Growth, Morphology and Respiratory Cost Responses to Salinity in the Mangrove Plant Rhizophora Stylosa Depend on Growth Temperature.

Mangrove plants, which have evolved to inhabit tidal flats, may adjust their physiological and morphological traits to optimize their growth in saline habitats. Furthermore, the confined distribution of mangroves within warm regions suggests that warm temperature is advantageous to their growth in saline environments. We analyzed growth, morphology and respiratory responses to moderate salinity and temperature in a mangrove species, Rhizophora stylosa. The growth of R. stylosa was accelerated in moderate salinity compared with its growth in fresh water. Under warm conditions, the increased growth is accompanied by increased specific leaf area (SLA) and specific root length. Low temperature resulted in a low relative growth rate due to a low leaf area ratio and small SLA, regardless of salinity. Salinity lowered the ratio of the amounts of alternative oxidase to cytochrome c oxidase in the mitochondrial respiratory chain in leaves. Salinity enhanced the leaf respiration rate for maintenance, but under warm conditions this enhancement was compensated by a low leaf respiration rate for growth. In contrast, salinity enhanced overall leaf respiration rates at low temperature. Our results indicate that under moderate saline conditions R. stylosa leaves require warm temperatures to grow with a high rate of resource acquisition without enhancing respiratory cost.

Alternative Oxidase Alleviates Mitochondrial Oxidative Stress during Limited Nitrate Reduction in Arabidopsis thaliana.

The conversion of nitrate to ammonium, i.e., nitrate reduction, is a major consumer of reductants in plants. Previous studies have reported that the mitochondrial alternative oxidase (AOX) is upregulated under limited nitrate reduction conditions, including no/low nitrate or when ammonium is the sole nitrogen (N) source. Electron transfer from ubiquinone to AOX bypasses the proton-pumping complexes III and IV, thereby consuming reductants efficiently. Thus, upregulated AOX under limited nitrate reduction may dissipate excessive reductants and thereby attenuate oxidative stress. Nevertheless, so far there is no firm evidence for this hypothesis due to the lack of experimental systems to analyze the direct relationship between nitrate reduction and AOX. We therefore developed a novel culturing system for A. thaliana that manipulates shoot activities of nitrate reduction and AOX separately without causing N starvation, ammonium toxicity, or lack of nitrate signal. Using shoots processed with this system, we examined genome-wide gene expression and growth to better understand the relationship between AOX and nitrate reduction. The results showed that, only when nitrate reduction was limited, AOX deficiency significantly upregulated genes involved in mitochondrial oxidative stress, reductant shuttles, and non-phosphorylating bypasses of the respiratory chain, and inhibited growth. Thus, we conclude that AOX alleviates mitochondrial oxidative stress and sustains plant growth under limited nitrate reduction.

Cooperation of an external carbonic anhydrase and HCO3- transporter supports underwater photosynthesis in submerged leaves of the amphibious plant Hygrophila difformis.

BACKGROUND AND AIMS: HCO3- can be a major carbon resource for photosynthesis in underwater environments. Here we investigate the underlying mechanism of uptake and membrane transport of HCO3- in submerged leaves of Hygrophila difformis, a heterophyllous amphibious plant. To characterize these mechanisms, we evaluated the sensitivity of underwater photosynthesis to an external carbonic anhydrase (CA) inhibitor and an anion exchanger protein inhibitor, and we attempted to identify components of the mechanism of HCO3- utilization. METHODS: We evaluated the effects of the external CA inhibitor and anion exchanger protein inhibitor on the NaHCO3 response of photosynthetic O2 evolution in submerged leaves of H. difformis. Furthermore, we performed a comparative transcriptomic analysis between terrestrial and submerged leaves. KEY RESULTS: Photosynthesis in the submerged leaves was decreased by both the external CA inhibitor and anion exchanger protein inhibitor, but no additive effect was observed. Among upregulated genes in submerged leaves, two α-CAs, Hdα-CA1 and Hdα-CA2, and one ß-carbonic anhydrase, Hdß-CA1, were detected. Based on their putative amino acid sequences, the α-CAs are predicted to be localized in the apoplastic region. Recombinant Hdα-CA1 and Hdß-CA1 showed dominant CO2 hydration activity over HCO3- dehydration activity. CONCLUSIONS: We propose that the use of HCO3- for photosynthesis in submerged leaves of H. difformis is driven by the cooperation between an external CA, Hdα-CA1, and an unidentified HCO3- transporter.

