The m-AAA Protease Associated with Neurodegeneration Limits MCU Activity in Mitochondria.

Mutations in subunits of mitochondrial m-AAA proteases in the inner membrane cause neurodegeneration in spinocerebellar ataxia (SCA28) and hereditary spastic paraplegia (HSP7). m-AAA proteases preserve mitochondrial proteostasis, mitochondrial morphology, and efficient OXPHOS activity, but the cause for neuronal loss in disease is unknown. We have determined the neuronal interactome of m-AAA proteases in mice and identified a complex with C2ORF47 (termed MAIP1), which counteracts cell death by regulating the assembly of the mitochondrial Ca2+ uniporter MCU. While MAIP1 assists biogenesis of the MCU subunit EMRE, the m-AAA protease degrades non-assembled EMRE and ensures efficient assembly of gatekeeper subunits with MCU. Loss of the m-AAA protease results in accumulation of constitutively active MCU-EMRE channels lacking gatekeeper subunits in neuronal mitochondria and facilitates mitochondrial Ca2+ overload, mitochondrial permeability transition pore opening, and neuronal death. Together, our results explain neuronal loss in m-AAA protease deficiency by deregulated mitochondrial Ca2+ homeostasis.

Trehalose accumulation enhances drought tolerance by modulating photosynthesis and ROS-antioxidant balance in drought sensitive and tolerant rice cultivars.

Trehalose being an integral part for plant growth, development and abiotic stress tolerance is accumulated in minute amounts in angiosperms with few exceptions from resurrection plants. In the current study, two rice cultivars differing in drought tolerance were used to analyse the role of trehalose in modulating photosynthesis and ROS-antioxidant balance leading to improvement in drought tolerance. Accumulation of trehalose in leaves of Vaisakh (drought-tolerant) and Aiswarya (drought-sensitive) rice cultivars was observed by spraying 50 mM trehalose and 100 µM validamycin A (trehalase inhibitor) followed by vacuum infiltration. Compared to stress sensitive Aiswarya cultivar, higher trehalose levels were observed in leaves of Vaisakh not only under control conditions but also under drought conditions corresponding with increased root length. The increase in leaf trehalose by treatment with trehalose or validamycin A corresponded well with a decrease in electrolyte leakage in sensitive and tolerant plants. Decreased ROS levels were reflected as increase in antioxidant enzyme activity and their gene expression in leaves of both the cultivars treated with trehalose or Validamycin A under control and drought conditions signifying the importance of trehalose in modulating the ROS-antioxidant balance for cellular protection. Further, higher chlorophyll, higher photosynthetic activity and modulation in other gas exchange parameters upon treatment with trehalose or validamycin A strongly suggested the beneficial role of trehalose for stress tolerance. Trehalose accumulation helped the tolerant cultivar adjust towards drought by maintaining higher water status and alleviating the ROS toxicity by effective activation and increment in antioxidant enzyme activity along with enhanced photosynthesis. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01404-7.

Exacerbation of drought-induced physiological and biochemical changes in leaves of Pisum sativum upon restriction of COX and AOX pathways of mitochondrial oxidative electron transport.

Drought stress affects photosynthesis, leading to significant decrease in crop productivity. In the current study, the importance of the cytochrome oxidase (COX) and alternative oxidase (AOX) pathways of themitochondrial oxidative electron transport chain (mETC) for photosynthesis and reactive oxygen species (ROS) homeostasis was evaluated in the leaves of Pisum sativum plants exposed to drought stress for 3 days (D3), 6 days (D6), and 9 days (D9). While drought stress resulted in decreased CO2 assimilation rates, leaf stomatal conductance, transpiration, and leaf intercellular CO2 concentration in a stress-dependent manner, superimposition with mETC inhibitors, antimycin A (AA) and salicylhydroxamic acid (SHAM), aggravated the responses. Decreased chlorophyll content, photosynthesis, and RubisCO (RbcL) degradation during progressive drought and their aggravation upon AOX pathway restriction indicated the importance of the AOX pathway for photosynthetic activity. Compared with COX pathway inhibition, higher intracellular H2O2 and O2.- levels, and increased cell death upon restriction of the AOX pathway during D6 and D9 stress conditions correlating with the modulation in antioxidant enzyme activities, signify the essentiality of the AOX pathway for ROS maintenance at optimal levels. Further, increased AOX1a expression during D6 and D9 conditions along with increasedAOXprotein levels indicated the activation of theAOXpathway during drought stress. Decline in Fv/Fm, actual quantum yield of PSII (ФPSII), photochemical quenching (qP), non-photochemical quenching (NPQ), and electron transport rate (ETR) upon restriction of the COX and AOX pathways indicated the requirement of mETC activity for optimal photochemical activities not only under normal conditions but also under progressive drought conditions.

