JAK inhibitor treatment for inborn errors of JAK/STAT signaling: An ESID/EBMT-IEWP retrospective study.

BACKGROUND: Inborn errors of immunity (IEI) with dysregulated JAK/STAT signaling present with variable manifestations of immune dysregulation and infections. Hematopoietic stem cell transplantation (HSCT) is potentially curative, but initially reported outcomes were poor. JAK inhibitors (JAKi) offer a targeted treatment option that may be an alternative or bridge to HSCT. However, data on their current use, treatment efficacy and adverse events are limited. OBJECTIVE: We evaluated the current off-label JAKi treatment experience for JAK/STAT inborn errors of immunity (IEI) among European Society for Immunodeficiencies (ESID)/European Society for Blood and Marrow Transplantation (EBMT) Inborn Errors Working Party (IEWP) centers. METHODS: We conducted a multicenter retrospective study on patients with a genetic disorder of hyperactive JAK/STAT signaling who received JAKi treatment for at least 3 months. RESULTS: Sixty-nine patients (72% children) were evaluated (45 STAT1 gain of function [GOF], 21 STAT3-GOF, 1 STAT5B-GOF, 1 suppressor of cytokine signaling 1 [aka SOCS1] loss of function, 1 JAK1-GOF). Ruxolitinib was the predominantly prescribed JAKi (80%). Overall, treatment resulted in improvement (partial or complete remission) of clinical symptoms in 87% of STAT1-GOF and in 90% of STAT3-GOF patients. We documented highly heterogeneous dosing and monitoring regimens. The response rate and time to response varied across different diseases and manifestations. Adverse events including infection and weight gain were frequent (38% of patients) but were mild (grade I-II) and transient in most patients. At last follow-up, 52 (74%) of 69 patients were still receiving JAKi treatment, and 11 patients eventually underwent HSCT after receipt of previous JAKi bridging therapy, with 91% overall survival. CONCLUSIONS: Our study suggests that JAKi may be highly effective to treat symptomatic JAK/STAT IEI patients. Prospective studies to define optimal JAKi dosing for the variable clinical presentations and age ranges should be pursued.

Abnormal biomarkers predict complex FAS or FADD defects missed by exome sequencing.

BACKGROUND: Elevated TCRαß+CD4-CD8- double-negative T cells (DNT) and serum biomarkers help identify FAS mutant patients with autoimmune lymphoproliferative syndrome (ALPS). However, in some patients with clinical features and biomarkers consistent with ALPS, germline or somatic FAS mutations cannot be identified on standard exon sequencing (ALPS-undetermined: ALPS-U). OBJECTIVE: We sought to explore whether complex genetic alterations in the FAS gene escaping standard sequencing or mutations in other FAS pathway-related genes could explain these cases. METHODS: Genetic analysis included whole FAS gene sequencing, copy number variation analysis, and sequencing of FAS cDNA and other FAS pathway-related genes. It was guided by FAS expression analysis on CD57+DNT, which can predict somatic loss of heterozygosity (sLOH). RESULTS: Nine of 16 patients with ALPS-U lacked FAS expression on CD57+DNT predicting heterozygous "loss-of-expression" FAS mutations plus acquired somatic second hits in the FAS gene, enriched in DNT. Indeed, 7 of 9 analyzed patients carried deep intronic mutations or large deletions in the FAS gene combined with sLOH detectable in DNT; 1 patient showed a FAS exon duplication. Three patients had reduced FAS expression, and 2 of them harbored mutations in the FAS promoter, which reduced FAS expression in reporter assays. Three of the 4 ALPS-U patients with normal FAS expression carried heterozygous FADD mutations with sLOH. CONCLUSION: A combination of serum biomarkers and DNT phenotyping is an accurate means to identify patients with ALPS who are missed by routine exome sequencing.

Injuries and Use of Safety Equipment in Stand-up Paddle Boarding.

