Critical computational analysis illuminates the reductive-elimination mechanism that activates nitrogenase for N2 reduction
Simone Raugei, Lance C. Seefeldt, and Brian M. Hoffman
This report critically evaluates the mechanism by which nitrogenase cleaves the N≡N triple bond. It assesses the thermodynamic driving force provided by the accompanying, apparently “wasteful,” reductive elimination of an H2, and explains how the enzyme mechanistically couples exothermic H2 formation to endothermic triple-bond cleavage in a nearly thermoneutral equilibrium process, thereby preventing the “futile” generation of two H2 without N2 reduction. This evaluation rests on a critical assessment of the density functional theory flavors needed to properly treat nitrogenase, and a demonstration that to prevent spurious disruption of FeMo-co upon 4[e−/H+] accumulation, one must employ a nitrogenase structural model that includes all residues interacting directly with FeMo-co, either via specific H-bond interactions, nonspecific electrostatic interactions, or steric confinement. (See pp. E10521–E10530.)
Nonequilibrium associative retrieval of multiple stored self-assembly targets
Gili Bisker and Jeremy L. England
Major experimental research efforts have gone into investigating general principles governing self-assembly under nonequilibrium driving. However, in contrast to equilibrium scenarios, where the system tends to find local minima in the free-energy landscape, there is no equivalent theoretical framework for systems operating far from equilibrium. Inspired by many examples of nonequilibrium self-assembly in living systems, we set out to explore the added benefits achieved by nonequilibrium driving and identify distinctive collective phenomena that emerge in this regime. We demonstrate the interplay between the assembly speed, kinetic stability, and relative population of dynamical attractors, aiming to provide insights into nonequilibrium self-assembly processes and lay the foundations for understanding biomolecular cases as well as for designing examples. (See pp. E10531–E10538.)
Intestinal barrier dysfunction orchestrates the onset of inflammatory host–microbiome cross-talk in a human gut inflammation-on-a-chip
Woojung Shin and Hyun Jung Kim
Identification of the trigger of human intestinal inflammation can be a compelling clinical strategy for developing effective and target-specific antiinflammatory therapeutics. The pathomimetic “gut inflammation-on-a-chip” inspired by dextran sodium sulfate (DSS)-induced colitis models in mice enabled the independent uncoupling of complex inflammatory cross-talks and the combinatorial recoupling of individual contributing factors one at a time to identify the initiator of inflammatory responses. Our discovery suggests that an intact epithelial barrier is necessary to maintain the “homeostatic tolerance” in response to physiological host–gut microbiome cross-talks. We also expound an insight of probiotic therapy that the undamaged epithelial barrier is a prerequisite for eliciting the probiotic efficacy. Finally, the gut inflammation-on-a-chip verifies how microphysiological systems can be successfully implemented to dissect the mechanisms of gastrointestinal diseases. (See pp. E10539–E10547.)
Structural and mechanistic insights into the function of the unconventional class XIV myosin MyoA from Toxoplasma gondii
Cameron J. Powell, Raghavendran Ramaswamy, Anne Kelsen, David J. Hamelin, David M. Warshaw, Jürgen Bosch, John E. Burke, Gary E. Ward, and Martin J. Boulanger
Class XIV myosins are promising therapeutic targets for the treatment of apicomplexan disease because of their importance in parasite fitness and divergence from conventional human myosins. In this study, we report the crystal structure of a class XIV myosin, MyoA from Toxoplasma gondii. Structural analysis complemented with mutagenesis, biochemical, and functional data support a model whereby unique sequence elements in class XIV motors result in unique mechanisms of force production and chemomechanical coupling. Notably, many of these elements are located in known binding sites for allosteric inhibitors of myosin function, highlighting the potential for the design of class-specific myosin inhibitors as treatments for apicomplexan disease. With the established structural and biochemical platforms, we are poised to advance detailed functional studies and support inhibitor development. (See pp. E10548–E10555.)