Halotolerant PGPB Staphylococcus sciuri ET101 protects photosynthesis through activation of redox dissipation pathways in Lycopersicon esculentum.

Photosynthesis is known to be seriously affected by salt stress. The stress induced membrane damage leads to disrupted photosynthetic components causing imbalance between production and utilization of ATP/NADPH with generation of ROS leading to photoinhibition and photodamage. In the current study, role of halotolerant plant growth promoting bacteria (PGPB) Staphylococcus sciuri ET101 in protection of photosynthesis in tomato plants during salinity stress was evaluated by analysing changes in antioxidant defense and activation of redox dissipation pathways. Inoculation of S. sciuri ET101 significantly enhanced the growth of tomato plants with significantly higher photosynthetic rates (PN) under normal and salinity stress conditions. Further, increased membrane stability, soluble sugar accumulation and significant decrease in malondialdehyde (MDA) content in leaves of ET101 inoculated tomato plants under normal and salinity were observed along with increased expression of antioxidant genes for efficient ROS detoxification and suppression of oxidative damage. Additionally, salinity induced decrease in rate of photosynthesis (PN) due to lowered chloroplastic CO2 concentration (Cc) attributed by low mesophyll conductance (gm) in uninoculated plants was alleviated by ET101 inoculation showing significantly higher carboxylation rate (Vcmax), RuBP generation (Jmax) and increased photorespiration (PR). The genes involved in photorespiratory process, cyclic electron flow (CEF), and alternative oxidase (AOX) pathway of mitochondrial respiration were abundantly expressed in leaves of ET101 inoculated plants indicating their involvement in protecting photosynthesis from salt stress induced photoinhibition. Collectively, our results indicated that S. sciuri ET101 has the potential in protecting photosynthesis of tomato plants under salinity stress through activation of redox dissipation pathways.

Differential modulation of photosynthesis, ROS and antioxidant enzyme activities in stress-sensitive and -tolerant rice cultivars during salinity and drought upon restriction of COX and AOX pathways of mitochondrial oxidative electron transport.

Drought and salt stresses are two major abiotic stress factors that hamper crop growth and productivity. Three rice cultivars with different sensitivity and tolerance towards abiotic stress were used in the current study. While cultivar Aiswarya is salt- and drought-sensitive, cultivar Vyttila is salt-tolerant and cultivar Vaisakh is drought-tolerant. We compared the physiological and biochemical responses of these rice cultivars under salt and drought stress conditions after restricting their cytochrome oxidase (COX) and alternative oxidase (AOX) pathways using antimycin A and salicylhydroxamic acid treatment. Further, changes in their expression of AOX genes and corresponding protein levels were compared and analysed. The sensitive and tolerant rice cultivars subjected to drought and salt stress showed differential responses in physiological and biochemical traits. Whereas Aiswarya showed clear phenotypic differences, such as stunted growth, leaf curling, and loss of greening in leaf tissues, with increase in salt content and progressive drought stress, Vyttila and Vaisakh showed no remarkable changes. Moreover, the drought-tolerant cultivar rehydrated after 10 days of drought exposure, whereas the sensitive variety did not show any rehydration of leaf tissue. The leaves of the tolerant cultivars showed lower reactive oxygen species (ROS) production than that of the sensitive plants under drought and salt stress conditions because of the activation of a stronger antioxidant defence. Although, the restriction of COX and AOX pathways increased the susceptibility of sensitive cultivars, it affected the tolerant varieties moderately. Higher photosynthetic rates, an efficient antioxidant system comprising higher superoxide dismutase, ascorbate peroxidase, and catalase activity along with higher AOX1a gene expression levels during drought and salt stress were observed in tolerant cultivars. The results suggest that an efficient antioxidant system and increased transcription of the AOX1a gene along with higher AOX protein levels are important for tolerant rice cultivars to maintain higher photosynthesis rates, lower ROS, and stress tolerance. Restriction of COX and AOX pathways impact the photosynthesis, ROS, and antioxidant enzymes in both sensitive and tolerant cultivars. The restriction of COX and AOX pathways have a stronger impact on gas exchange and fluorescence parameters of the sensitive cultivar than on that of the tolerant cultivars owing to the higher photosynthetic rates in tolerant cultivars.

