Regulatory cascade and biological activity of Beauveria bassiana oosporein that limits bacterial growth after host death
Yanhua Fan, Xi Liu, Nemat O. Keyhani, Guirong Tang, Yan Pei, Wenwen Zhang, and Sheng Tong
Since the discovery of oosporein more than 70 years ago, there have been conflicting reports on its potential antimicrobial and insecticidal activities. Our results indicate that oosporein is unlikely to function as an insect toxin or to be involved in early to mid-infection processes, including penetration and immune evasion. Instead, oosporein most likely functions after death of the host to thwart bacterial competition on a host cadaver, allowing the fungus to maximally use host nutrients and complete its life cycle. Our data also reveal that oosporein production is regulated by a cascade of transcription factors, with BbSmr1 acting as an upstream negative regulator, targeting the expression of OpS3, which in turn acts as a positive regulator of the oosporein biosynthetic gene cluster. (See pp. E1578–E1586.)
Vulnerability of primitive human placental trophoblast to Zika virus
Megan A. Sheridan, Dinar Yunusov, Velmurugan Balaraman, Andrei P. Alexenko, Shinichiro Yabe, Sergio Verjovski-Almeida, Danny J. Schust, Alexander W. Franz, Yoel Sadovsky, Toshihiko Ezashi, and R. Michael Roberts
We have tested the hypothesis that the placenta of early pregnancy might be more easily breached by the Zika virus (ZIKV) than the relatively resistant outer cells of the mature placenta. Colonies of placental lineage cells derived from embryonic stem cells, which are probably analogous to the primitive placenta at implantation, were lysed more rapidly by an African strain of ZIKV, considered relatively benign, than by an Asian strain linked to fetal brain abnormalities. We conclude that the human fetus may be most vulnerable to ZIKV very early in pregnancy and that the African strain may threaten a pregnancy more strongly than previously believed. (See pp. E1587–E1596.)
DYNC1H1 mutations associated with neurological diseases compromise processivity of dynein–dynactin–cargo adaptor complexes
Ha Thi Hoang, Max A. Schlager, Andrew P. Carter, and Simon L. Bullock
Mutations in microtubule motors and their cofactors are associated with several neurological diseases in humans. How disease-associated mutations affect motor function, and consequently have neuropathological effects, is largely unknown. We take advantage of recent advances in the ability to activate transport of the human dynein complex in vitro to functionally characterize 17 disease-associated mutations in the gene encoding its heavy chain. The vast majority of mutations do not affect the ability of dynein to bind a cargo adaptor but do compromise long-range motion of the motor complex. Our findings suggest that defective motility of cargo–motor complexes contributes to dynein-associated neurological phenotypes and highlight several regions of the enigmatic motor that orchestrate its ability to walk over long distances. (See pp. E1597–E1606.)
Investigations of human myosin VI targeting using optogenetically controlled cargo loading
Alexander R. French, Tobin R. Sosnick, and Ronald S. Rock
Myosins are a broad class of motor proteins that generate force on actin filaments and fulfill contractile, transport, and anchoring roles. Myosin VI, the only myosin to walk toward the pointed end of actin filaments, is implicated in cancer metastasis and deafness. Intriguingly, myosin VI may play both transport and anchoring roles, depending on where it is activated in the cell. Here we develop an optogenetic tool for studying myosin VI activation with high spatial and temporal resolution. Our approach photoactivates unmodified myosin VI through its native cargo pathway, enabling investigation of motor function and activation partners with minimal perturbation. This approach allows us to detect how and where myosin VI integrates multiple protein and second messenger signals to activate. (See pp. E1607–E1616.)
Modeling the two-way feedback between contractility and matrix realignment reveals a nonlinear mode of cancer cell invasion
Hossein Ahmadzadeh, Marie R. Webster, Reeti Behera, Angela M. Jimenez Valencia, Denis Wirtz, Ashani T. Weeraratna, and Vivek B. Shenoy
The mechanical cross talk between intracellular and extracellular forces can promote the invasive potential of tumor cells in tumors. Using a quantitative model, we elucidate the two-way feedback loop between stress-dependent cell contractility and matrix fiber realignment and strain stiffening, which enables the cells to polarize and enhance their contractility to break free from the tumor and invade into the matrix. Our model predicts that intermediate matrix stiffness is optimal for invasion, and we find a positive correlation between cell elongation and alignment of fibers in the matrix. Importantly, our model can be used to explain how morphological and structural changes in the tumor microenvironment, such as elevated rigidity and fiber alignment prior to cell invasion, are prognostic of the malignant phenotype. (See pp. E1617–E1626.)