Arrhythmia mutations in calmodulin cause conformational changes that affect interactions with the cardiac voltage-gated calcium channel
Kaiqian Wang, Christian Holt, Jocelyn Lu, Malene Brohus, Kamilla Taunsig Larsen, Michael Toft Overgaard, Reinhard Wimmer, and Filip Van Petegem
Calmodulin is a ubiquitous Ca2+-sensing protein that can bind to more than a 100 different targets. In doing so, it endows many of these with Ca2+-dependent modulation. As one of the most conserved proteins throughout evolutionary history, all calmodulin genes in vertebrates are identical. However, several disease-associated mutations have been uncovered in human calmodulin genes, all of which are linked to inherited cardiac arrhythmia syndromes. This report shows high-resolution glimpses into calmodulin disease mutations and their effect on binding the L-type voltage-gated calcium channel, a channel involved in the cardiac action potential that receives calcium-dependent feedback. Although each mutant is able to affect calcium-dependent inactivation, the structures show that they adopt different mechanisms. (See pp. E10556–E10565.)
Small-molecule CaVα1⋅CaVβ antagonist suppresses neuronal voltage-gated calcium-channel trafficking
Xingjuan Chen, Degang Liu, Donghui Zhou, Yubing Si, David Xu, Christopher W. Stamatkin, Mona K. Ghozayel, Matthew S. Ripsch, Alexander G. Obukhov, Fletcher A. White, and Samy O. Meroueh
Voltage-gated ion channels, such as CaV2.2, consist of pore-forming and auxiliary subunits that interact through protein–protein interactions. We develop a small-molecule antagonist of the protein–protein interaction between the calcium channel alpha pore-forming domain (CaVα) and beta subunits (CaVβ). The compound suppresses trafficking of CaV2.2 channels to the cell membrane and inhibits CaV2.2 activity by acting intracellularly. This allows peripheral access and eliminates the need of intrathecal administration. Indeed, in vivo systemic administration of the small molecule reduces neuropathic pain behavior in animal models. Our compounds serve as chemical tools to explore the CaVα⋅CaVβ interaction in vivo and as a starting point for the development of therapeutics for the treatment of a range of disorders associated with calcium channels. (See pp. E10566–E10575.)
SHOC2–MRAS–PP1 complex positively regulates RAF activity and contributes to Noonan syndrome pathogenesis
Lucy C. Young, Nicole Hartig, Isabel Boned del Río, Sibel Sari, Benjamin Ringham-Terry, Joshua R. Wainwright, Greg G. Jones, Frank McCormick, and Pablo Rodriguez-Viciana
We demonstrate a mechanism whereby germline mutations in MRAS, SHOC2, and PPP1CB contribute directly to Noonan syndrome by enhancing formation of a ternary complex, which specifically dephosphorylates an inhibitory site on RAF kinases, activating downstream signaling. SHOC2 is required for tumorigenic properties of tumor-derived cell lines with RAS mutations and has more recently been identified by others in a synthetic lethal screen as a gene essential for viability of RAS mutant but not RAS wild-type cells. A thorough analysis of this complex at the biochemical and structural level has demonstrated the remarkable ability of this complex to dictate specificity for RAF and suggests possible strategies to inhibit the complex as a way of targeting the RAS–ERK pathway. (See pp. E10576–E10585.)
Specificity landscapes unmask submaximal binding site preferences of transcription factors
Devesh Bhimsaria, José A. Rodríguez-Martínez, Junkun Pan, Daniel Roston, Elif Nihal Korkmaz, Qiang Cui, Parameswaran Ramanathan, and Aseem Z. Ansari
Several experimental platforms and computational methods have been developed to identify DNA binding sites of over 1,000 transcription factors. Often, high-affinity (maximal) binding sites are reported as consensus motifs. Differences between experimental platforms contribute to uncertainty in ascribing binding to submaximal sites. However, biological studies emphasize the importance of submaximal binding sites in shaping regulatory functions of transcription factors. To bridge this gap, we developed Differential Specificity and Energy Landscapes to unmask differences between experimental and computational methods as well as capture distinct submaximal binding site preferences of transcription factors. Our results suggest that subtle variation in protein structure can allosterically confer homolog-specific differences in binding to submaximal affinity sites. (See pp. E10586–E10595.)
