共查询到20条相似文献,搜索用时 46 毫秒
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Maria Bernabeu Samuel A. Danziger Marion Avril Marina Vaz Prasad H. Babar Andrew J. Brazier Thurston Herricks Jennifer N. Maki Ligia Pereira Anjali Mascarenhas Edwin Gomes Laura Chery John D. Aitchison Pradipsinh K. Rathod Joseph D. Smith 《Proceedings of the National Academy of Sciences of the United States of America》2016,113(23):E3270-E3279
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Sarit Avrani Debbie Lindell 《Proceedings of the National Academy of Sciences of the United States of America》2015,112(17):E2191-E2200
Prochlorococcus is an abundant marine cyanobacterium that grows rapidly in the environment and contributes significantly to global primary production. This cyanobacterium coexists with many cyanophages in the oceans, likely aided by resistance to numerous co-occurring phages. Spontaneous resistance occurs frequently in Prochlorococcus and is often accompanied by a pleiotropic fitness cost manifested as either a reduced growth rate or enhanced infection by other phages. Here, we assessed the fate of a number of phage-resistant Prochlorococcus strains, focusing on those with a high fitness cost. We found that phage-resistant strains continued evolving toward an improved growth rate and a narrower resistance range, resulting in lineages with phenotypes intermediate between those of ancestral susceptible wild-type and initial resistant substrains. Changes in growth rate and resistance range often occurred in independent events, leading to a decoupling of the selection pressures acting on these phenotypes. These changes were largely the result of additional, compensatory mutations in noncore genes located in genomic islands, although genetic reversions were also observed. Additionally, a mutator strain was identified. The similarity of the evolutionary pathway followed by multiple independent resistant cultures and clones suggests they undergo a predictable evolutionary pathway. This process serves to increase both genetic diversity and infection permutations in Prochlorococcus populations, further augmenting the complexity of the interaction network between Prochlorococcus and its phages in nature. Last, our findings provide an explanation for the apparent paradox of a multitude of resistant Prochlorococcus cells in nature that are growing close to their maximal intrinsic growth rates.Large bacterial populations are present in the oceans, playing important roles in primary production and the biogeochemical cycling of matter. These bacterial communities are highly diverse (1–4) yet form stable and reproducible bacterial assemblages under similar environmental conditions (5–7).These bacteria are present together with high abundances of viruses (phages) that have the potential to infect and kill them (8–11). Although studied only rarely in marine organisms (12–16), this coexistence is likely to be the result of millions of years of coevolution between these antagonistic interacting partners, as has been well documented for other systems (17–20). From the perspective of the bacteria, survival entails the selection of cells that are resistant to infection, preventing viral production and enabling the continuation of the cell lineage. Resistance mechanisms include passively acquired spontaneous mutations in cell surface molecules that prevent phage entry into the cell and other mechanisms that actively terminate phage infection intracellularly, such as restriction–modification systems and acquired resistance by CRISPR-Cas systems (21, 22). Mutations in the phage can also occur that circumvent these host defenses and enable the phage to infect the recently emerged resistant bacterium (23).Acquisition of resistance by bacteria is often associated with a fitness cost. This cost is frequently, but not always, manifested as a reduction in growth rate (24–27). Recently, an additional type of cost of resistance was identified, that of enhanced infection whereby resistance to one phage leads to greater susceptibility to other phages (14, 15, 28).Over the years, a number of models have been developed to explain coexistence in terms of the above coevolutionary processes and their costs (16, 29–32). In the arms race model, repeated cycles of host mutation and virus countermutation occur, leading to increasing breadths of host resistance and viral infectivity. However, experimental evidence generally indicates that such directional arms race dynamics do not continue indefinitely (25, 33, 34). Therefore, models of negative density-dependent fluctuations due to selective trade-offs, such as kill-the-winner, are often invoked (20, 33, 35, 36). In these models, fluctuations are generally considered to occur between rapidly growing competition specialists that are susceptible to infection and more slowly growing resistant strains that are considered defense specialists. Such negative density-dependent fluctuations