BACKGROUND: Stand-up paddleboarding (SUP) is a water sport that has gained in popularity. Still, very little is known about its injury profile. PURPOSE: To analyze the incidence, mechanisms, and risk factors for SUP-related injuries in mainly calm waters and the use of safety equipment. STUDY DESIGN: Descriptive epidemiology study. METHODS: The authors distributed an online survey that gathered information on characteristics, injury history over the past 12 months, use of (safety) equipment, and health issues of SUP riders in German-speaking countries. RESULTS: A total of 438 participants completed the survey. The mean participant age was 45 years (range, 8-82 years), 48% were female athletes, and 19.6% took part in competitions. Over the past 12 months, 72 (17%) of 431 respondents experienced a SUP-related injury, resulting in an overall incidence rate (IR) of 1.95 injuries per 1000 hours of activity. Male competitive athletes using a hardboard had the highest risk of injury. The IR for competitive riders was higher (3.21 injuries per 1000 hours) than that for recreational riders (1.41 injuries per 1000 hours). Mechanisms of injury were overuse (n = 29) and contact with the board (n = 18) or ground (n = 12). Injuries affected mainly muscles/tendons and joints of the upper arm/shoulder (n = 20), followed by wrist/hand (n = 18), knee (n = 16), elbow/forearm (n = 12), and foot (n = 11). The highest risk of injury occurred when paddling in wild water (33.3%), followed by SUP surfing (29.4%), and the lowest risk occurred while paddling on a lake (16.1%). A life vest was always used by 27, depending on the situation by 201, and never by 202 participants; a leash was always used by 161, depending on the situation by 244, and never by 26 participants. Of the 272 participants who paddled in winter, 253 wore a drysuit or wetsuit, whereas 19 paddled without any special clothing. CONCLUSION: SUP-related injuries predominantly happened in wild water or while SUP surfing, and male competitive athletes using a hardboard had the highest risk of injury. The main mechanisms of injury were overuse and hitting the board or ground. The upper extremity was more prone to injury, followed by the knee and foot. More effort should be made to educate paddlers on the necessity and correct use of the life vest, leash, and specialized outfits.

Co-purification of nitrate reductase 1 with components of the cytochrome bcc-aa3 oxidase supercomplex from spores of Streptomyces coelicolor A3(2).

In order to reduce nitrate in vivo, the spore-specific respiratory nitrate reductase, Nar1, of Streptomyces coelicolor relies on an active cytochrome bcc-aa3 oxidase supercomplex (bcc-aa3 supercomplex). This suggests that membrane-associated Nar1, comprising NarG1, NarH1, and NarI1 subunits, might not act as a classical menaquinol oxidase but could either receive electrons from the bcc-aa3 supercomplex, or require the supercomplex to stabilize the reductase in the membrane to allow it to function. To address the biochemical basis for this dependence on the bcc-aa3 supercomplex, we purified two different Strep-tagged variants of Nar1 and enriched the native enzyme complex from spore extracts using different chromatographic and electrophoretic procedures. Polypeptides associated with the isolated Nar1 complexes were identified using mass spectrometry and included components of the bcc-aa3 supercomplex, along with an alternative, spore-specific cytochrome b component, QcrB3. Surprisingly, we also co-enriched the Nar3 enzyme with Nar1 from the wild-type strain of S. coelicolor. Two differentially migrating active Nar1 complexes could be identified after clear native polyacrylamide gel electrophoresis; these had masses of approximately 450 and 250 kDa. The distribution of active Nar1 in these complexes was influenced by the presence of cytochrome bd oxidase and by QcrB3; the presence of the latter shifted Nar1 into the larger complex. Together, these data suggest that several respiratory complexes can associate in the spore membrane, including Nar1, Nar3, and the bcc-aa3 supercomplex. Moreover, these findings provide initial support for the hypothesis that Nar1 and the bcc-aa3 supercomplex physically associate.

Tumor-derived TGF-ß inhibits mitochondrial respiration to suppress IFN-γ production by human CD4+ T cells.