Loss of peroxisomal NAD kinase 3 (NADK3) affects photorespiration metabolism in Arabidopsis.

Nicotinamide adenine dinucleotides (NAD+ and NADP+) are electron mediators involved in various metabolic pathways. NADP(H) are produced by NAD kinase (NADK) through the phosphorylation of NAD(H). The Arabidopsis NADK3 (AtNADK3) is reported to preferentially phosphorylate NADH to NADPH and is localized in the peroxisome. To elucidate the biological function of AtNADK3 in Arabidopsis, we compared metabolites of nadk1, nadk2 and nadk3 Arabidopsis T-DNA inserted mutants. Metabolome analysis revealed that glycine and serine, which are intermediate metabolites of photorespiration, both increased in the nadk3 mutants. Plants grown for 6 weeks under short-day conditions showed increased NAD(H), indicating a decrease in the phosphorylation ratio in the NAD(P)(H) equilibrium. Furthermore, high CO2 (0.15%) treatment induced a decrease in glycine and serine in nadk3 mutants. The nadk3 showed a significant decrease in post-illumination CO2 burst, suggesting that the photorespiratory flux was disrupted in the nadk3 mutant. In addition, an increase in CO2 compensation points and a decrease in CO2 assimilation rate were observed in the nadk3 mutants. These results indicate that the lack of AtNADK3 causes a disruption in the intracellular metabolism, such as in amino acid synthesis and photorespiration.

Dynamic seasonal changes in photosynthesis systems in leaves of Asarum tamaense, an evergreen understorey herbaceous species.

BACKGROUND AND AIMS: Evergreen herbaceous species in the deciduous forest understorey maintain their photosystems in long-lived leaves under dynamic seasonal changes in light and temperature. However, in evergreen understorey herbs, it is unknown how photosynthetic electron transport acclimates to seasonal changes in forest understorey environments, and what photoprotection systems function in excess energy dissipation under high-light and low-temperature environments in winter. METHODS: Here, we used Asarum tamaense, an evergreen herbaceous species in the deciduous forest understorey with a single-flush and long-lived leaves, and measured photosynthetic CO2 assimilation and electron transport in leaves throughout the year. The contents of photosynthetic proteins, pigments and primary metabolites were determined from regularly collected leaves. KEY RESULTS: Both the rates of CO2 assimilation and electron transport under saturated light were kept low in summer, but increased in autumn and winter in A. tamaense leaves. Although the contents of photosynthetic proteins including Rubisco did not increase in autumn and winter, the proton motive force and ΔpH across the thylakoid membrane were high in summer and decreased from summer to winter to a great extent. These decreases alleviated the suppression by lumen acidification and increased the electron transport rate in winter. The content and composition of carotenoids changed seasonally, which may affect changes in non-photochemical quenching from summer to winter. Winter leaves accumulated proline and malate, which may support cold acclimation. CONCLUSIONS: In A. tamaense leaves, the increase in photosynthetic electron transport rates in winter was not due to an increase in photosynthetic enzyme contents, but due to the activation of photosynthetic enzymes and/or release of limitation of photosynthetic electron flow. These seasonal changes in the regulation of electron transport and also the changes in several photoprotection systems should support the acclimation of photosynthetic C gain under dynamic environmental changes throughout the year.

Tight relationship between two photosystems is robust in rice leaves under various nitrogen conditions.

Leaf nitrogen (N) level affects not only photosynthetic CO2 assimilation, but also two photosystems of the photosynthetic electron transport. The quantum yield of photosystem II [Y(II)] and the non-photochemical yield due to the donor side limitation of photosystem I [Y(ND)], which denotes the fraction of oxidized P700 (P700+) to total P700, oppositely change depending on leaf N level, and the negative correlation between these two parameters has been reported in leaves of plants cultivated at various N levels in growth chambers. Here, we aimed to clarify whether this correlation is maintained after short-term changes in leaf N level, and what parameters are the most responsive to the changes in leaf N level under field conditions. We cultivated rice varieties at two N fertilization levels in paddy fields, treated additional N fertilization to plants grown at low N, and measured parameters of two photosystems of mature leaves. In rice leaves under low N condition, the Y(ND) increased and the photosynthetic linear electron flow was suppressed. In this situation, the accumulation of P700+ can function as excess energy dissipation. After the N addition, both Y(ND) and Y(II) changed, and the negative correlation between them was maintained. We used a newly-developed device to assess the photosystems. This device detected the similar changes in Y(ND) after the N addition, and the negative correlation between Y(ND) and photosynthetic O2 evolution rates was observed in plants under various N conditions. This study has provided strong field evidence that the Y(ND) largely changes depending on leaf N level, and that the Y(II) and Y(ND) are negatively correlated with each other irrespective of leaf N level, varieties and annual variation. The Y(ND) can stably monitor the leaf N status and the linear electron flow under field conditions.