Evidence for the principle of minimal frustration in the evolution of protein folding landscapes
Franco O. Tzul, Daniel Vasilchuk, and George I. Makhatadze
A detailed understanding of how different sequences that fold into the same structure affect the folding energy landscape presents one of the current challenges of the protein folding field. The principle of minimal frustration suggests that naturally evolved proteins with the same structure should have similar folding rates and that modulation of thermodynamic stability should occur via unfolding rates. We experimentally tested this hypothesis using 15 different thioredoxins, with sequences either obtained from the extant organisms or resurrected using ancestral sequence reconstruction. We show that all of these proteins fold with similar rates and that their dramatic differences in stability are because of the differences in the unfolding rates. (See pp. E1627–E1632.)
Improved regulatory element prediction based on tissue-specific local epigenomic signatures
Yupeng He, David U. Gorkin, Diane E. Dickel, Joseph R. Nery, Rosa G. Castanon, Ah Young Lee, Yin Shen, Axel Visel, Len A. Pennacchio, Bing Ren, and Joseph R. Ecker
In mammals, when and where a gene is transcribed are primarily regulated by the activity of regulatory DNA elements, or enhancers. Genetic mutation disrupting enhancer function is emerging as one of the major causes of human diseases. However, our knowledge remains limited about the location and activity of enhancers in the numerous and distinct cell types and tissues. Here, we develop a computational approach, regulatory element prediction based on tissue-specific local epigenetic marks (REPTILE), to precisely locate enhancers based on genome-wide DNA methylation and histone modification profiling. We systematically tested REPTILE on a variety of human and mouse cell types and tissues. Compared with existing methods, we found that enhancer predictions from REPTILE are more likely to be active in vivo and the predicted locations are more accurate. (See pp. E1633–E1640.)
Intrinsically disordered proteins drive enamel formation via an evolutionarily conserved self-assembly motif
Tomas Wald, Frantisek Spoutil, Adriana Osickova, Michaela Prochazkova, Oldrich Benada, Petr Kasparek, Ladislav Bumba, Ophir D. Klein, Radislav Sedlacek, Peter Sebo, Jan Prochazka, and Radim Osicka
Formation of the hardest mineralized tissue in vertebrates, tooth enamel, relies on a unique set of enamel matrix proteins (EMPs). These EMPs assemble into a 3D extracellular organic matrix that directs the deposition of calcium and phosphate ions into hydroxyapatite crystallites. However, the molecular basis of EMP assembly into the organic matrix remains poorly understood. This study shows that self-assembly of the key EMPs, ameloblastin and amelogenin, involves a short linear amino acid motif that is evolutionarily conserved from the first tetrapods to man. Functionality of this motif in ameloblastin is shown to be essential for organization of the enamel organic matrix and for proper organization of hydroxyapatite crystallites into the compact bundles that determine the structure and mechanical resistance of enamel. (See pp. E1641–E1650.)
The ETS-5 transcription factor regulates activity states in Caenorhabditis elegans by controlling satiety
Vaida Juozaityte, David Pladevall-Morera, Agnieszka Podolska, Steffen Nørgaard, Brent Neumann, and Roger Pocock
Animals constantly monitor their internal energy levels and modify their eating and foraging behavior as required. Our work defines a role for the ETS-5 transcription factor in the control of body fat levels and thereby the activity of animals. We have defined the responses controlled by ETS-5 at the genetic, cellular, and organismal levels and identified how ETS-5 interacts with known pathways that regulate food-regulated behavioral states. These findings provide insight into how fat levels are regulated and how satiety controls organismal activity. (See pp. E1651–E1658.)
NFκB–Pim-1–Eomesodermin axis is critical for maintaining CD8 T-cell memory quality
Karin M. Knudson, Curtis J. Pritzl, Vikas Saxena, Amnon Altman, Mark A. Daniels, and Emma Teixeiro
Mice and humans whose T cells are deficient in NFκB signaling lack memory T cells, but the mechanism behind this is unclear. We found that NFκB signaling is required during the resolution phase of the immune response to maintain long-term CD8 memory. NFκB signaling is necessary for preserving expression of Eomesodermin and prosurvival Bcl-2 in memory T cells, in a cell-intrinsic process where T-cell receptor (TCR) signals and Pim-1 kinase are involved. Our study defines an unexpected role of NFκB and Pim-1 signaling in the maintenance of T-cell memory quality. Furthermore, it identifies targets and specific times of intervention where protective T-cell memory could be reinforced in vaccines and cancer immunotherapies by manipulation of the NFκB–Pim-1–Eomesodermin axis. (See pp. E1659–E1667.)