Transforming growth factor-ß (TGF-ß) is produced by tumors, and increased amounts of this cytokine in the tumor microenvironment and serum are associated with poor patient survival. TGF-ß-mediated suppression of antitumor T cell responses contributes to tumor growth and survival. However, TGF-ß also has tumor-suppressive activity; thus, dissecting cell type-specific molecular effects may inform therapeutic strategies targeting this cytokine. Here, using human peripheral and tumor-associated lymphocytes, we investigated how tumor-derived TGF-ß suppresses a key antitumor function of CD4+ T cells, interferon-γ (IFN-γ) production. Suppression required the expression and phosphorylation of Smad proteins in the TGF-ß signaling pathway, but not their nuclear translocation, and depended on oxygen availability, suggesting a metabolic basis for these effects. Smad proteins were detected in the mitochondria of CD4+ T cells, where they were phosphorylated upon treatment with TGF-ß. Phosphorylated Smad proteins were also detected in the mitochondria of isolated tumor-associated lymphocytes. TGF-ß substantially impaired the ATP-coupled respiration of CD4+ T cells and specifically inhibited mitochondrial complex V (ATP synthase) activity. Last, inhibition of ATP synthase alone was sufficient to impair IFN-γ production by CD4+ T cells. These results, which have implications for human antitumor immunity, suggest that TGF-ß targets T cell metabolism directly, thus diminishing T cell function through metabolic paralysis.

Hypoxia-induced synthesis of respiratory nitrate reductase 2 of Streptomyces coelicolor A3(2) depends on the histidine kinase OsdK in mycelium but not in spores.

The three nitrate reductases (Nar) of the saprophytic aerobic actinobacterium Streptomyces coelicolor A3(2) contribute to survival when oxygen becomes limiting. In the current study, we focused on synthesis of the Nar2 enzyme, which is the main Nar enzyme present and active in exponentially growing mycelium. Synthesis of Nar2 can, however, also be induced in spores after extended periods of anoxic incubation. The osdRK genes (oxygen stress and development) were recently identified to encode a two-component system important for expression of the nar2 operon in mycelium. OsdK is a predicted histidine kinase and we show here that an osdK mutant completely lacks Nar2 enzyme activity in mycelium. Recovery of Nar2 enzyme activity was achieved by re-introduction of the osdRK genes into the mutant on an integrative plasmid. In anoxically incubated spores, however, the osdK mutant retained the ability to synthesize NarG2, the catalytic subunit of Nar2. We could also demonstrate that synthesis of NarG2 in spores occurred only under hypoxic conditions; anoxia, as well as O2 concentrations significantly higher than 1 % in the gas-phase, failed to result in induction of NarG2 synthesis. Together, these findings indicate that, although Nar2 synthesis in both mycelium and spores is induced by oxygen limitation, different mechanisms control these processes and only Nar2 synthesis in mycelium is under the control of the OsdKR two-component system.

Anaerobic nitrate respiration in the aerobe Streptomyces coelicolor A3(2): helping maintain a proton gradient during dormancy.

Respiratory nitrate reductases (Nar) catalyse the reduction of nitrate to nitrite, coupling this process to energy conservation. The obligate aerobic actinobacterium Streptomyces coelicolor synthesizes three Nar enzymes that contribute to maintenance of a membrane potential when either the mycelium or the spores become hypoxic or anoxic. No growth occurs under such conditions but the bacterium survives the lack of O2 by remaining metabolically active; reducing nitrate is one means whereby this process is aided. Nar1 is exclusive to spores, Nar2 to vegetative mycelium and Nar3 to stationary-phase mycelium, each making a distinct contribution to energy conservation. While Nar2 and Nar3 appear to function like conventional menaquinol oxidases, unusually, Nar1 is completely dependent for its activity on a cytochrome bcc-aa 3 oxidase supercomplex. This suggest that electrons within this supercomplex are diverted to Nar1 during O2 limitation. Receiving electrons from this supercomplex potentially allows nitrate reduction to be coupled to the Q-cycle of the cytochrome bcc complex. This modification likely improves the efficiency of energy conservation, extending longevity of spores under O2 limitation. Knowledge gained on the bioenergetics of NO3 - respiration in the actinobacteria will aid our understanding of how many microorganisms survive under conditions of extreme nutrient and energy restriction.

Activity of Spore-Specific Respiratory Nitrate Reductase 1 of Streptomyces coelicolor A3(2) Requires a Functional Cytochrome bcc-aa3 Oxidase Supercomplex.