Temperature dependence of O2 respiration in mangrove leaves and roots: implications for seedling dispersal phenology.

Seasonal differences in diaspore dispersal of three mangrove species, Kandelia obovata, Bruguiera gymnorrhiza and Rhizophora stylosa, suggest that respiratory energy production and demand may differ as a result of interspecific differences in temperature dependence of growth and maintenance processes during seedling establishment. We analyzed growth, temperature dependencies of respiratory O2 consumption and amounts of respiratory chain enzymes in seedlings of these species grown at various temperatures. Respiration rates measured at the low reference temperature, RREF , were highest in leaves of 15°C-grown K. obovata, whose dispersal occurs in the cold season, while root RREF of 15°C-grown R. stylosa was 60% those of the other species, possibly because of warm conditions during its establishment phase. In leaves and roots of K. obovata and leaves of R. stylosa, the overall activation energy, Eo , changed with growth temperature associated with changes in the ratios of the amount of protein in the two respiratory pathways. However, Eo of seedlings of B. gymnorrhiza, which has a long dispersal phase, were constant and independent of growth temperature. The different temperature responses of seedling respiration and growth among these three species may reflect the seasonal temperature range of seedling dispersal and establishment in each species.

Cost-benefit analysis of mesophyll conductance: diversities of anatomical, biochemical and environmental determinants.

BACKGROUND: Plants invest photosynthates in construction and maintenance of their structures and functions. Such investments are considered costs. These costs are recovered by the CO2 assimilation rate (A) in the leaves, and thus A is regarded as the immediate, short-term benefit. In photosynthesizing leaves, CO2 diffusion from the air to the carboxylation site is hindered by several structural and biochemical barriers. CO2 diffusion from the intercellular air space to the chloroplast stroma is obstructed by the mesophyll resistance. The inverses is the mesophyll conductance (gm). Whether various plants realize an optimal gm, and how much investment is needed for a relevant gm, remain unsolved. SCOPE: This review examines relationships among leaf construction costs (CC), leaf maintenance costs (MC) and gm in various plants under diverse growth conditions. Through a literature survey, we demonstrate a strong linear relationship between leaf mass per area (LMA) and leaf CC. The overall correlation of CC vs. gm across plant phylogenetic groups is weak, but significant trends are evident within specific groups and/or environments. Investment in CC is necessary for an increase in LMA and mesophyll cell surface area (Smes). This allows the leaf to accommodate more chloroplasts, thus increasing A. However, increases in LMA and/or Smes often accompany other changes, such as cell wall thickening, which diminishes gm. Such factors that make the correlations of CC and gm elusive are identified. CONCLUSIONS: For evaluation of the contribution of gm to recover CC, leaf life span is the key factor. The estimation of MC in relation to gm, especially in terms of costs required to regulate aquaporins, could be essential for efficient control of gm over the short term. Over the long term, costs are mainly reflected in CC, while benefits also include ultimate fitness attributes in terms of integrated carbon gain over the life of a leaf, plant survival and reproductive output.

Low N level increases the susceptibility of PSI to photoinhibition induced by short repetitive flashes in leaves of different rice varieties.

The recovery from photoinhibition is much slower in photosystem (PS) I than in PSII; therefore, the susceptibility of PSI to photoinhibition is important with respect to photosynthetic production under special physiological conditions. Previous studies have shown that repetitive short-pulse (rSP) illumination selectively induces PSI photoinhibition. Depending on the growth light intensity or the variety/species of the plant, PSI photoinhibition is different, but the underlying mechanisms remain unknown. Here, we aimed to clarify whether the differences in the susceptibility of PSI to photoinhibition depend on environmental factors or on rice varieties and which physiological properties of the plant are related to this susceptibility. We exposed mature leaves of rice plants to rSP illumination. We examined the effects of elevated CO2 concentration and low N during growth on the susceptibility of PSI to photoinhibition and compared it in 12 different varieties. We fitted the decrease in the quantum yield of PSI during rSP illumination and estimated a parameter indicating susceptibility. Low N level increased susceptibility, whereas elevated CO2 concentration did not. The susceptibility differed among different rice varieties, and many indica varieties showed higher susceptibility than the temperate japonica varieties. Susceptibility was negatively correlated with the total chlorophyll content and N content. However, the decrease in P m ' value, an indicator of damaged PSI, was positively correlated with chlorophyll content. This suggests that in leaves with a larger electron transport capacity, the overall PSI activity may be less susceptible to photoinhibition, but more damaged PSI may accumulate during rSP illumination.

