The novel mitochondrial matrix protease Ste23 is required for efficient presequence degradation and processing.

Approximately 70% of mitochondrial precursor proteins are imported from the cytosol via N-terminal presequences, which are cleaved upon exposure to the mitochondrial processing protease MPP in the matrix. Cleaved presequence peptides then need to be efficiently degraded, and impairment of this clearance step, for example, by amyloid ß peptides, causes feedback inhibition of MPP, leading ultimately to accumulation of immature precursor proteins within mitochondria. Degradation of mitochondrial peptides is performed by Cym1 in yeast and its homologue, PreP, in humans. Here we identify the novel mitochondrial matrix protease Ste23 in yeast, a homologue of human insulin-degrading enzyme, which is required for efficient peptide degradation. Ste23 and Cym1 tightly cooperate to ensure the correct functioning of the essential presequence processing machinery.

C-terminally mutated tubby protein accumulates in aggresomes.

The tubby protein (Tub), a putative transcription factor, plays important roles in the maintenance and function of neuronal cells. A splicing defect-causing mutation in the 3'-end of the tubby gene, which is predicted to disrupt the carboxy-terminal region of the Tub protein, causes maturity-onset obesity, blindness, and deafness in mice. Although this pathological Tub mutation leads to a loss of function, the precise mechanism has not yet been investigated. Here, we found that the mutant Tub proteins were mostly localized to puncta found in the perinuclear region and that the C-terminus was important for its solubility. Immunocytochemical analysis revealed that puncta of mutant Tub co-localized with the aggresome. Moreover, whereas wild-type Tub was translocated to the nucleus by extracellular signaling, the mutant forms failed to undergo such translocation. Taken together, our results suggest that the malfunctions of the Tub mutant are caused by its misfolding and subsequent localization to aggresomes. [BMB Reports 2017; 50(1): 37-42].

Oxidative stress and neurodegenerative diseases: a neurotrophic approach.

Neurotrophins are important neurotrophic factors involved in the survival, differentiation and function of a wide variety of neuron populations. A common feature for most neurotrophins is that they are synthesized as precursor proteins (pro-neurotrophins) that upon being processed by proteolysis render the mature active form responsible for most of their trophic functions. However, some of the pro-neurotrophin form of these proteins, such as the precursor form of NGF (pro-NGF), have been shown to induce opposite effects and trigger apoptosis on neurons through the p75NTR receptor. This suggests that the balance between the levels of proneurotrophin and neurotrophin must be tightly controlled. In this context, it has been shown that in conditions of oxidative stress due for instance to aging or the development of some neurodegenerative disease, neurotrophins are oxidatively modified at least by advanced glycation/lipoxidation end products (AGE/ALEs) which makes pro-NGF refractary to be processed. The lack of maturation and the imbalance in favor of the precursor form may change the pattern of active signaling pathways towards cell death, thus exacerbating the deleterious alterations, for instance during the development of neurodegenerative diseases. Besides that, AGE/ALEs also induce the processing of the pro-NGF receptor p75NTR by α- secretase which is followed by the processing by γ -secretase and the release of the intracellular domain of p75NTR (p75NTRICD). Once cleaved, p75NTRICD recruits two intracellular interactors, NRIF and TRAF6, which allows NRIF phosphorylation by JNK. The phosphorylated form of NRIF then translocates to the nucleus and induces the expression of pro-apoptotic proteins. In this chapter we will summarize the mechanisms by which ROS- induce protein modifications, which proteins are susceptible to be modified, how these modifications affect function and signaling and, finally, how they can be related to neurodegenerative diseases.

Oxygen restriction as challenge test reveals early high-fat-diet-induced changes in glucose and lipid metabolism.

Challenge tests stress homeostasis and may reveal deviations in health that remain masked under unchallenged conditions. Ideally, challenge tests are non-invasive and applicable in an early phase of an animal experiment. Oxygen restriction (OxR; based on ambient, mild normobaric hypoxia) is a non-invasive challenge test that measures the flexibility to adapt metabolism. Metabolic inflexibility is one of the hallmarks of the metabolic syndrome. To test whether OxR can be used to reveal early diet-induced health effects, we exposed mice to a low-fat (LF) or high-fat (HF) diet for only 5 days. The response to OxR was assessed by calorimetric measurements, followed by analysis of gene expression in liver and epididymal white adipose tissue (eWAT) and serum markers for e.g. protein glycation and oxidation. Although HF feeding increased body weight, HF and LF mice did not differ in indirect calorimetric values under normoxic conditions and in a fasting state. Exposure to OxR; however, increased oxygen consumption and lipid oxidation in HF mice versus LF mice. Furthermore, OxR induced gluconeogenesis and an antioxidant response in the liver of HF mice, whereas it induced de novo lipogenesis and an antioxidant response in eWAT of LF mice, indicating that HF and LF mice differed in their adaptation to OxR. OxR also increased serum markers of protein glycation and oxidation in HF mice, whereas these changes were absent in LF mice. Cumulatively, OxR is a promising new method to test food products on potential beneficial effects for human health.

Salivary type I collagen degradation end-products and related matrix metalloproteinases in periodontitis.

AIM: Type I collagen degradation end-products and related matrix metalloproteinases (MMPs) were examined aiming to detect potential markers of periodontitis in saliva, with high sensitivity and specificity. MATERIALS AND METHODS: The salivary concentrations of MMP-8, MMP-9 and MMP-13, tartrate-resistant acid phosphatase serum type 5b, C-terminal cross-linked telopeptide of type I collagen (CTx), N-terminal cross-linked telopeptide of type I collagen (NTx) and cross-linked carboxyterminal telopeptide of type I collagen were analysed in 230 subjects. Oral health examination included panoramic radiography. RESULTS: The concentrations of MMP-8, MMP-9 and MMP-13 in saliva were higher in subjects with generalized periodontitis than in controls. Of the tested salivary markers, MMP-8 was the only marker capable of differentiating subjects with severe alveolar bone loss from those with slight bone loss (p < 0.001). The association between the salivary MMP-8 levels and periodontitis remained significant after the adjustment with age, gender and smoking. In addition, significant correlations were found between the tested markers and periodontal parameters. CONCLUSION: Enzymes and end-products of type I collagen degradation have different associations with each other and with periodontal status that may reflect their roles in the cascade leading to alveolar bone loss. MMP-8 is a strong biomarker candidate for detecting alveolar bone destruction.