Noncanonical role for the binding protein in substrate uptake by the MetNI methionine ATP Binding Cassette (ABC) transporter
Phong T. Nguyen, Jeffrey Y. Lai, Allen T. Lee, Jens T. Kaiser, and Douglas C. Rees
The high-affinity methionine importer MetNI belongs to the ATP Binding Cassette (ABC) family of transporters that carry out the ATP-dependent uptake of substrates into cells. As with other ABC importers, MetNI requires a soluble binding protein (MetQ) that in the canonical mechanistic model delivers substrates to the transporter. We made the unexpected observation that a MetQ variant with significantly impaired ligand-binding properties supports d-selenomethionine uptake at a higher rate than wild-type MetQ. A crystal structure of MetNIQ in the outward-facing conformation reveals access channels through the binding protein to the transmembrane translocation pathway. These studies support a noncanonical role for the binding protein in facilitating the uptake of certain substrates directly through the transporter–binding protein complex. (See pp. E10596–E10604.)
Hox5 genes direct elastin network formation during alveologenesis by regulating myofibroblast adhesion
Steven M. Hrycaj, Leilani Marty-Santos, Cristina Cebrian, Andrew J. Rasky, Catherine Ptaschinski, Nicholas W. Lukacs, and Deneen M. Wellik
Hox5 genes play critical roles in embryonic lung development, but mutants die at birth, preventing investigation of potential postnatal functions for these genes. Surprisingly, we show that the highest expression levels of Hox5 genes occur 1–2 weeks after birth. We created a conditional allele for Hoxa5 that allowed us to generate and study mutants for all three Hox5 genes during postnatal development. Hox5 mutants have poorly developed alveoli and expanded distal airspaces resulting from an abrogated elastin network. These defects arise from the inability of Hox5 mutant fibroblasts to adhere to the fibronectin matrix due to loss of integrins Itga5/b1. Thus, our data highlight redundant roles for all three Hox5 genes in regulating fibroblast adhesion and elastogenesis during alveologenesis. (See pp. E10605–E10614.)
Evolutionarily conserved Tbx5–Wnt2/2b pathway orchestrates cardiopulmonary development
Jeffrey D. Steimle, Scott A. Rankin, Christopher E. Slagle, Jenna Bekeny, Ariel B. Rydeen, Sunny Sun-Kin Chan, Junghun Kweon, Xinan H. Yang, Kohta Ikegami, Rangarajan D. Nadadur, Megan Rowton, Andrew D. Hoffmann, Sonja Lazarevic, William Thomas, Erin A. T. Boyle Anderson, Marko E. Horb, Luis Luna-Zurita, Robert K. Ho, Michael Kyba, Bjarke Jensen, Aaron M. Zorn, Frank L. Conlon, and Ivan P. Moskowitz
In the 20 years since the discovery of the genetic link between the transcription factor TBX5 and congenital heart defects, few direct targets of TBX5 in cardiac morphogenesis have been identified. In this work, we demonstrate that TBX5 directly regulates canonical Wnt ligands required for initiation of lung development. Lung endoderm forms a Hedgehog signaling source required for morphogenesis of both the lungs and the cardiac inflow septum. Our work expands the role of TBX5 to include a non–cell-autonomous component for atrial septation. We find the mesoderm–endoderm–mesoderm signaling loop initiated by TBX5 is evolutionarily conserved from amphibians to mammals. This work suggests that the evolutionary origin of lungs may have involved the recruitment of cardiac TBX5. (See pp. E10615–E10624.)
Epidemiology of the silent polio outbreak in Rahat, Israel, based on modeling of environmental surveillance data
Andrew F. Brouwer, Joseph N. S. Eisenberg, Connor D. Pomeroy, Lester M. Shulman, Musa Hindiyeh, Yossi Manor, Itamar Grotto, James S. Koopman, and Marisa C. Eisenberg
The 2013–2014 silent polio epidemic in Israel was a setback to global eradication efforts because Israel had previously been certified as polio-free by the World Health Organization. Fortunately, Israel has a robust environmental surveillance program that detected the epidemic and allowed rapid mobilization of a vaccine campaign before any cases of acute flaccid paralysis. This kind of silent (caseless) epidemic will be increasingly common as we approach global eradication, demonstrating the need for both enhanced environmental surveillance and an accompanying inference framework to translate environmental data into public health metrics. We incorporate environmental data into a population-level disease transmission model, generating insights into the epidemiology of the outbreak. This framework can be used to guide future interventions. (See pp. E10625–E10633.)
