Enforced lysosomal biogenesis rescues erythromycin- and clindamycin-induced mitochondria-mediated cell death in human cells

Molecular and Cellular Biochemistry - Antibiotics are the front-line treatment against many bacterial infectious diseases in human. The excessive and long-term use of antibiotics in human cause...     

Roles of the NH2-terminal Domains of Cardiac Ryanodine Receptor in Ca2+ Release Activation and Termination

The NH₂-terminal region (residues 1–543) of the cardiac ryanodine receptor (RyR2) harbors a large number of mutations associated with cardiac arrhythmias and cardiomyopathies. Functional studies have revealed that the NH₂-terminal region is involved in the activation and termination of Ca²⁺ release. The three-dimensional structure of the NH₂-terminal region has recently been solved. It is composed of three domains (A, B, and C). However, the roles of these individual domains in Ca²⁺ release activation and termination are largely unknown. To understand the functional significance of each of these NH₂-terminal domains, we systematically deleted these domains and assessed their impact on caffeine- or Ca²⁺-induced Ca²⁺ release and store overload-induced Ca²⁺ release (SOICR) in HEK293 cells. We found that all deletion mutants were capable of forming caffeine- and ryanodine-sensitive functional channels, indicating that the NH₂-terminal region is not essential for channel gating. Ca²⁺ release measurements revealed that deleting domain A markedly reduced the threshold for SOICR termination but had no effect on caffeine or Ca²⁺ activation or the threshold for SOICR activation, whereas deleting domain B substantially enhanced caffeine and Ca²⁺ activation and lowered the threshold for SOICR activation and termination. Conversely, deleting domain C suppressed caffeine activation, abolished Ca²⁺ activation and SOICR, and diminished protein expression. These results suggest that domain A is involved in channel termination, domain B is involved in channel suppression, and domain C is critical for channel activation and expression. Our data shed new insights into the structure-function relationship of the NH₂-terminal domains of RyR2 and the action of NH₂-terminal disease mutations.     

Easy Come,Easy Go: Capillary Forces Enable Rapid Refilling of Embolized Primary Xylem Vessels

Protoxylem plays an important role in the hydraulic function of vascular systems of both herbaceous and woody plants, but relatively little is known about the processes underlying the maintenance of protoxylem function in long-lived tissues. In this study, embolism repair was investigated in relation to xylem structure in two cushion plant species, Azorella macquariensis and Colobanthus muscoides, in which vascular water transport depends on protoxylem. Their protoxylem vessels consisted of a primary wall with helical thickenings that effectively formed a pit channel, with the primary wall being the pit channel membrane. Stem protoxylem was organized such that the pit channel membranes connected vessels with paratracheal parenchyma or other protoxylem vessels and were not exposed directly to air spaces. Embolism was experimentally induced in excised vascular tissue and detached shoots by exposing them briefly to air. When water was resupplied, embolized vessels refilled within tens of seconds (excised tissue) to a few minutes (detached shoots) with water sourced from either adjacent parenchyma or water-filled vessels. Refilling occurred in two phases: (1) water refilled xylem pit channels, simplifying bubble shape to a rod with two menisci; and (2) the bubble contracted as the resorption front advanced, dissolving air along the way. Physical properties of the protoxylem vessels (namely pit channel membrane porosity, hydrophilic walls, vessel dimensions, and helical thickenings) promoted rapid refilling of embolized conduits independent of root pressure. These results have implications for the maintenance of vascular function in both herbaceous and woody species, because protoxylem plays a major role in the hydraulic systems of leaves, elongating stems, and roots.There is a pressing need to understand how plants manage the maintenance of water transport from roots through leaves under changing environmental conditions (Allen et al., 2010; Choat et al., 2012). The problem arises because water is transported through the xylem under tension (i.e. under negative absolute pressure). As tension increases, conduits become increasingly vulnerable to cavitation, which causes the conduits to lose their ability to transport water. Conduits can become embolized during normal diurnal function as a result of tensions induced by transpiration and in response to environmental conditions such as drought or freezing stress (Zimmermann and Tyree, 2002). Vulnerability to cavitation and embolism formation suggests that plants have mechanisms to regain lost hydraulic capacity, either through the formation of new conduits or by refilling embolized ones.The vulnerability of conduits to embolisms and the capacity for repair are related to the structural diversity of xylem tissue (Zwieniecki and Holbrook, 2009; Lens et al., 2011; Cai et al., 2014). In vascular plants, the classification of xylem tissues depends on the meristem that produced them (Evert and Eichhorn, 2006). Primary xylem is produced by apical meristems and includes both protoxylem and metaxylem conduits, which are distinguished by their wall structure and the timing of their development. Protoxylem matures during organ elongation, which results in loss of function due to stretching in some tissues and species, while in many others, functionality is maintained throughout the life of the organ. In contrast, metaxylem matures in elongated tissue. In herbaceous plants, primary xylem is the major hydraulic system of the roots, stems, and leaves. In woody plants, the primary xylem remains the main hydraulic system of the leaves, while the radial growth of stems occurs through the activity of a vascular cambium, which produces secondary xylem with only metaxylem conduits. As a woody plant grows, the secondary xylem (and hence the metaxylem) thus becomes of increasing importance to stem hydraulic function. However, protoxylem remains an integral component of the plant hydraulic system due to its function in leaves and elongating stems and roots.As discussed in a recent review (Brodersen and McElrone, 2013), refilling of embolized vessels has been shown to depend on the generation of positive pressure by roots in many monocots, herbaceous plants, and a few woody species. However, many species lack root pressure; thus, attention has focused on so-called novel refilling, which involves adjacent living cells in the repair of embolized metaxylem or secondary xylem in stems of mature plants. Novel refilling has been studied with a variety of methods to visualize temporal variation in the presence and subsequent absence of embolized vessels, including cryo-scanning electron microscopy (Cryo-SEM; Canny, 1997; McCully et al., 2014), double staining (Zwieniecki and Holbrook, 1998; Zwieniecki et al., 2000), NMR imaging (Holbrook et al., 2001; Zwieniecki et al., 2013), and high-resolution x-ray computed tomography (Lee and Kim, 2008; Brodersen et al., 2010; Kim and Lee, 2010; Lee et al., 2013; Suuronen et al., 2013). These observations, in combination with other measurements, led to a working hypothesis of an osmotically driven repair mechanism in which sugars pumped into embolized vessels by adjacent paratracheal parenchyma provide the osmotic pressure difference that refills the vessel (Nardini et al., 2011).Little is known about embolism and its repair in protoxylem, which has structural features that make it potentially more vulnerable to embolism than metaxylem in the same plant or tissue (Choat et al., 2005). These include a greater exposed area of the primary cell wall with annular or helical thickenings instead of secondary walls. This could enhance stretching of the primary wall when large pressure differences develop between functional and embolized vessels, thereby decreasing the pressure required for air seeding of bubbles (Choat et al., 2004). Choat et al. (2005) suggested that greater vulnerability of protoxylem to embolism might underpin the roles of petioles, leaves, and small stems in the hydraulic segmentation hypothesis of Zimmermann (1983), in which sacrifice of the most easily replaceable tissues protects the function of the main structure of a plant during water stress. If ease of protoxylem embolism were to contribute to the function of hydraulic fuses during mild water stress, then ease of refilling would be required to rapidly reset the system.This study focuses on embolism repair in two distantly related, vascular species, Azorella macquariensis (Apiaceae) and Colobanthus muscoides (Caryophyllaceae), that depend exclusively on protoxylem for vascular water transport. Both species form cushions, with the former being an endemic, keystone species in the alpine zone of subantarctic Macquarie Island and the latter being a regional endemic that plays a major role in rocky coastal areas often within the supralittoral zone (Selkirk et al., 1990; Orchard, 1993). Both species are of ecological interest, because the subantarctic region is under increasing threat from climate change (Adams, 2009). Specifically, the climate on Macquarie Island is progressively changing from one that is perpetually wet and misty to one with increased exposure to periodic drying (Bergstrom et al., 2015). Dieback of alpine vegetation was first observed in 2008, and by 2010, extensive and unprecedented decline of A. macquariensis led to its listing as critically endangered (Bricher et al., 2013).In this study, protoxylem structure was studied in relation to embolism repair. Refilling of gas-filled vessels was compared between excised tissue and that in intact, detached shoots. The results showed that the physical properties of the protoxylem facilitated refilling by capillary forces and that rapid refilling in detached shoots supplied with water occurred without root pressure.     