Growth temperature affects O2 consumption rates and plasticity of respiratory flux to support shoot growth at various growth temperatures.

The temperature dependence of respiration rates and their acclimation to growth temperature vary among species/ecotypes, but the details remain unclear. Here, we compared the temperature dependence of shoot O2 consumption rates among Arabidopsis thaliana ecotypes to clarify how the temperature dependence and their acclimation to temperature differ among ecotypes, and how these differences relate to shoot growth. We examined growth analysis, temperature dependence of O2 consumption rates, and protein amounts of the respiratory chain components in shoots of twelve ecotypes of A. thaliana grown at three different temperatures. The temperature dependence of the O2 consumption rates were fitted to the modified Arrhenius model. The dynamic response of activation energy to measurement temperature was different among growth temperatures, suggesting that the plasticity of respiratory flux to temperatures differs among growth temperatures. The similar values of activation energy at growth temperature among ecotypes suggest that a similar process may determine the O2 consumption rates at the growth temperature in any ecotype. These results suggest that the growth temperature affects not only the absolute rate of O2 consumption but also the plasticity of respiratory flux in response to temperature, supporting the acclimation of shoot growth to various temperatures.

Distinct responses of growth and respiration to growth temperatures in two mangrove species.

BACKGROUND AND AIMS: Mangrove plants are mostly found in tropical and sub-tropical tidal flats, and their limited distribution may be related to their responses to growth temperatures. However, the mechanisms underlying these responses have not been clarified. Here, we measured the dependencies of the growth parameters and respiration rates of leaves and roots on growth temperatures in typical mangrove species. METHODS: We grew two typical species of Indo-Pacific mangroves, Bruguiera gymnorrhiza and Rhizophora stylosa, at four different temperatures (15, 20, 25 and 30 °C) by irrigating with fresh water containing nutrients, and we measured growth parameters, chemical composition, and leaf and root O2 respiration rates. We then estimated the construction costs of leaves and roots and the respiration rates required for maintenance and growth. KEY RESULTS: The relative growth rates of both species increased with growth temperature due to changes in physiological parameters such as net assimilation rate and respiration rate rather than to changes in structural parameters such as leaf area ratio. Both species required a threshold temperature for growth (12.2 °C in B. gymnorrhiza and 18.1 °C in R. stylosa). At the low growth temperature, root nitrogen uptake rate was lower in R. stylosa than in B. gymnorrhiza, leading to a slower growth rate in R. stylosa. This indicates that R. stylosa is more sensitive than B. gymnorrhiza to low temperature. CONCLUSIONS: Our results suggest that the mangrove species require a certain warm temperature to ensure respiration rates sufficient for maintenance and growth, particularly in roots. The underground temperature probably limits their growth under the low-temperature condition. The lower sensitivity of B. gymnorrhiza to low temperature shows its potential to adapt to a wider habitat temperature range than R. stylosa. These growth and respiratory features may explain the distribution patterns of the two mangrove species.

Manganese toxicity disrupts indole acetic acid homeostasis and suppresses the CO2 assimilation reaction in rice leaves.

Despite the essentiality of Mn in terrestrial plants, its excessive accumulation in plant tissues can cause growth defects, known as Mn toxicity. Mn toxicity can be classified into apoplastic and symplastic types depending on its onset. Symplastic Mn toxicity is hypothesised to be more critical for growth defects. However, details of the relationship between growth defects and symplastic Mn toxicity remain elusive. In this study, we aimed to elucidate the molecular mechanisms underlying symplastic Mn toxicity in rice plants. We found that under excess Mn conditions, CO2 assimilation was inhibited by stomatal closure, and both carbon anabolic and catabolic activities were decreased. In addition to stomatal dysfunction, stomatal and leaf anatomical development were also altered by excess Mn accumulation. Furthermore, indole acetic acid (IAA) concentration was decreased, and auxin-responsive gene expression analyses showed IAA-deficient symptoms in leaves due to excess Mn accumulation. These results suggest that excessive Mn accumulation causes IAA deficiency, and low IAA concentrations suppress plant growth by suppressing stomatal opening and leaf anatomical development for efficient CO2 assimilation in leaves.

