 signaling could mediate the CNS effects.

BACE1 across species: a comparison of the in vivo consequences of BACE1 deletion in mice and rats.

Assessing BACE1 (ß-site APP cleaving enzyme 1) knockout mice for general health and neurological function may be useful in predicting risks associated with prolonged pharmacological BACE1 inhibition, a treatment approach currently being developed for Alzheimer's disease. To determine whether BACE1 deletion-associated effects in mice generalize to another species, we developed a novel Bace1-/- rat line using zinc-finger nuclease technology and compared Bace1-/- mice and rats with their Bace1+/+ counterparts. Lack of BACE1 was confirmed in Bace1-/- animals from both species. Removal of BACE1 affected startle magnitude, balance beam performance, pain response, and nerve myelination in both species. While both mice and rats lacking BACE1 have shown increased mortality, the increase was smaller and restricted to early developmental stages for rats. Bace1-/- mice and rats further differed in body weight, spontaneous locomotor activity, and prepulse inhibition of startle. While the effects of species and genetic background on these phenotypes remain difficult to distinguish, our findings suggest that BACE1's role in myelination and some sensorimotor functions is consistent between mice and rats and may be conserved in other species. Other phenotypes differ between these models, suggesting that some effects of BACE1 inhibition vary with the biological context (e.g. species or background strain).

BACE1 dependent neuregulin processing: review.

Neuregulin-1 (NRG1), known also as heregulin, acetylcholine receptor inducing activity (ARIA), glial growth factor (GGF), or sensory and motor neuron derived factor (SMDF), plays essential roles in several developmental processes, and is required also later in life. Many variants of NRG1 are produced via alternative splicing and usage of distinct promoters. All contain an epidermal growth factor (EGF)-like domain, which alone is sufficient to bind and activate the cognate receptors, members of the ErbB family. NRG1 mediated signaling is crucial for cardiogenesis and the development of the mammary gland and ErbB2 (HER2), an orphan co-receptor for NRG1 is the target of the drug Herceptin� (trastuzumab) used for treatment of metastatic breast cancer. In the nervous system, NRG1 controls the early development of subpopulations of neural crest cells. In particular, NRG1 acts as an essential paracrine signaling molecule expressed on the axonal surface, where it signals to Schwann cells throughout development and regulates the thickness of the myelin sheath. NRG1 is required also by other cell types in the nervous system, for instance as an axonal signal released by proprioceptive afferents to induce development of the muscle spindle, and it controls aspects of cortical interneuron development as well as the formation of thalamocortical projections. Work from several laboratories implicates dysregulation of NRG1/ErbB4 signaling in the etiology of schizophrenia. Biochemical studies have shown that the precursor proteins of NRG1 can be released from the membrane through limited proteolysis. In addition, most NRG1 isoforms contain a transmembrane domain, which is processed by γ-secretase after shedding. Thereby the intracellular domain is released into the cytoplasm. Despite this, the importance of NRG1 cleavage for its functions in vivo remained unclear until recently. ß- Secretase (ß-site amyloid precursor protein cleaving enzyme 1, BACE1) was first identified through its function as the rate limiting enzyme of amyloid-ß-peptide (Aß) production. Aß is the major component of amyloid plaques in Alzheimer's disease (AD). More recently it was shown that Neuregulin-1 is a major physiological substrate of BACE1 during early postnatal development. Mutant mice lacking BACE1 display severe hypomyelination of peripheral nerves similar to that seen in mice lacking NRG1/ErbB signaling in Schwann cells, and a BACE1-dependent activation of NRG1 in the process of peripheral myelination was proposed. Here we summarize the current knowledge about the role of NRG1 proteolysis for ErbB receptor mediated signaling during development and in Alzheimer's disease.