are also likely to occur between strains that have differences in viral susceptibility ranges, such as those that would result from enhanced infection (30).The above coevolutionary processes are considered to be among the major mechanisms that have led to and maintain diversity within bacterial communities (32, 35, 37–39). These processes also influence genetic microdiversity within populations of closely related bacteria. This is especially the case for cell surface-related genes that are often localized to genomic islands (14, 40, 41), regions of high gene content, and gene sequence variability among members of a population. As such, populations in nature display an enormous degree of microdiversity in phage susceptibility regions, potentially leading to an assortment of subpopulations with different ranges of susceptibility to coexisting phages (4, 14, 30, 40).Prochlorococcus is a unicellular cyanobacterium that is the numerically dominant photosynthetic organism in vast oligotrophic expanses of the open oceans, where it contributes significantly to primary production (42, 43). Prochlorococcus consists of a number of distinct ecotypes (44–46) that form stable and reproducible population structures (7). These populations coexist in the oceans with tailed double-stranded DNA phage populations that infect them (47–49).Previously, we found that resistance to phage infection occurs frequently in two high-light–adapted Prochlorococcus ecotypes through spontaneous mutations in cell surface-related genes (14). These genes are primarily localized to genomic island 4 (ISL4) that displays a high degree of genetic diversity in environmental populations (14, 40). Although about a third of Prochlorococcus-resistant strains had no detectable associated cost, the others came with a cost manifested as either a slower growth rate or enhanced infection by other phages (14). In nature, Prochlorococcus seems to be growing close to its intrinsic maximal growth rate (50–52). This raises the question as to the fate of emergent resistant Prochlorococcus lineages in the environment, especially when resistance is accompanied with a high growth rate fitness cost.To begin addressing this question, we investigated the phenotype of Prochlorococcus strains with time after the acquisition of resistance. We found that resistant strains evolved toward an improved growth rate and a reduced resistance range. Whole-genome sequencing and PCR screening of many of these strains revealed that these phenotypic changes were largely due to additional, compensatory mutations, leading to increased genetic diversity. These findings suggest that the oceans are populated with rapidly growing Prochlorococcus cells with varying degrees of resistance and provide an explanation for how a multitude of presumably resistant Prochlorococcus cells are growing close to their maximal known growth rate in nature. 相似文献
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Arun K. Haldar Clémence Foltz Ryan Finethy Anthony S. Piro Eric M. Feeley Danielle M. Pilla-Moffett Masaki Komatsu Eva-Maria Frickel J?rn Coers 《Proceedings of the National Academy of Sciences of the United States of America》2015,112(41):E5628-E5637
Many microbes create and maintain pathogen-containing vacuoles (PVs) as an intracellular niche permissive for microbial growth and survival. The destruction of PVs by IFNγ-inducible guanylate binding protein (GBP) and immunity-related GTPase (IRG) host proteins is central to a successful immune response directed against numerous PV-resident pathogens. However, the mechanism by which IRGs and GBPs cooperatively detect and destroy PVs is unclear. We find that host cell priming with IFNγ prompts IRG-dependent association of Toxoplasma- and Chlamydia-containing vacuoles with ubiquitin through regulated translocation of the E3 ubiquitin ligase tumor necrosis factor (TNF) receptor associated factor 6 (TRAF6). This initial ubiquitin labeling elicits p62-mediated escort and deposition of GBPs to PVs, thereby conferring cell-autonomous immunity. Hypervirulent strains of Toxoplasma gondii evade this process via specific rhoptry protein kinases that inhibit IRG function, resulting in blockage of downstream PV ubiquitination and GBP delivery. Our results define a ubiquitin-centered mechanism by which host cells deliver GBPs to PVs and explain how hypervirulent parasites evade GBP-mediated immunity.Pathogen-containing vacuoles (PVs) provide a safe haven to many intracellular bacterial and protozoan pathogens (1). Within the vacuolar enclosure of PVs, these pathogens can accumulate nutrients required for microbial growth. Moreover, life within the vacuolar niche shields microbes from cytoplasmic immune sensors that, once activated, can trigger proinflammatory and cell-autonomous immune responses (1). Accordingly, many intracellular pathogens such as the bacterium Chlamydia trachomatis and the protozoan Toxoplasma gondii have successfully adapted