Nucleic acid oxidation: an early feature of Alzheimer's disease

Studies of oxidative damage during the progression of Alzheimer's disease (AD) suggest its central role in disease pathogenesis. To investigate levels of nucleic acid oxidation in both early and late stages of AD, levels of multiple base adducts were quantified in nuclear and mitochondrial DNA from the superior and middle temporal gyri (SMTG), inferior parietal lobule (IPL), and cerebellum (CER) of age‐matched normal control subjects, subjects with mild cognitive impairment, preclinical AD, late‐stage AD, and non‐AD neurological disorders (diseased control; DC) using gas chromatography/mass spectrometry. Median levels of multiple DNA adducts in nuclear and mitochondrial DNA were significantly (p ≤ 0.05) elevated in the SMTG, IPL, and CER in multiple stages of AD and in DC subjects. Elevated levels of fapyguanine and fapyadenine in mitochondrial DNA suggest a hypoxic environment early in the progression of AD and in DC subjects. Overall, these data suggest that oxidative damage is an early event not only in the pathogenesis of AD but is also present in neurodegenerative diseases in general.

     

High‐throughput sequencing reveals neustonic and planktonic microbial eukaryote diversity in coastal waters

Neustonic organisms inhabit the sea surface microlayer (SML) and have important roles in marine ecosystem functioning. Here, we use high‐throughput 18S rRNA gene sequencing to characterize protist and fungal diversity in the SML at a coastal time‐series station and compare with underlying plankton assemblages. Protist diversity was higher in February (pre‐bloom) compared to April (spring bloom), and was lower in the neuston than in the plankton. Major protist groups, including Stramenopiles and Alveolata, dominated both neuston and plankton assemblages. Chrysophytes and diatoms were enriched in the neuston in April, with diatoms showing distinct changes in community composition between the sampling periods. Pezizomycetes dominated planktonic fungi assemblages, whereas fungal diversity in the neuston was more varied. This is the first study to utilize a molecular‐based approach to characterize neustonic protist and fungal assemblages, and provides the most comprehensive diversity assessment to date of this ecosystem. Variability in the SML microeukaryote assemblage structure has potential implications for biogeochemical and food web processes at the air‐sea interface.