Marker reconstitution mutagenesis: a simple and efficient reverse genetic approach.

A novel reverse genetic approach termed 'marker reconstitution mutagenesis' was designed to generate mutational allelic series in genes of interest. This approach consists of two simple steps which utilize two selective markers. First, using one selective marker, a partial fragment of another selective marker gene is inserted adjacently to a gene of interest by homologous recombination. Second, random mutations are introduced precisely into the gene of interest, together with the reconstitution of the latter selective marker by homologous recombination. This approach was successfully tested for several genes in the fission yeast Schizosaccharomyces pombe. It circumvents the problems encountered with other methods and should be adaptable to any organism that incorporates exogenous DNA by homologous recombination.

Cytochrome oxidase and alternative oxidase pathways of mitochondrial electron transport chain are important for the photosynthetic performance of pea plants under salinity stress conditions.

The flexible plant mitochondrial electron transport chain with cytochrome c oxidase (COX) and alternative oxidase (AOX) pathways is known to be modulated by abiotic stress conditions. The effect of salinity stress on the mitochondrial electron transport chain and the importance of COX and AOX pathways for optimization of photosynthesis under salinity stress conditions is not clearly understood. In the current study, importance of COX and AOX pathways for photosynthetic performance of pea plants (Pisum sativum L. Pea Arkel cv) was analysed by using the mitochondrial electron transport chain inhibitors Antimycin A (AA) and salicylhydroxamic acid (SHAM) which restrict the electron flow through COX and AOX pathways respectively. Salinity stress resulted in decreased CO2 assimilation rates, leaf stomatal conductance, transpiration and leaf intercellular CO2 concentration in a stress dependent manner. Superimposition of leaves of salt stressed plants with AA and SHAM caused cellular H2O2 and O2- accumulation along with cell death. Additionally, aggravation in decrease of CO2 assimilation rates, leaf stomatal conductance, transpiration and leaf intercellular CO2 concentration upon superimposition with AA and SHAM during salinity stress suggests the importance of mitochondrial oxidative electron transport for photosynthesis. Increased expression of AOX1a and AOX2 transcripts along with AOX protein levels indicated up regulation of AOX pathway in leaves during salinity stress. Chlorophyll fluorescence measurements revealed enhanced damage to Photosystem (PS) II in the presence of AA and SHAM during salinity stress. Results suggested the beneficial role of COX and AOX pathways for optimal photosynthetic performance in pea leaves during salinity stress conditions.

IQGAP-related Rng2p organizes cortical nodes and ensures position of cell division in fission yeast.

Correct positioning of the cell division machinery is crucial for genomic stability and cell fate determination. The fission yeast Schizosaccharomyces pombe, like animal cells, divides using an actomyosin ring and is an attractive model to study eukaryotic cytokinesis. In S. pombe, positioning of the actomyosin ring depends on the anillin-related protein Mid1p. Mid1p arrives first at the medial cortex and recruits actomyosin ring components to node-like structures, although how this is achieved is unknown. Here we show that the IQGAP-related protein Rng2p, an essential component of the actomyosin ring, is a key element downstream of Mid1p. Rng2p physically interacts with Mid1p and is required for the organization of other actomyosin ring components into cortical nodes. Failure of localization of Rng2p to the nodes prevents medial retention of Mid1p and leads to actomyosin ring assembly in a node-independent manner at nonmedial locations. We conclude that Mid1p recruits Rng2p to cortical nodes at the division site and that Rng2p, in turn, recruits other components of the actomyosin ring to cortical nodes, thereby ensuring correct placement of the division site.