to a vacuolar lifestyle.For the host to successfully combat infections with PV-resident microbes, the innate immune system must target PVs and its inhabitants for destruction. Critical mediators of host-directed attacks on PVs are two families of IFNγ-inducible GTPases: immunity-related GTPases (IRGs) and guanylate binding proteins (GBPs) (2). Members of both GTPase families play roles in host-mediated lysis of PVs, a process resulting in the release of microbes into the host cell cytoplasm, subsequent killing of PV-expelled microbes, and host cell death (3–8). Additionally, GBPs help deliver cytosolic subunits of the antimicrobial NADPH oxidase NOX2 for assembly on phagosomal membranes, orchestrate the capture of PV-resident microbes inside degradative autophagolysosomes, and promote the activation of canonical and noncanonical inflammasome pathways (5, 8–12). As a critical first step underlying most if not all of these known GBP-controlled cell-autonomous immune responses, GBPs must locate to their intracellular microbial targets.GBPs belong to the dynamin superfamily of large GTPases (13). Similar to other members of the dynamin superfamiliy, GBPs can assemble as oligomers in a nucleotide-dependent fashion (13). Binding of GTP results in dimer formation; subsequent GTP hydrolysis prompts conformational changes that enable GBPs to assemble as tetramers (14, 15). Mutations in the G domain that reduce nucleotide binding affinities and hydrolytic activity block GBP oligomerization, constrain the localization of GBPs to the cytoplasm, and prevent GBPs from binding to PV membranes (9, 15–18). These observations support a model in which GBP monomers are diffusely distributed in the cytoplasm and GBP oligomers associate with membranes. However, these observations fail to account for the specificity with which oligomeric GBPs agglomerate on PV membranes.PVs formed by C. trachomatis and T. gondii recruit not only GBPs but also members of the IRG family of IFNγ-inducible GTPase (4, 19). The IRG protein family can be divided into two subgroups: IRGM and GKS proteins (20). Whereas GKS proteins feature the canonical glycine–lysine–serine (GKS) P-loop sequence, IRGM proteins have a substitution of a lysine for a methionine in their P-loop sequence (20). IRGM and GKS proteins also differ in their subcellular localization: IRGM proteins associate with endomembranes, whereas monomeric GDP-bound GKS proteins predominantly reside within the host cell cytoplasm (4, 17, 21, 22). Once GKS proteins transition into a GTP-bound active state, they can bind to PV membranes (21). IRGM proteins inhibit this activation step and thereby guard IRGM-decorated membranes against GKS protein targeting (17, 21). Because PV membranes surrounding either C. trachomatis or T. gondii are largely devoid of IRGM proteins, they are the preferred GKS binding substrate following a “missing-self” principle of immune targeting (17, 23). In IRGM-deficient cells, however, GKS proteins enter the active state prematurely, form protein aggregates, mislocalize, and thus fail to bind to PVs (17, 21). Although these previous observations help explain how IRGM proteins promote the delivery of GKS proteins to PVs, IRGM proteins also control the subcellular localization of GBPs through an uncharacterized mechanism (6, 17, 24–26).Here, we report a previously unidentified host-directed ubiquitination pathway involved in innate immunity. We demonstrate that Chlamydia- and Toxoplasma-containing vacuoles become ubiquitin-decorated upon IFNγ priming of their host cells. IFNγ-dependent association of ubiquitin with PVs requires IFNγ-inducible IRG proteins and the E3 ligase tumor necrosis factor (TNF) receptor associated factor 6 (TRAF6). Experimental removal of the IFNγ-inducible ubiquitination pathway dramatically diminishes the p62-dependent delivery of GBPs to PVs and thereby renders host cells more susceptible to infections. Thus, our observations imply that ubiquitin serves as a host-induced pattern that marks intracellular structures as immune targets for members of the GBP family of host defense proteins. 相似文献
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Michael R. Yeaman Scott G. Filler Siyang Chaili Kevin Barr Huiyuan Wang Deborah Kupferwasser John P. Hennessey Jr. Yue Fu Clint S. Schmidt John E. Edwards Jr. Yan Q. Xiong Ashraf S. Ibrahim 《Proceedings of the National Academy of Sciences of the United States of America》2014,111(51):E5555-E5563