Senescent cells expose and secrete an oxidized form of membrane-bound vimentin as revealed by a natural polyreactive antibody
David Frescas, Christelle M. Roux, Semra Aygun-Sunar, Anatoli S. Gleiberman, Peter Krasnov, Oleg V. Kurnasov, Evguenia Strom, Lauren P. Virtuoso, Michelle Wrobel, Andrei L. Osterman, Marina P. Antoch, Vadim Mett, Olga B. Chernova, and Andrei V. Gudkov
Understanding the mechanisms underlying the development of senescence and the consequences related to the accumulation of senescent cells is a major focus of ongoing research. Our report shows that senescent cells express a form of oxidized vimentin on their cell surface and that oxidized vimentin is secreted into the blood of senescence-prone senescence-accelerated mouse prone 8 mice. Given the growing evidence that oxidized proteins are involved in the development of human diseases, the detection and monitoring of secreted proteins like malondialdehyde-modified vimentin is certain to become a vital and noninvasive biomarker for studying senescence and monitoring age-related illnesses. (See pp. E1668–E1677.)
Functional genomics analysis of vitamin D effects on CD4+ T cells in vivo in experimental autoimmune encephalomyelitis
Manuel Zeitelhofer, Milena Z. Adzemovic, David Gomez-Cabrero, Petra Bergman, Sonja Hochmeister, Marie N'diaye, Atul Paulson, Sabrina Ruhrmann, Malin Almgren, Jesper N. Tegnér, Tomas J. Ekström, André Ortlieb Guerreiro-Cacais, and Maja Jagodic
Vitamin D has been suggested to be associated with beneficial immunomodulation in autoimmune diseases. We demonstrate that the protective effect of vitamin D in an animal model of multiple sclerosis (MS) is linked to multiple signaling and metabolic pathways critical for T-cell activation and differentiation into pathogenic T helper (Th) 1 and Th17 subsets in vivo. This effect is mediated by epigenetic mechanisms as reflected by genome-wide reduction of DNA methylation and upregulation of microRNAs, with concomitant downregulation of their protein-coding target genes. Our data support the role of vitamin D in modulating risk for human disease, because orthologues of nearly 50% of MS candidate risk genes changed their expression in vivo in CD4+ T cells upon vitamin D supplementation. (See pp. E1678–E1687.)
AMH/MIS as a contraceptive that protects the ovarian reserve during chemotherapy
Motohiro Kano, Amanda E. Sosulski, LiHua Zhang, Hatice D. Saatcioglu, Dan Wang, Nicholas Nagykery, Mary E. Sabatini, Guangping Gao, Patricia K. Donahoe, and David Pépin
All current reversible hormonal contraceptives rely on modulating gonadotropins or sex steroids by acting on the hypothalamic–pituitary–gonadal axis. Primordial follicle activation, the first step of folliculogenesis, is independent of gonadotropins or steroids. In this study we show that Müllerian inhibiting substance (MIS) can completely block primordial follicle activation, representing a unique mechanism of contraception that spares the pool of quiescent primordial follicles (ovarian reserve). Chemotherapy is thought to cause the over-recruitment of primordial follicles. Here we show that treatment with MIS during cycles of carboplatin, doxorubicin, or cyclophosphamide can significantly protect the ovarian reserve in mice. Thus, MIS may provide a paradigm of a reversible contraceptive that could mitigate damage to the ovarian reserve associated with gonadotoxic chemotherapeutics. (See pp. E1688–E1697.)
Galectin-3 directs antimicrobial guanylate binding proteins to vacuoles furnished with bacterial secretion systems
Eric M. Feeley, Danielle M. Pilla-Moffett, Erin E. Zwack, Anthony S. Piro, Ryan Finethy, Joseph P. Kolb, Jennifer Martinez, Igor E. Brodsky, and Jörn Coers
To combat infections with bacterial pathogens that reside and replicate inside specialized intracellular vacuoles, the innate immune system must be able to distinguish these pathogen-containing vacuoles (PVs) from endogenous vesicles. Here, we demonstrate that the host can detect the presence of bacterial secretion systems as hallmarks of PVs. The insertion of bacterial protein secretion pores destabilizes vesicular membranes, allowing host carbohydrate-binding proteins to access the sugar-decorated luminal side of vacuolar membranes. We show that a specific carbohydrate-binding protein recruits members of a family of antimicrobial GTPases to PVs. Our findings reveal how host cells deliver antimicrobial defenses specifically to intracellular vesicles occupied by pathogens. (See pp. E1698–E1706.)