Photochemistry of Photosystems II and I in Rice Plants Grown under Different N Levels at Normal and High Temperature.

Although N levels affect leaf photosynthetic capacity, the effects of N levels on the photochemistry of photosystems II and I (PSII and PSI, respectively) are not well-understood. In the present study, we examined this aspect in rice (Oryza sativa L. 'Hitomebore') plants grown under three different N levels at normal or high temperatures that can occur during rice culture and do not severely suppress photosynthesis. At both growth temperatures, the quantum efficiency of PSII [Y(II)] and the fraction of the primary quinone electron acceptor in its oxidized state were positively correlated with the amount of total leaf-N, whereas the quantum yields of non-photochemical quenching and donor-side limitation of PSI [Y(ND)] were negatively correlated with the amount of total leaf-N. These changes in PSII and PSI parameters were strongly correlated with each other. Growth temperatures scarcely affected these relationships. These results suggest that the photochemistry of PSII and PSI is coordinately regulated primarily depending on the amount of total leaf-N. When excess light energy occurs in low N-acclimated plants, oxidation of the reaction center chlorophyll of PSI is thought to be stimulated to protect PSI from excess light energy. It is also suggested that PSII and PSI normally operate at high temperature used in the present study. In addition, as the relationships between Y(II) and Y(ND) were found to be almost identical to those observed in osmotically stressed rice plants, common regulation is thought to be operative when excess light energy occurs due to different causes.

Temperature-dependent fasciation mutants provide a link between mitochondrial RNA processing and lateral root morphogenesis.

Although mechanisms that activate organogenesis in plants are well established, much less is known about the subsequent fine-tuning of cell proliferation, which is crucial for creating properly structured and sized organs. Here we show, through analysis of temperature-dependent fasciation (TDF) mutants of Arabidopsis, root redifferentiation defective 1 (rrd1), rrd2, and root initiation defective 4 (rid4), that mitochondrial RNA processing is required for limiting cell division during early lateral root (LR) organogenesis. These mutants formed abnormally broadened (i.e. fasciated) LRs under high-temperature conditions due to extra cell division. All TDF proteins localized to mitochondria, where they were found to participate in RNA processing: RRD1 in mRNA deadenylation, and RRD2 and RID4 in mRNA editing. Further analysis suggested that LR fasciation in the TDF mutants is triggered by reactive oxygen species generation caused by defective mitochondrial respiration. Our findings provide novel clues for the physiological significance of mitochondrial activities in plant organogenesis.

Effects of root morphology, respiration and carboxylate exudation on carbon economy in two non-mycorrhizal lupines under phosphorus deficiency.

Under phosphorus (P) deficiency, Lupinus albus develops cluster roots that allow efficient P acquisition, while L. angustifolius without cluster roots also grows well. Both species are non-mycorrhizal. We quantitatively examined the carbon budgets to investigate the different strategies of these species. Biomass allocation, respiratory rates, protein amounts and carboxylate exudation rates were examined in hydroponically-grown plants treated with low (1 µM; P1) or high (100 µM; P100) P. At P1, L. albus formed cluster roots, and L. angustifolius increased biomass allocation to the roots. The respiratory rates of the roots were faster in L. albus than in L. angustifolius. The protein amounts of the non-phosphorylating alternative oxidase and uncoupling protein were greater in the cluster roots of L. albus at P1 than in the roots at P100, but similar between the P treatments in L. angustifolius roots. At P1, L. albus exuded carboxylates at a faster rate than L. angustifolius. The carbon budgets at P1 were surprisingly similar between the two species, which is attributed to the contrasting root growth and development strategies. L. albus developed cluster roots with rapid respiratory and carboxylate exudation rates, while L. angustifolius developed a larger root system with slow respiratory and exudation rates.

Theoretical analysis of a temperature-dependent model of respiratory O2 consumption using the kinetics of the cytochrome and alternative pathways.