Increasing rates of life-threatening infections and decreasing susceptibility to antibiotics urge development of an effective vaccine targeting Staphylococcus aureus. This study evaluated the efficacy and immunologic mechanisms of a vaccine containing a recombinant glycoprotein antigen (NDV-3) in mouse skin and skin structure infection (SSSI) due to methicillin-resistant S. aureus (MRSA). Compared with adjuvant alone, NDV-3 reduced abscess progression, severity, and MRSA density in skin, as well as hematogenous dissemination to kidney. NDV-3 induced increases in CD3+ T-cell and neutrophil infiltration and IL-17A, IL-22, and host defense peptide expression in local settings of SSSI abscesses. Vaccine induction of IL-22 was necessary for protective mitigation of cutaneous infection. By comparison, protection against hematogenous dissemination required the induction of IL-17A and IL-22 by NDV-3. These findings demonstrate that NDV-3 protective efficacy against MRSA in SSSI involves a robust and complementary response integrating innate and adaptive immune mechanisms. These results support further evaluation of the NDV-3 vaccine to address disease due to S. aureus in humans.The bacterium Staphylococcus aureus is the leading cause of skin and skin structure infections (SSSIs), including cellulitis, furunculosis, and folliculitis (1–4), and a common etiologic agent of impetigo (5), erysipelas (6), and superinfection in atopic dermatitis (7). This bacterium is a significant cause of surgical or traumatic wound infections (8, 9), as well as decuibitus and diabetic skin lesions (10). Moreover, SSSI is an important risk factor for systemic infection. The skin is a key portal of entry for hematogenous dissemination, particularly in association with i.v. catheters. S. aureus is now the second most common bloodstream isolate in healthcare settings (11), and SSSI is a frequent source of invasive infections such as pneumonia or endocarditis (12, 13). Despite a recent modest decline in rates of methicillin-resistant S. aureus (MRSA) infection in some cohorts (13), infections due to S. aureus remain a significant problem (14, 15). Even with appropriate therapy, up to one-third of patients diagnosed with S. aureus bacteremia succumb—accounting for more attributable annual deaths than HIV, tuberculosis, and viral hepatitis combined (16).The empiric use of antibiotics in healthcare-associated and community-acquired settings has increased S. aureus exposure to these agents, accelerating selection of resistant strains. As a result, resistance to even the most recently developed agents is emerging at an alarming pace (17, 18). The impact of this trend is of special concern in light of high rates of mortality associated with invasive MRSA infection (e.g., 15–40% in bacteremia or endocarditis), even with the most recently developed antistaphylococcal therapeutics (19, 20). Moreover, patients who experience SSSI due to MRSA exhibit high 1-y recurrence rates, often prompting surgical debridement (21) and protracted antibiotic treatment.Infections due to MRSA are a special concern in immune-vulnerable populations, including hemodialysis (22), neutropenic (23, 24), transplantation (25), and otherwise immunosuppressed patients (26, 27), and in patients with inherited immune dysfunctions (28–31) or cystic fibrosis (32). Patients having deficient interleukin 17 (IL-17) or IL-22 responses (e.g., signal transduction mediators STAT3, DOCK8, or CARD9 deficiencies) exhibit chronic or “cold” abscesses, despite high densities of pathogens such as S. aureus (33, 34). For example, patients with Chronic Granulomatous Disease (CGD; deficient Th1 and oxidative burst response) have increased risk of disseminated S. aureus infection. In contrast, patients with Job’s Syndrome (deficient Th17 response) typically have increased risk to SSSI and lung infections, but less so for systemic S. aureus bacteremia (35, 36). This pattern contrasts that observed in neutropenic or CGD patients (37). These themes suggest efficacious host defenses against MRSA skin and invasive infections involve complementary but distinct molecular and cellular immune responses.From these perspectives, vaccines or immunotherapeutics that prevent or lessen severity of MRSA infections, or that enhance antibiotic efficacy, would be significant advances in patient care and public health. However, to date, there are no licensed prophylactic or therapeutic vaccine immunotherapies for S. aureus or MRSA infection. Unfortunately, efforts to develop vaccines targeting S. aureus capsular polysaccharide type 5 or 8 conjugates, or the iron-regulated surface determinant B protein, have not been successful thus far (38, 39). Likewise, passive immunization using monoclonal antibodies targeting the S. aureus adhesin clumping factor A (ClfA, tefibazumab) (40) or lipoteichoic acid (pagibaximab) (41) have not shown efficacy against invasive infections in human clinical studies to date. Moreover, the striking recurrence rates of SSSI due to MRSA imply that natural exposure does not induce optimal preventive immunity or durable anamnestic response to infection or reinfection. Thus, significant challenges exist in the development of an efficacious vaccine targeting diseases caused by S. aureus (42) that are perhaps not optimally addressed by conventional approaches.The NDV-3 vaccine reflects a new strategy to induce durable immunity targeting S. aureus. Its immunogen is engineered from the agglutinin-like sequence 3 (Als3) adhesin/invasin of Candida albicans, which we discovered to be a structural homolog of S. aureus adhesins (43). NDV-3 is believed to cross-protect against S. aureus and C. albicans due to sequence (T-cell) and conformational (B-cell) epitopes paralleled in both organisms (44). Our prior data have shown that NDV-3 is efficacious in murine models of hematogenous and mucosal candidiasis (45), as well as S. aureus bacteremia (46–48). Recently completed phase I clinical trials demonstrate the safety, tolerability, and immunogenicity of NDV-3 in humans (49). 相似文献