Species groups distributed across elevational gradients reveal convergent and continuous genetic adaptation to high elevations
Yan-Bo Sun, Ting-Ting Fu, Jie-Qiong Jin, Robert W. Murphy, David M. Hillis, Ya-Ping Zhang, and Jing Che
Organisms living in extreme environments are useful for studying the process of adaptation. We studied two distantly related groups that are distributed across a broad elevational gradient on and near the Qinghai-Tibetan Plateau and identified molecular adaptations to increasing elevations. We show that high-elevation adaptation (HEA) emerged soon after a split from low-elevation lineages, and adaptations continue to evolve in species that inhabit increasingly high elevations. Genes related to DNA repair and energy metabolism evolved rapidly, suggesting a crucial role of these genes in HEA. Moreover, we observed common patterns of HEA for similar functions between distantly related lineages, although these functional changes often involved different specific genes. (See pp. E10634–E10641.)
BRCA1 ensures genome integrity by eliminating estrogen-induced pathological topoisomerase II–DNA complexes
Hiroyuki Sasanuma, Masataka Tsuda, Suguru Morimoto, Liton Kumar Saha, Md Maminur Rahman, Yusuke Kiyooka, Haruna Fujiike, Andrew D. Cherniack, Junji Itou, Elsa Callen Moreu, Masakazu Toi, Shinichiro Nakada, Hisashi Tanaka, Ken Tsutsui, Shintaro Yamada, Andre Nussenzweig, and Shunichi Takeda
BRCA1 plays a key role in homology-directed repair (HDR) in S/G2-phase cells. It remains unclear why BRCA1 mutation carriers develop cancer predominantly in breast and ovarian tissues. We revealed that a physiological concentration (10 nM) of estrogens efficiently induce TOP2β-dependent DSBs in the absence of BRCA1 in breast cancer cells arrested in G1 phase. This genotoxicity was confirmed also in G0/G1-phase epithelial cells of mouse mammary glands. These findings indicated that BRCA1 contributes to DSB repair independent of HDR. Our data suggested that BRCA1 promotes the removal of TOP2 adducts from DSBs by the nucleolytic activity of MRE11 for subsequent DSB repair by nonhomologous end-joining. This function of BRCA1 may help explain the female-organ-specific carcinogenesis of BRCA1-mutation carriers. (See pp. E10642–E10651.)
RNA polymerase II CTD interactome with 3′ processing and termination factors in fission yeast and its impact on phosphate homeostasis
Ana M. Sanchez, Stewart Shuman, and Beate Schwer
The phosphorylation pattern of the Pol2 carboxy-terminal domain (CTD) Y1S2P3T4S5P6S7 repeats comprises an informational code coordinating transcription and RNA processing. We exploited fission yeast CTD phospho-site mutants and synthetic genetic arraying to illuminate opposing roles for Ser7 and Thr4 in transcription termination whereby: S7A elicits precocious termination via cleavage-polyadenylation factor (CPF) subunits and Rhn1; and T4A reduces termination and is lethal absent CPF subunits Ppn1 and Swd22. The findings that Y1F, S2A, and T4A are concordantly lethal with ppn1∆ and swd22∆ provide insights into a CTD vocabulary, implicating Tyr1-Ser2-Thr4 as a three-letter CTD word. This work underscores how the effects of mutating ostensibly inessential CTD marks are genetically buffered by other cellular factors that are functionally redundant to those marks. (See pp. E10652–E10661.)