Spores have strongly reduced metabolic activity and are produced during the complex developmental cycle of the actinobacterium Streptomyces coelicolor Resting spores can remain viable for decades, yet little is known about how they conserve energy. It is known, however, that they can reduce either oxygen or nitrate using endogenous electron sources. S. coelicolor uses either a cytochrome bd oxidase or a cytochrome bcc-aa3 oxidase supercomplex to reduce oxygen, while nitrate is reduced by Nar-type nitrate reductases, which typically oxidize quinol directly. Here, we show that in resting spores the Nar1 nitrate reductase requires a functional bcc-aa3 supercomplex to reduce nitrate. Mutants lacking the complete qcr-cta genetic locus encoding the bcc-aa3 supercomplex showed no Nar1-dependent nitrate reduction. Recovery of Nar1 activity was achieved by genetic complementation but only when the complete qcr-cta locus was reintroduced to the mutant strain. We could exclude that the dependence on the supercomplex for nitrate reduction was via regulation of nitrate transport. Moreover, the catalytic subunit, NarG1, of Nar1 was synthesized in the qcr-cta mutant, ruling out transcriptional control. Constitutive synthesis of Nar1 in mycelium revealed that the enzyme was poorly active in this compartment, suggesting that the Nar1 enzyme cannot act as a typical quinol oxidase. Notably, nitrate reduction by the Nar2 enzyme, which is active in growing mycelium, was not wholly dependent on the bcc-aa3 supercomplex for activity. Together, our data suggest that Nar1 functions together with the proton-translocating bcc-aa3 supercomplex to increase the efficiency of energy conservation in resting spores.IMPORTANCEStreptomyces coelicolor forms spores that respire with either oxygen or nitrate, using only endogenous electron donors. This helps maintain a membrane potential and, thus, viability. Respiratory nitrate reductase (Nar) usually receives electrons directly from reduced quinone species; however, we show that nitrate respiration in spores requires a respiratory supercomplex comprising cytochrome bcc oxidoreductase and aa3 oxidase. Our findings suggest that the Nar1 enzyme in the S. coelicolor spore functions together with the proton-translocating bcc-aa3 supercomplex to help maintain the membrane potential more efficiently. Dissecting the mechanisms underlying this survival strategy is important for our general understanding of bacterial persistence during infection processes and of how bacteria might deal with nutrient limitation in the natural environment.

Plasmonic mid-infrared third harmonic generation in germanium nanoantennas.

We demonstrate third harmonic generation in plasmonic antennas consisting of highly doped germanium grown on silicon substrates and designed to be resonant in the mid-infrared frequency range that is inaccessible with conventional nonlinear plasmonic materials. Owing to the near-field enhancement, the result is an ultrafast, subdiffraction, coherent light source with a wavelength tunable between 3 and 5 µm, and ideally overlapping with the fingerprint region of molecular vibrations. To observe the nonlinearity in this challenging spectral window, a high-power femtosecond laser system equipped with parametric frequency conversion in combination with an all-reflective confocal microscope setup is employed. We demonstrate spatially resolved maps of the linear scattering cross section and the nonlinear emission of single isolated antenna structures. A clear third-order power dependence as well as mid-infrared emission spectra prove the nonlinear nature of the light emission. Simulations support the observed resonance length of the double-rod antenna and demonstrate that the field enhancement inside the antenna material is responsible for the nonlinear frequency mixing.

Early effector maturation of naïve human CD8+ T cells requires mitochondrial biogenesis.

The role of mitochondrial biogenesis during naïve to effector differentiation of CD8+ T cells remains ill explored. In this study, we describe a critical role for early mitochondrial biogenesis in supporting cytokine production of nascent activated human naïve CD8+ T cells. Specifically, we found that prior to the first round of cell division activated naïve CD8+ T cells rapidly increase mitochondrial mass, mitochondrial respiration, and mitochondrial reactive oxygen species (mROS) generation, which were all inter-linked and important for CD8+ T cell effector maturation. Inhibition of early mitochondrial biogenesis diminished mROS dependent IL-2 production - as well as subsequent IL-2 dependent TNF, IFN-γ, perforin, and granzyme B production. Together, these findings point to the importance of mitochondrial biogenesis during early effector maturation of CD8+ T cells.