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Danica A. Helb Kevin K. A. Tetteh Philip L. Felgner Jeff Skinner Alan Hubbard Emmanuel Arinaitwe Harriet Mayanja-Kizza Isaac Ssewanyana Moses R. Kamya James G. Beeson Jordan Tappero David L. Smith Peter D. Crompton Philip J. Rosenthal Grant Dorsey Christopher J. Drakeley Bryan Greenhouse 《Proceedings of the National Academy of Sciences of the United States of America》2015,112(32):E4438-E4447
Tools to reliably measure Plasmodium falciparum (Pf) exposure in individuals and communities are needed to guide and evaluate malaria control interventions. Serologic assays can potentially produce precise exposure estimates at low cost; however, current approaches based on responses to a few characterized antigens are not designed to estimate exposure in individuals. Pf-specific antibody responses differ by antigen, suggesting that selection of antigens with defined kinetic profiles will improve estimates of Pf exposure. To identify novel serologic biomarkers of malaria exposure, we evaluated responses to 856 Pf antigens by protein microarray in 186 Ugandan children, for whom detailed Pf exposure data were available. Using data-adaptive statistical methods, we identified combinations of antibody responses that maximized information on an individual’s recent exposure. Responses to three novel Pf antigens accurately classified whether an individual had been infected within the last 30, 90, or 365 d (cross-validated area under the curve = 0.86–0.93), whereas responses to six antigens accurately estimated an individual’s malaria incidence in the prior year. Cross-validated incidence predictions for individuals in different communities provided accurate stratification of exposure between populations and suggest that precise estimates of community exposure can be obtained from sampling a small subset of that community. In addition, serologic incidence predictions from cross-sectional samples characterized heterogeneity within a community similarly to 1 y of continuous passive surveillance. Development of simple ELISA-based assays derived from the successful selection strategy outlined here offers the potential to generate rich epidemiologic surveillance data that will be widely accessible to malaria control programs.Many countries have extensive programs to reduce the burden of Plasmodium falciparum (Pf), the parasite responsible for most malaria morbidity and mortality (1). Effectively using limited resources for malaria control or elimination and evaluating interventions require accurate measurements of the risk of being infected with Pf (2–15). To reflect the rate at which individuals are infected with Pf in a useful way, metrics used to estimate exposure in a community need to account for dynamic changes over space and time, especially in response to control interventions (16–18).A variety of metrics can be used to estimate Pf exposure, but tools that are more precise and low cost are needed for population surveillance. Existing metrics have varying intrinsic levels of precision and accuracy and are subject to a variety of extrinsic factors, such as cost, time, and availability of trained personnel (19). For example, entomological measurements provide information on mosquito to human transmission for a community but are expensive, require specially trained staff, and lack standardized procedures, all of which reduce precision and/or make interpretation difficult (19–22). Parasite prevalence can be measured by detecting parasites in the blood of individuals from a cross-sectional sample of a community and is, therefore, relatively simple and inexpensive to perform, but results may be imprecise, especially in areas of low transmission (19, 23), and biased by a number of factors, including immunity and access to antimalarial treatment (5, 6, 19, 23–25). The burden of symptomatic disease in a community can be estimated from routine health systems data; however, such data are frequently unreliable (5, 26–28) and generally underestimate the prevalence of Pf infection in areas of intense transmission. Precise and quantitative information about exposure at an individual level can be reliably obtained from cohort studies by measuring the incidence of asymptomatic and/or symptomatic Pf infection (i.e., by measuring the molecular force of