Lipoteichoic acid anchor triggers Mincle to drive protective immunity against invasive group A Streptococcus infection
Takashi Imai, Takayuki Matsumura, Sabine Mayer-Lambertz, Christine A. Wells, Eri Ishikawa, Suzanne K. Butcher, Timothy C. Barnett, Mark J. Walker, Akihiro Imamura, Hideharu Ishida, Tadayoshi Ikebe, Tomofumi Miyamoto, Manabu Ato, Shouichi Ohga, Bernd Lepenies, Nina M. van Sorge, and Sho Yamasaki
Group A Streptococcus (GAS) causes invasive streptococcal infections in humans, resulting in high mortality. Thus, GAS is also known as “killer bacteria” or “flesh-eating bacteria.” The mechanism by which the immune system recognizes this potent pathogen remains elusive. In this study, we showed that the innate immune receptor Mincle (macrophage inducible C-type lectin) plays pivotal roles against invasive GAS infection through the recognition of monoglucosyldiacylglycerol (MGDG), a component of the lipoteichoic acid anchor. MGDG induced proinflammatory cytokines, ROS, and NO production in a Mincle-dependent manner. In an invasive GAS infection model, Mincle-deficient mice exhibited severe bacteremia and rapid lethality. These results indicate that Mincle plays a central role in protective immunity against acute GAS infection. (See pp. E10662–E10671.)
Identification and validation of a tumor-infiltrating Treg transcriptional signature conserved across species and tumor types
Angela M. Magnuson, Evgeny Kiner, Ayla Ergun, Jun Seok Park, Natasha Asinovski, Adriana Ortiz-Lopez, Aoife Kilcoyne, Elisa Paoluzzi-Tomada, Ralph Weissleder, Diane Mathis, and Christophe Benoist
FOXP3+ T regulatory cells (Tregs) dampen immune responses in many environments, particularly in tumors, where they contribute to cancer’s resistance to immunologic defenses. This very broad analysis of tumor-infiltrating Tregs has identified a set of genes that are preferentially expressed by these Tregs in different species, tumor models or cohorts, and types or stages of tumors. This striking commonality suggests that there are core mechanisms that tumors use to attract and mold Tregs, whose perturbation should unleash antitumor immunity. Experimental validation by genome editing provides a proof of concept for the relevance of these genes in TITRs. (See pp. E10672–E10681.)
Akt-mediated platelet apoptosis and its therapeutic implications in immune thrombocytopenia
Mengxing Chen, Rong Yan, Kangxi Zhou, Xiaodong Li, Yang Zhang, Chunliang Liu, Mengxiao Jiang, Honglei Ye, Xingjun Meng, Ningbo Pang, Lili Zhao, Jun Liu, Weiling Xiao, Renping Hu, Qingya Cui, Wei Zhong, Yunxiao Zhao, Mingqing Zhu, Anning Lin, Changgeng Ruan, and Kesheng Dai
Immune thrombocytopenia (ITP) patients with antiplatelet glycoprotein (GP) Ib-IX autoantibodies appear refractory to conventional treatments; however, the mechanism remains elusive. Here we show that the platelets undergo apoptosis in ITP patients with anti-GPIbα autoantibodies. We demonstrate that anti-GPIbα antibody binding activates Akt, which elicits platelet apoptosis through activation of phosphodiesterase (PDE3A) and PDE3A-mediated PKA inhibition. Phosphatidylserine (PS) exposure results in phagocytosis of anti-GPIbα antibody-bound platelets by macrophages in the liver. Notably, inhibition or genetic ablation of Akt or Akt-regulated apoptotic signaling or blockage of PS exposure rescues the platelets from clearance. Therefore, our findings reveal pathogenic mechanisms of ITP with anti-GPIbα autoantibodies and, more importantly, suggest therapeutic strategies for thrombocytopenia caused by autoantibodies or other pathogenic factors. (See pp. E10682–E10691.)
Mastocytosis-derived extracellular vesicles exhibit a mast cell signature, transfer KIT to stellate cells, and promote their activation
Do-Kyun Kim, Young-Eun Cho, Hirsh D. Komarow, Geethani Bandara, Byoung-Joon Song, Ana Olivera, and Dean D. Metcalfe
Pathological findings in systemic mastocytosis (SM) are generally attributed to an increase in the mast cell burden and associated production of mast cell mediators. We now describe that serum from patients with SM contains extracellular vesicles (EVs) with a mast cell signature and that their concentrations correlate with surrogate markers of disease. These SM-EVs have the ability to alter hepatic stellate cell function including promoting a fibrotic phenotype, partly through the introduction of activated KIT. Tyrosine kinase inhibitors used to treat SM reversed these effects. These observations provide additional insight into how mast cells influence other organ systems in mastocytosis and suggest that targeting the release and/or effects of EVs might be of value in conjunction with existing therapeutic approaches. (See pp. E10692–E10701.)