Cytochrome bd Oxidase Has an Important Role in Sustaining Growth and Development of Streptomyces coelicolor A3(2) under Oxygen-Limiting Conditions.

Streptomyces coelicolor A3(2) is a filamentously growing, spore-forming, obligately aerobic actinobacterium that uses both a copper aa3 -type cytochrome c oxidase and a cytochrome bd oxidase to respire oxygen. Using defined knockout mutants, we demonstrated that either of these terminal oxidases was capable of allowing the bacterium to grow and complete its developmental cycle. The genes encoding the bcc complex and the aa3 oxidase are clustered at a single locus. Using Western blot analyses, we showed that the bcc-aa3 oxidase branch is more prevalent in spores than the bd oxidase. The level of the catalytic subunit, CydA, of the bd oxidase was low in spore extracts derived from the wild type, but it was upregulated in a mutant lacking the bcc-aa3 supercomplex. This indicates that cytochrome bd oxidase can compensate for the lack of the other respiratory branch. Components of both oxidases were abundant in growing mycelium. Growth studies in liquid medium revealed that a mutant lacking the bcc-aa3 oxidase branch grew approximately half as fast as the wild type, while the oxygen reduction rate of the mutant remained close to that of the wild type, indicating that the bd oxidase was mainly functioning in controlling electron flux. Developmental defects were observed for a mutant lacking the cytochrome bd oxidase during growth on buffered rich medium plates with glucose as the energy substrate. Evidence based on using the redox-cycling dye methylene blue suggested that cytochrome bd oxidase is essential for the bacterium to grow and complete its developmental cycle under oxygen limitation.IMPORTANCE Respiring with oxygen is an efficient means of conserving energy in biological systems. The spore-forming, filamentous actinobacterium Streptomyces coelicolor grows only aerobically, synthesizing two enzyme complexes for O2 reduction, the cytochrome bcc-aa3 cytochrome oxidase supercomplex and the cytochrome bd oxidase. We show in this study that the bacterium can survive with either of these respiratory pathways to oxygen. Immunological studies indicate that the bcc-aa3 oxidase is the main oxidase present in spores, but the bd oxidase compensates if the bcc-aa3 oxidase is inactivated. Both oxidases are active in mycelia. Growth conditions were identified, revealing that cytochrome bd oxidase is essential for aerial hypha formation and sporulation, and this was linked to an important role of the enzyme under oxygen-limiting conditions.

Mitochondria-Endoplasmic Reticulum Contact Sites Function as Immunometabolic Hubs that Orchestrate the Rapid Recall Response of Memory CD8+ T Cells.

Glycolysis is linked to the rapid response of memory CD8+ T cells, but the molecular and subcellular structural elements enabling enhanced glucose metabolism in nascent activated memory CD8+ T cells are unknown. We found that rapid activation of protein kinase B (PKB or AKT) by mammalian target of rapamycin complex 2 (mTORC2) led to inhibition of glycogen synthase kinase 3ß (GSK3ß) at mitochondria-endoplasmic reticulum (ER) junctions. This enabled recruitment of hexokinase I (HK-I) to the voltage-dependent anion channel (VDAC) on mitochondria. Binding of HK-I to VDAC promoted respiration by facilitating metabolite flux into mitochondria. Glucose tracing pinpointed pyruvate oxidation in mitochondria, which was the metabolic requirement for rapid generation of interferon-γ (IFN-γ) in memory T cells. Subcellular organization of mTORC2-AKT-GSK3ß at mitochondria-ER contact sites, promoting HK-I recruitment to VDAC, thus underpins the metabolic reprogramming needed for memory CD8+ T cells to rapidly acquire effector function.

Cytochrome bcc-aa3 Oxidase Supercomplexes in the Aerobic Respiratory Chain of Streptomyces coelicolor A3(2).