infection) (29–35). Unfortunately, the expense of cohort studies limits their use to research settings. The end result is that most malaria-endemic regions lack reliable, timely data on Pf exposure, limiting the capabilities of malaria control programs to guide and evaluate interventions.Serologic assays offer the potential to provide incidence estimates for symptomatic and asymptomatic Pf infection, which are currently obtained from cohort studies, at the cost of cross-sectional studies (36–38). Although Pf infections are transient, a record of infection remains detectable in an individual’s antibody profile. Thus, appropriately chosen antibody measurements integrated with age can provide information about an individual’s exposure history. Antibodies can be measured by simple ELISAs and obtained from dried blood spots, which are easy to collect, transport, and store (39–41). Serologic responses to Pf antigens have been explored as potential epidemiological tools (42–45), and estimated rates of seroconversion to well-characterized Pf antigens accurately reflect stable rates of exposure in a community, whereas distinct changes in these rates are obtained from successful interventions (22, 39, 41, 46–53). However, current serologic assays are not designed to detect short-term or gradual changes in Pf exposure or measure exposure to infection at an individual level. The ability to calibrate antibody responses to estimates of exposure in individuals could allow for more flexible sampling of a population (e.g., not requiring age stratification), improve accuracy of exposure estimates from small sample sizes, and better characterize heterogeneity in exposure within a community.Different Pf antigens elicit antibody responses with different magnitudes and kinetics, providing a large and diverse set of potential biomarkers for exposure (38, 54–58). We hypothesized that new and more highly informative serologic biomarkers better able to characterize an individual’s recent exposure history could be identified by analyzing antibody responses to a large number of candidate Pf antigens in participants with well-characterized exposure histories. To test this hypothesis, we probed plasma from participants in two cohort studies in Uganda against a protein microarray containing 856 Pf antigens. The primary aim of this analysis was to identify responses to select antigens that were most informative of recent exposure using robust, data-adaptive statistical methods. Each participant’s responses to these selected antigens were used as predictors for two primary outcomes of their recent exposure to Pf: (i) days since last Pf infection and (ii) the incidence of symptomatic malaria in the last year. These individual-level estimates were then aggregated across a population to assess community-level malaria exposure. The selection strategy presented here identified accurate biomarkers of exposure for children living in areas of moderate to high Pf exposure and illustrates the utility of this flexible and broadly applicable approach. 相似文献
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Xiaofan Li Hansi Liu Jose Chu Luo Sarah A. Rhodes Liana M. Trigg Damian B. van Rossum Andriy Anishkin Fortunay H. Diatta Jessica K. Sassic David K. Simmons Bishoy Kamel Monica Medina Mark Q. Martindale Timothy Jegla 《Proceedings of the National Academy of Sciences of the United States of America》2015,112(9):E1010-E1019
We examined the origins and functional evolution of the Shaker and KCNQ families of voltage-gated K+ channels to better understand how neuronal excitability evolved. In bilaterians, the Shaker family consists of four functionally distinct gene families (Shaker, Shab, Shal, and Shaw) that share a subunit structure consisting of a voltage-gated K+ channel motif coupled to a cytoplasmic domain that mediates subfamily-exclusive assembly (T1). We traced the origin of this unique Shaker subunit structure to a common ancestor of ctenophores and parahoxozoans (cnidarians, bilaterians, and placozoans). Thus, the Shaker family is metazoan specific but is likely to have evolved in a basal metazoan. Phylogenetic analysis suggested that the Shaker subfamily could predate the divergence of ctenophores and parahoxozoans, but that the Shab, Shal, and Shaw subfamilies are parahoxozoan specific. In support of this, putative ctenophore Shaker subfamily channel subunits coassembled with cnidarian and mouse Shaker subunits, but not with cnidarian Shab, Shal, or Shaw subunits. The KCNQ family, which has a distinct subunit structure, also appears solely within the parahoxozoan lineage. Functional analysis indicated that the characteristic properties of Shaker, Shab, Shal, Shaw, and KCNQ currents evolved before the divergence of cnidarians and bilaterians. These results show that a major diversification of voltage-gated K+ channels occurred in ancestral parahoxozoans and imply that many fundamental mechanisms for the regulation of action potential