Isolation and characterization of NY-ESO-1–specific T cell receptors restricted on various MHC molecules
Michael T. Bethune, Xiao-Hua Li, Jiaji Yu, Jami McLaughlin, Donghui Cheng, Colleen Mathis, Blanca Homet Moreno, Katherine Woods, Ashley J. Knights, Angel Garcia-Diaz, Stephanie Wong, Siwen Hu-Lieskovan, Cristina Puig-Saus, Jonathan Cebon, Antoni Ribas, Lili Yang, Owen N. Witte, and David Baltimore
T immune cells can be engineered to express tumor-specific T cell receptor (TCR) genes and thereby kill cancer cells. This approach—termed TCR gene therapy—is effective but can cause serious adverse events if the target is also expressed in healthy, noncancerous tissue. NY-ESO-1 is a tumor-specific antigen that has been targeted successfully and safely through TCR gene therapies for melanoma, synovial sarcoma, and myeloma. However, trials to date have focused exclusively on a single NY-ESO-1–derived epitope presented on HLA-A*02:01, limiting application to patients expressing that allele. In this work, we isolate TCRs that collectively recognize multiple NY-ESO-1–derived epitopes presented by multiple MHC alleles. We thereby outline a general approach for expanding targeted immunotherapies to more diverse MHC haplotypes. (See pp. E10702–E10711.)
Redox, amino acid, and fatty acid metabolism intersect with bacterial virulence in the gut
Reed Pifer, Regan M. Russell, Aman Kumar, Meredith M. Curtis, and Vanessa Sperandio
Enteric pathogens have to gauge the intestinal environment and adapt their metabolism and virulence strategies to establish themselves within the host. Here we show using a high-throughput screen that several metabolic pathways intertwine with virulence gene expression in enterohemorrhagic Escherichia coli. This screen identified transcriptional regulators that respond to fluctuations in amino and fatty acids as playing an important role in virulence gene expression both in vitro and during mammalian infection. Our study has fundamental implications for how differential gut metabolite compositions may affect disease outcome and susceptibility to pathogens. (See pp. E10712–E10719.)
Median nerve stimulation induces analgesia via orexin-initiated endocannabinoid disinhibition in the periaqueductal gray
Yi-Hung Chen, Hsin-Jung Lee, Ming Tatt Lee, Ya-Ting Wu, Yen-Hsien Lee, Ling-Ling Hwang, Ming-Shiu Hung, Andreas Zimmer, Ken Mackie, and Lih-Chu Chiou
Pain remains an unmet medical need due to the ineffectiveness or significant side effects of current pain therapies. Peripheral neurostimulation has been used to relieve pain for decades, but its mechanism(s) remain unsettled. Here, we established an animal model of median nerve stimulation by electrically stimulating the PC6 (Neiguan) acupoint of mice (MNS-PC6). We found MNS-PC6 can release an endogenous neuropeptide (orexin) from the hypothalamus to inhibit pain responses in mice through an endocannabinoid (an endogenous lipid functioning like chemicals from cannabis) that reduces the inhibitory (GABAergic) control in a midbrain pain-control region (the periaqueductal gray). Importantly, MNS-PC6–induced pain relief is endogenous opioid independent. Thus, MNS-PC6 may provide an alternative strategy for pain management in opioid-tolerant patients. (See pp. E10720–E10729.)
Dopamine D2 receptor-mediated circuit from the central amygdala to the bed nucleus of the stria terminalis regulates impulsive behavior
Bokyeong Kim, Sehyoun Yoon, Ryuichi Nakajima, Hyo Jin Lee, Hee Jeong Lim, Yeon-Kyung Lee, June-Seek Choi, Bong-June Yoon, George J. Augustine, and Ja-Hyun Baik
Impulsivity is a tendency to act with little or no forethought or consideration of the consequences and is a major component of various psychiatric disorders. However, little is known about the brain circuits that regulate reward-related impulsivity. Our genetic manipulations reveal that dopamine D2 receptors in the central nucleus of the amygdala have a crucial role in reward-related impulsive behavior. By using optogenetics to control the neurons that possess these receptors, we have identified elements of a neural circuit that contributes to regulating impulsivity. This information should enable approaches to managing impulsivity associated with neuropsychiatric disorders such as attention-deficit/hyperactivity disorder, bipolar disorder, and addiction-related disorders. (See pp. E10730–E10739.)