Streptomyces coelicolor A3(2), an obligately aerobic, oxidase-positive, and filamentous soil bacterium, lacks a soluble cytochrome c in its respiratory chain, having instead a membrane-associated diheme c-type cytochrome, QcrC. This necessitates complex formation to allow electron transfer between the cytochrome bcc and aa3 oxidase respiratory complexes. Combining genetic complementation studies with in-gel cytochrome oxidase activity staining, we demonstrate that the complete qcrCAB-ctaCDFE gene locus on the chromosome, encoding, respectively, the bcc and aa3 complexes, is required to manifest a cytochrome oxidase enzyme activity in both spores and mycelium of a qcr-cta deletion mutant. Blue-native-PAGE identified a cytochrome aa3 oxidase complex of approximately 270 kDa, which catalyzed oxygen-dependent diaminobenzidine oxidation without the requirement for exogenously supplied cytochrome c, indicating association with QcrC. Furthermore, higher molecular mass complexes were identified upon addition of soluble cytochrome c, suggesting the supercomplex is unstable and readily dissociates into subcomplexes lacking QcrC. Immunological and mass spectrometric analyses of active, high-molecular mass oxidase-containing complexes separated by clear-native PAGE identified key subunits of both the bcc complex and the aa3 oxidase, supporting supercomplex formation. Our data also indicate that the cytochrome b QcrB of the bcc complex is less abundant in spores compared with mycelium.

Coherent field transients below 15 THz from phase-matched difference frequency generation in 4H-SiC.

We experimentally demonstrate tunable, phase-matched difference frequency generation covering the spectral region below 15 THz using 4H-SiC as a nonlinear crystal. This material combines a non-centrosymmetric lattice and strong birefringence with broadband transparency at low optical frequencies. Thorough refractive index measurements in the terahertz spectral range allow us to calculate phase-matching conditions for any near-infrared pump laser source. 4H-SiC is also exploited as a detector crystal for electro-optic sampling. The results allow us to estimate the effective second-order nonlinear coefficient.

Phosphate and oxygen limitation induce respiratory nitrate reductase 3 synthesis in stationary-phase mycelium of Streptomyces coelicolor A3(2).

The saprophytic actinobacterium Streptomyces coelicolor A3(2) requires oxygen for filamentous growth. Surprisingly, the bacterium also synthesizes three active respiratory nitrate reductases (Nar), which are believed to contribute to survival, or general fitness, of the bacterium in soil when oxygen becomes limiting. In this study, we analysed Nar3 and showed that activity of the enzyme is restricted to stationary-phase mycelium of S. coelicolor. Phosphate limitation was shown to be necessary for induction of enzyme synthesis. Nar3 synthesis was inhibited by inclusion of 20 mM phosphate in a defined 'switch assay' in which highly dispersed mycelium from exponentially growing cultures was shifted to neutral MOPS-glucose buffer to induce Nar3 synthesis and activity. Quantitative assessment of nar3 transcripts revealed a 30-fold induction of gene expression in stationary-phase mycelium. Transcript levels in stationary-phase mycelium incubated with phosphate were reduced by a little more than twofold, suggesting that the negative influence of phosphate on Nar3 synthesis was mainly at the post-transcriptional level. Furthermore, it was demonstrated that oxygen limitation was necessary to induce high levels of Nar3 activity. However, an abrupt shift from aerobic to anaerobic conditions prevented appearance of Nar3 activity. This suggests that the bacterium regulates Nar3 synthesis in response to the energy status of the mycelium. Nitrate had little impact on regulation of the Nar3 level. Together, these data identify Nar3 as a stationary-phase nitrate reductase in S. coelicolor and demonstrate that enzyme synthesis is induced in response to both phosphate limitation and hypoxia.

Optical Activation of Germanium Plasmonic Antennas in the Mid-Infrared.

Impulsive interband excitation with femtosecond near-infrared pulses establishes a plasma response in intrinsic germanium structures fabricated on a silicon substrate. This direct approach activates the plasmonic resonance of the Ge structures and enables their use as optical antennas up to the mid-infrared spectral range. The optical switching lasts for hundreds of picoseconds until charge recombination redshifts the plasma frequency. The full behavior of the structures is modeled by the electrodynamic response established by an electron-hole plasma in a regular array of antennas.

Memory CD8(+) T Cells Require Increased Concentrations of Acetate Induced by Stress for Optimal Function.