propagation evolved at this time. Our results further suggest that there are likely to be substantial differences in the regulation of neuronal excitability between ctenophores and parahoxozoans.Voltage-gated K+ channels are highly conserved among bilaterian metazoans and play a central role in the regulation of excitation in neurons and muscle. Understanding the functional evolution of these channels may therefore provide important insights into how neuromuscular excitation evolved within the Metazoa. Three major gene families, Shaker, KCNQ, and Ether-a-go-go (EAG) encode all voltage-gated K+ channels in bilaterians (1, 2). In this study, we examine the functional evolution and origins of the Shaker and KCNQ gene families. Shaker family channels can be definitively identified by a unique subunit structure that includes both a voltage-gated K+ channel core and a family-specific cytoplasmic domain within the N terminus known as the T1 domain. T1 mediates assembly of Shaker family subunits into functional tetrameric channels (3, 4). KCNQ channels are also tetrameric but lack a T1 domain and use a distinct coiled-coil assembly domain in the C terminus (5, 6). KCNQ channels can be identified by the presence of this family-specific assembly motif and high amino acid conservation within the K+ channel core. Both channel families are found in cnidarians (1, 7) and thus predate the divergence of cnidarians and bilaterians, but their ultimate evolutionary origins have not yet been defined.Shaker family K+ channels serve diverse roles in the regulation of neuronal firing and can be divided into four gene subfamilies based on function and sequence homology: Shaker, Shab, Shal, and Shaw (8, 9). The T1 assembly domain is only compatible between subunits from the same gene subfamily (4, 10) and thus serves to keep the subfamilies functionally segregated. Shaker subfamily channels activate rapidly near action potential threshold and range from rapidly inactivating to noninactivating. Multiple roles for Shaker channels in neurons and muscles have been described, but their most unique and fundamental role may be that of axonal action potential repolarization. Shaker channels are clustered to the axon initial segment and nodes of Ranvier in vertebrate neurons (11–13) and underlie the delayed rectifier in squid giant axons (14). The Shaker subfamily is diverse in cnidarians (15, 16), and the starlet sea anemone Nematostella vectensis has functional orthologs of most identified Shaker current types observed in bilaterians (16).The Shab and Shal gene subfamilies encode somatodendritic delayed rectifiers and A currents, respectively (17–20). Shab channels are important for maintaining sustained firing (21, 22), whereas the Kv4-based A current modulates spike threshold and frequency (17). Shab and Shal channels are present in cnidarians, but cnidarian Shab channels have not been functionally characterized, and the only cnidarian Shal channels expressed to date display atypical voltage dependence and kinetics compared with bilaterian channels (23). Shaw channels are rapid, high-threshold channels specialized for sustaining fast firing in vertebrates (24, 25) but have a low activation threshold and may contribute to resting potential in Drosophila (19, 26, 27). A Caenorhabditis elegans Shaw has slow kinetics but a high activation threshold (28), and a single expressed cnidarian Shaw channel has the opposite: a low activation threshold but relatively fast kinetics (29). Thus, the ancestral properties and function of Shaw channels is not yet understood. Further functional characterization of cnidarian Shab, Shal, and Shaw channels would provide a better understanding of the evolutionary status of the Shaker family in early parahoxozoans.KCNQ family channels underlie the M current in vertebrate neurons (30) that regulates subthreshold excitability (31). The M current provides a fundamental mechanism for regulation of firing threshold through the Gq G-protein pathway because KCNQ channels require phosphatidylinositol 4,5-bisphosphate (PIP2) for activation (32, 33). PIP2 hydrolysis and subsequent KCNQ channel closure initiated by Gq-coupled receptors produces slow excitatory postsynaptic potentials, during which the probability of firing is greatly increased (32, 33). The key functional adaptations of KCNQ channels for this physiological role that can be observed in vitro are (i) a requirement for PIP2 to couple voltage-sensor activation to pore opening (34, 35), and (ii) a hyperpolarized voltage–activation curve that allows channels to open below typical action potential thresholds. Both key features are found in vertebrate (30, 34, 36–38), Drosophila (39), and C. elegans (40) KCNQ channels, suggesting they may have been present in KCNQ channels in a bilaterian ancestor. Evolution of the M current likely represented a major advance in the ability to modulate the activity of neuronal circuits, but it is not yet clear