Activation of orexin system facilitates anesthesia emergence and pain control
Wei Zhou, Kevin Cheung, Steven Kyu, Lynn Wang, Zhonghui Guan, Philip A. Kurien, Philip E. Bickler, and Lily Y. Jan
Although millions of procedures are performed under general anesthesia every year, we do not fully understand the mechanisms underlying anesthesia. Orexin neurons in the hypothalamus form one of the centers in the central nervous system involved in sleep–wake control. The orexin system has also been implicated in anesthesia. In this study, we specifically activated orexin neurons via a designer receptor that is exclusively activated by a designer drug, to test how activation of orexin neurons affects anesthesia recovery. We found that orexin neuronal activation can speed up wakeup and improve pain control as well. Our study suggests that the orexin system has the potential as a target for drug discovery to facilitate postprocedural recovery. (See pp. E10740–E10747.)
Coiled-coil structure-dependent interactions between polyQ proteins and Foxo lead to dendrite pathology and behavioral defects
Min Jee Kwon, Myeong Hoon Han, Joshua A. Bagley, Do Young Hyeon, Byung Su Ko, Yun Mi Lee, In Jun Cha, Seung Yeol Kim, Dong Young Kim, Ho Min Kim, Daehee Hwang, Sung Bae Lee, and Yuh Nung Jan
It remains unclear how the structural properties of polyglutamine (polyQ) proteins, which underlie several neurodegenerative disorders, including Huntington’s disease and spinocerebellar ataxias (SCAs), translate into the toxicity of these proteins. Here, we demonstrate that coiled-coil structures in expanded polyQ regions of SCA type 3 (SCA3) proteins cause dendrite defects in Drosophila neurons, as well as behavioral abnormalities. Moreover, interactions of SCA3 with Foxo mediated by coiled-coil domains of these two proteins resulted in functional impairment of this transcription factor, whereas its overexpression significantly rescued the SCA3-induced defects. Our study expanded the current understanding of neuronal pathology mediated by polyQ proteins via the coiled-coil–mediated interactions. These results may have important implications in therapeutic strategies for polyQ protein-related diseases. (See pp. E10748–E10757.)
Metabolic regulation of female puberty via hypothalamic AMPK–kisspeptin signaling
Juan Roa, Alexia Barroso, Francisco Ruiz-Pino, Maria Jesus Vázquez, Patricia Seoane-Collazo, Noelia Martínez-Sanchez, David García-Galiano, Tuncay Ilhan, Rafael Pineda, Silvia León, Maria Manfredi-Lozano, Violeta Heras, Matti Poutanen, Juan M. Castellano, Francisco Gaytan, Carlos Diéguez, Leonor Pinilla, Miguel López, and Manuel Tena-Sempere
The age of puberty in humans is changing via unknown mechanisms, although metabolic alterations in childhood are blamed as a major contributing factor. Perturbations in pubertal timing are posed with increased risk of later cardiometabolic diseases and reduced life expectancy, urging a better understanding of the molecular basis for these phenomena. We describe a mechanism whereby the main cellular energy sensor, AMPK, operates in a population of hypothalamic neurons, named Kiss1, which produce the puberty-activating signal, kisspeptin, to metabolically control puberty onset. This neuroendocrine circuit provides a molecular link between conditions of negative energy balance and delayed pubertal timing, via a repressive AMPK–Kiss1 pathway, which may become a druggable target in conditions of disordered puberty, especially of metabolic origin. (See pp. E10758–E10767.)