AlphaB-crystallin regulates remyelination after peripheral nerve injury
Erin-Mai F. Lim, Stan T. Nakanishi, Vahid Hoghooghi, Shane E. A. Eaton, Alexandra L. Palmer, Ariana Frederick, Jo A. Stratton, Morgan G. Stykel, Patrick J. Whelan, Douglas W. Zochodne, Jeffrey Biernaskie, and Shalina S. Ousman
Regeneration and full behavioral recovery after injury to human peripheral nerves are often incomplete. To identify factors that could improve this situation, we focused on alphaB-crystallin (αBC), a small heat shock protein that has been associated with survival and differentiation of glial cells as well as neuroprotection in the central nervous system. We report that αBC, which is expressed by both peripheral axons and Schwann cells, is important for remyelination of damaged, peripheral axons in mice. Its absence resulted in thinner myelin sheaths and fewer myelinating Schwann cells. As a consequence, nerve conduction and sensory and motor behaviors were negatively impacted. Our work, therefore, suggests that administration of αBC can improve the regenerative capacity of the peripheral nervous system. (See pp. E1707–E1716.)
Ca2+-binding protein 2 inhibits Ca2+-channel inactivation in mouse inner hair cells
Maria Magdalena Picher, Anna Gehrt, Sandra Meese, Aleksandra Ivanovic, Friederike Predoehl, SangYong Jung, Isabelle Schrauwen, Alberto Giulio Dragonetti, Roberto Colombo, Guy Van Camp, Nicola Strenzke, and Tobias Moser
Ca2+ channels mediate excitation-secretion coupling and show little inactivation at sensory ribbon synapses, enabling reliable synaptic information transfer during sustained stimulation. Studies of Ca2+-channel complexes in HEK293 cells indicated that Ca2+-binding proteins (CaBPs) antagonize their calmodulin-dependent inactivation. Although human mutations affecting CABP2 were shown to cause hearing impairment, the role of CaBP2 in auditory function and the precise disease mechanism remained enigmatic. Here, we disrupted CaBP2 in mice and showed that CaBP2 is required for sound encoding at inner hair cell synapses, likely by suppressing Ca2+-channel inactivation. We propose that the number of activatable Ca2+ channels at the active zone is reduced when CaBP2 is lacking, as is likely the case with the newly described human CABP2 mutation. (See pp. E1717–E1726.)
Differential modulation of global and local neural oscillations in REM sleep by homeostatic sleep regulation
Bowon Kim, Bernat Kocsis, Eunjin Hwang, Youngsoo Kim, Robert E. Strecker, Robert W. McCarley, and Jee Hyun Choi
This study demonstrates that slow and fast cortical oscillations undergo different adaptations to homeostatic challenge of chronic sleep deprivation, which may benefit different functions of sleep. When mice sleep only 6 h/d for 5 d, rapid eye movement (REM) sleep settles on a persistently elevated level, even though sleep debt continues to accumulate. Using high-density EEG, we found that different forms of slow oscillations follow this general pattern, whereas all high-frequency oscillations showed progressive daily increases. Slow and fast oscillations play distinct roles in coordination of brain cell activity on different scales, and thus our results help to reconcile two seemingly opposite functions of sleep in synaptic homeostasis and sleep-dependent memory consolidation. (See pp. E1727–E1736.)
Hemoglobin phase of oxygenation and deoxygenation in early brain development measured using fNIRS
Hama Watanabe, Yoshihiko Shitara, Yoshinori Aoki, Takanobu Inoue, Shinya Tsuchida, Naoto Takahashi, and Gentaro Taga
We propose time-averaged hemoglobin phase of oxygenation and deoxygenation (hPod) measured using functional near-infrared spectroscopy as a method for the detection of the developmental status of the hemodynamic and metabolic processes of the brain. hPod values exhibit initially rapid changes from in-phase to antiphase patterns in neonates and infants. These changes become more gradual in later development. We also describe the impact of early preterm birth on brain development. (See pp. E1737–E1744.)
Phosphorylation of αB-crystallin supports reactive astrogliosis in demyelination
Hedwich F. Kuipers, Jane Yoon, Jack van Horssen, May H. Han, Paul L. Bollyky, Theo D. Palmer, and Lawrence Steinman
αB-crystallin (CRYAB) is a protein involved in the protection of cells from stress and cell death, and is very highly produced in brain lesions in multiple sclerosis (MS). Astrocytes are the main non-neuronal cell type in the central nervous system. Although they are involved in many processes in health and injury, their precise role in MS is poorly understood. Using a mouse model of MS, as well as brain tissue from MS patients, we found that CRYAB enables astrocytes to become activated and that these activated astrocytes play a pathogenic role in MS, worsening injury. Furthermore, we found that a specific stress-induced modification of CRYAB supports its role in astrocyte activation and affects the intracellular signaling pathways it regulates. (See pp. E1745–E1754.)