How systemic metabolic alterations during acute infections impact immune cell function remains poorly understood. We found that acetate accumulates in the serum within hours of systemic bacterial infections and that these increased acetate concentrations are required for optimal memory CD8(+) T cell function in vitro and in vivo. Mechanistically, upon uptake by memory CD8(+) T cells, stress levels of acetate expanded the cellular acetyl-coenzyme A pool via ATP citrate lyase and promoted acetylation of the enzyme GAPDH. This context-dependent post-translational modification enhanced GAPDH activity, catalyzing glycolysis and thus boosting rapid memory CD8(+) T cell responses. Accordingly, in a murine Listeria monocytogenes model, transfer of acetate-augmented memory CD8(+) T cells exerted superior immune control compared to control cells. Our results demonstrate that increased systemic acetate concentrations are functionally integrated by CD8(+) T cells and translate into increased glycolytic and functional capacity. The immune system thus directly relates systemic metabolism with immune alertness.

The Immune-Metabolic Basis of Effector Memory CD4+ T Cell Function under Hypoxic Conditions.

Effector memory (EM) CD4(+) T cells recirculate between normoxic blood and hypoxic tissues to screen for cognate Ag. How mitochondria of these cells, shuttling between normoxia and hypoxia, maintain bioenergetic efficiency and stably uphold antiapoptotic features is unknown. In this study, we found that human EM CD4(+) T cells had greater spare respiratory capacity (SRC) than did naive counterparts, which was immediately accessed under hypoxia. Consequently, hypoxic EM cells maintained ATP levels, survived and migrated better than did hypoxic naive cells, and hypoxia did not impair their capacity to produce IFN-γ. EM CD4(+) T cells also had more abundant cytosolic GAPDH and increased glycolytic reserve. In contrast to SRC, glycolytic reserve was not tapped under hypoxic conditions, and, under hypoxia, glucose metabolism contributed similarly to ATP production in naive and EM cells. However, both under normoxic and hypoxic conditions, glucose was critical for EM CD4(+) T cell survival. Mechanistically, in the absence of glycolysis, mitochondrial membrane potential (ΔΨm) of EM cells declined and intrinsic apoptosis was triggered. Restoring pyruvate levels, the end product of glycolysis, preserved ΔΨm and prevented apoptosis. Furthermore, reconstitution of reactive oxygen species (ROS), whose production depends on ΔΨm, also rescued viability, whereas scavenging mitochondrial ROS exacerbated apoptosis. Rapid access of SRC in hypoxia, linked with built-in, oxygen-resistant glycolytic reserve that functionally insulates ΔΨm and mitochondrial ROS production from oxygen tension changes, provides an immune-metabolic basis supporting survival, migration, and function of EM CD4(+) T cells in normoxic and hypoxic conditions.

Complement Regulates Nutrient Influx and Metabolic Reprogramming during Th1 Cell Responses.

Expansion and acquisition of Th1 cell effector function requires metabolic reprogramming; however, the signals instructing these adaptations remain poorly defined. Here we found that in activated human T cells, autocrine stimulation of the complement receptor CD46, and specifically its intracellular domain CYT-1, was required for induction of the amino acid (AA) transporter LAT1 and enhanced expression of the glucose transporter GLUT1. Furthermore, CD46 activation simultaneously drove expression of LAMTOR5, which mediated assembly of the AA-sensing Ragulator-Rag-mTORC1 complex and increased glycolysis and oxidative phosphorylation (OXPHOS), required for cytokine production. T cells from CD46-deficient patients, characterized by defective Th1 cell induction, failed to upregulate the molecular components of this metabolic program as well as glycolysis and OXPHOS, but IFN-γ production could be reinstated by retrovirus-mediated CD46-CYT-1 expression. These data establish a critical link between the complement system and immunometabolic adaptations driving human CD4(+) T cell effector function.

Nonlinear photoluminescence spectrum of single gold nanostructures.

We investigate the multiphoton photoluminescence characteristics of gold nanoantennas fabricated from single crystals and polycrystalline films. By exciting these nanostructures with ultrashort pulses tunable in the near-infrared range, we observe distinct features in the broadband photoluminescence spectrum. By comparing antennas of different crystallinity and shape, we demonstrate that the nanoscopic geometry of plasmonic devices determines the shape of the emission spectra. Our findings rule out the contribution of the gold band structure in shaping the photoluminescence.