Temperature dependence of plant respiratory O2 -consumption has been empirically described by the Arrhenius equation. The slope of the Arrhenius plot (which is proportional to activation energy) sometimes deviates from a constant value. We conducted kinetic model simulations of mitochondrial electron flow dynamics to clarify factors affecting the shape of the Arrhenius plot. We constructed a kinetic model of respiration in which competitive O2 -consumption by the cytochrome pathway (CP) and the alternative pathway (AP) were considered, and we used this model to describe the temperature dependence of respiratory O2 -consumption of Arabidopsis. The model indicated that the electron partitioning and activation energy differences between CP and AP were reflected in the slope and magnitude of the dependent variables of the Arrhenius plot. When the electron partitioning and activation energies of CP and AP were constant with temperature change, our model suggested that the Arrhenius plot would be almost linear. When the electron partitioning or activation energy of CP, or both, rapidly changed with temperature, the Arrhenius plot deviated from linearity, as reported in previous experimental studies. Our simulation analysis quantitatively linked the kinetic model parameters with physiological mechanisms underlying the instantaneous temperature dependence of plant respiration rate.

Atmospheric CO2 Concentration and N Availability Affect the Balance of the Two Photosystems in Mature Leaves of Rice Plants Grown at a Free-Air CO2 Enrichment Site.

Atmospheric CO2 concentration ([CO2]) has been substantially increasing. Responses of leaf photosynthesis to elevated [CO2] have been intensively investigated because leaf photosynthesis is one of the most important determinants of crop yield. The responses of photosynthesis to elevated [CO2] can depend on nitrogen (N) availability. Here, we aimed to investigate the significance of the appropriate balance between two photosystems [photosystem I (PSI) and photosystem II (PSII)] under various [CO2] and N levels, and thus to clarify if responses of photosynthetic electron transport rates (ETRs) of the two photosystems to elevated [CO2] are altered by N availability. Thus, we examined parameters of the two photosystems in mature leaves of rice plants grown under two [CO2] levels (ambient and 200 µmol mol-1 above ambient) and three N fertilization levels at the Tsukuba free-air CO2 enrichment experimental facility in Japan. Responses of ETR of PSII (ETRII) and ETR of PSI (ETRI) to [CO2] levels differed among N levels. When moderate levels of N were applied (MN), ETRI was higher under elevated [CO2], whereas at high levels of N were applied (HN), both ETRII and ETRI were lower under elevated [CO2] compared with ambient [CO2]. Under HN, the decreases in ETRII and ETRI under elevated [CO2] were due to increases in the non-photochemical quenching of PSII [Y(NPQ)] and the donor side limitation of PSI [Y(ND)], respectively. The relationship between the effective quantum yields of PSI [Y(I)] and PSII [Y(II)] changed under elevated [CO2] and low levels of N (LN). Under both conditions, the ratio of Y(I) to Y(II) was higher than under other conditions. The elevated [CO2] and low N changed the balance of the two photosystems. This change may be important because it can induce the cyclic electron flow around PSI, leading to induction of non-photochemical quenching to avoid photoinhibition.

Rare Neurologic Disease-Associated Mutations of AIMP1 are Related with Inhibitory Neuronal Differentiation Which is Reversed by Ibuprofen.

BACKGROUND: Hypomyelinating leukodystrophy 3 (HLD3), previously characterized as a congenital diseases associated with oligodendrocyte myelination, is increasingly regarded as primarily affecting neuronal cells. METHODS: We used N1E-115 cells as the neuronal cell model to investigate whether HLD3-associated mutant proteins of cytoplasmic aminoacyl-tRNA synthase complex-interacting multifunctional protein 1 (AIMP1) aggregate in organelles and affect neuronal differentiation. RESULTS: 292CA frame-shift type mutant proteins harboring a two-base (CA) deletion at the 292th nucleotide are mainly localized in the lysosome where they form aggregates. Similar results are observed in mutant proteins harboring the Gln39-to-Ter (Q39X) mutation. Interestingly, the frame-shift mutant-specific peptide specifically interacts with actin to block actin fiber formation. The presence of actin with 292CA mutant proteins, but not with wild type or Q39X ones, in the lysosome is detectable by immunoprecipitation of the lysosome. Furthermore, expression of 292CA or Q39X mutants in cells inhibits neuronal differentiation. Treatment with ibuprofen reverses mutant-mediated inhibitory differentiation as well as the localization in the lysosome. CONCLUSIONS: These results not only explain the cell pathological mechanisms inhibiting phenotype differentiation in cells expressing HLD3-associated mutants but also identify the first chemical that restores such cells in vitro.