JAZ repressors of metabolic defense promote growth and reproductive fitness in Arabidopsis
Qiang Guo, Yuki Yoshida, Ian T. Major, Kun Wang, Koichi Sugimoto, George Kapali, Nathan E. Havko, Christoph Benning, and Gregg A. Howe
The plant hormone jasmonate promotes resistance to plant-eating organisms, ranging from pathogenic microbes to mammals. Jasmonate reprograms metabolism to fuel the production of diverse defense compounds and simultaneously inhibits plant growth. Understanding how growth is influenced across a range of defense levels remains unclear, but has important implications for optimizing crop productivity. Using a genetic approach to “tune” the jasmonate response, we assessed the physiological consequences of discrete levels of defense throughout the plant life cycle. Overactivation of jasmonate response led to carbon starvation, near loss of seed production and, under extreme conditions, lethality. Our findings explain the emergence of diverse strategies to keep jasmonate responses at bay and provide new insights into metabolic processes that underlie growth–defense trade-offs. (See pp. E10768–E10777.)
ATP compartmentation in plastids and cytosol of Arabidopsis thaliana revealed by fluorescent protein sensing
Chia Pao Voon, Xiaoqian Guan, Yuzhe Sun, Abira Sahu, May Ngor Chan, Per Gardeström, Stephan Wagner, Philippe Fuchs, Thomas Nietzel, Wayne K. Versaw, Markus Schwarzländer, and Boon Leong Lim
By studying in vivo changes of ATP levels in the plastids and cytosol of Arabidopsis thaliana using a FRET-based ATP sensor, we show that the plastidic ATP concentrations in cotyledon, hypocotyl, and root of 10-day-old seedlings are significantly lower than the cytosolic concentrations. We show that chloroplasts consume ATP rapidly and the import of ATP into mature chloroplasts is impeded by the low density of NTT transporter. Hence, unlike in diatoms, where ATP is imported into chloroplasts to support the Calvin–Benson–Bassham (CBB) cycle, mature chloroplasts of Arabidopsis do not balance the ATP:NADPH ratio by importing ATP from the cytosol. Rather, chloroplasts can export surplus reducing equivalents, which can be used by the mitochondria to supply ATP to the cytosol. (See pp. E10778–E10787.)
More than $1 billion needed annually to secure Africa’s protected areas with lions
Peter A. Lindsey, Jennifer R. B. Miller, Lisanne S. Petracca, Lauren Coad, Amy J. Dickman, Kathleen H. Fitzgerald, Michael V. Flyman, Paul J. Funston, Philipp Henschel, Samuel Kasiki, Kathryn Knights, Andrew J. Loveridge, David W. Macdonald, Roseline L. Mandisodza-Chikerema, Sean Nazerali, Andrew J. Plumptre, Riko Stevens, Hugo W. Van Zyl, and Luke T. B. Hunter
Protected areas (PAs) are the cornerstone of conservation yet face funding inadequacies that undermine their effectiveness. Using the conservation needs of lions as a proxy for those of wildlife more generally, we compiled a dataset of funding in Africa’s PAs with lions and estimated a minimum target for conserving the species and managing PAs effectively. PAs with lions require $1.2 to $2.4 billion or $1,000 to $2,000/km2 annually, yet receive just $381 million or $200/km2 (median) annually. Nearly all PAs with lions are inadequately funded; deficits total $0.9 to $2.1 billion. Governments and donors must urgently and significantly invest in PAs to prevent further declines of lions and other wildlife and to capture the economic, social, and environmental benefits that healthy PAs can confer. (See pp. E10788–E10796.)
Multiscale effects of heating and cooling on genes and gene networks
Daniel A. Charlebois, Kevin Hauser, Sylvia Marshall, and Gábor Balázsi
Fluctuating environments such as changes in ambient temperature represent a fundamental challenge to life. Cells must protect gene networks that protect them from such stresses, making it difficult to understand how temperature affects gene network function in general. Here, we focus on single genes and small synthetic network modules to reveal four key effects of nonoptimal temperatures at different biological scales: (i) a cell fate choice between arrest and resistance, (ii) slower growth rates, (iii) Arrhenius reaction rates, and (iv) protein structure changes. We develop a multiscale computational modeling framework that captures and predicts all of these effects. These findings promote our understanding of how temperature affects living systems and enables more robust cellular engineering for real-world applications. (See pp. E10797–E10806.)