when PIP2-dependent KCNQ channels first evolved.Here, we examine the origins and functional evolution of the Shaker and KCNQ gene families. If we assume the evolution of neuronal signaling provided a major selective pressure for the functional diversification of voltage-gated K+ channels, then we can hypothesize that the appearance of these gene families might accompany the emergence of the first nervous systems or a major event in nervous system evolution. Recent phylogenies that place the divergence of ctenophores near the root of the metazoan tree suggest that the first nervous systems, or at least the capacity to make neurons, may have been present in a basal metazoan ancestor (41–43) (Fig. S1). One hypothesis then is that much of the diversity of metazoan voltage-gated channels should be shared between ctenophores and parahoxozoans [cnidarians, bilaterians, and placozoans (44)]. However, genome analysis indicates that many “typical” neuronal genes are missing in ctenophores and the sponges lack a nervous system, leading to the suggestion that extant nervous systems may have evolved independently in ctenophores and parahoxozoans (42, 45). Thus, a second hypothesis is that important steps in voltage-gated K+ channel evolution might have occurred separately in ctenophores and parahoxozoans. We tested these hypotheses by carefully examining the phylogenetic distribution and functional evolution of Shaker and KCNQ family K+ channels. Our results support a model in which major innovations in neuromuscular excitability occurred specifically within the parahoxozoan lineage. 相似文献
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Yichun Shuai Areekul Hirokawa Yulian Ai Min Zhang Wanhe Li Yi Zhong 《Proceedings of the National Academy of Sciences of the United States of America》2015,112(48):E6663-E6672
Recent studies have identified molecular pathways driving forgetting and supported the notion that forgetting is a biologically active process. The circuit mechanisms of forgetting, however, remain largely unknown. Here we report two sets of Drosophila neurons that account for the rapid forgetting of early olfactory aversive memory. We show that inactivating these neurons inhibits memory decay without altering learning, whereas activating them promotes forgetting. These neurons, including a cluster of dopaminergic neurons (PAM-β′1) and a pair of glutamatergic neurons (MBON-γ4>γ1γ2), terminate in distinct subdomains in the mushroom body and represent parallel neural pathways for regulating forgetting. Interestingly, although activity of these neurons is required for memory decay over time, they are not required for acute forgetting during reversal learning. Our results thus not only establish the presence of multiple neural pathways for forgetting in Drosophila but also suggest the existence of diverse circuit mechanisms of forgetting in different contexts.Although forgetting commonly has a negative connotation, it is a functional process that shapes memory and cognition (1–4). Recent studies, including work in relatively simple invertebrate models, have started to reveal basic biological mechanisms underlying forgetting (5–15). In Drosophila, single-session Pavlovian conditioning by pairing an odor (conditioned stimulus, CS) with electric shock (unconditioned stimulus, US) induces aversive memories that are short-lasting (16). The memory performance of fruit flies is observed to drop to a negligible level within 24 h, decaying rapidly early after training and slowing down thereafter (17). Memory decay or forgetting requires the activation of the small G protein Rac, a signaling protein involved in actin remodeling, in the mushroom body (MB) intrinsic neurons (6). These so-called Kenyon cells (KCs) are the neurons that integrate CS–US information (18, 19) and support aversive memory formation and retrieval (20–22). In addition to Rac, forgetting also requires the DAMB dopamine receptor (7), which has highly enriched expression in the MB (23). Evidence suggests that the dopamine-mediated forgetting signal is conveyed to the MB by dopamine neurons (DANs) in the protocerebral posterior lateral 1 (PPL1) cluster (7, 24). Therefore, forgetting of olfactory aversive memory in Drosophila depends on a particular set of intracellular molecular pathways within KCs, involving Rac, DAMB, and possibly others (25), and also receives modulation from extrinsic neurons. Although important cellular evidence supporting the hypothesis that memory traces are erased under these circumstances is still lacking, these findings lend support to the notion that forgetting is an active, biologically regulated process (17, 26).Although existing studies point to the MB circuit as essential for forgetting, several questions remain to be answered. First, whereas the molecular pathways for learning and forgetting of olfactory aversive memory are distinct and separable (6, 7), the neural circuits seem to overlap. Rac-mediated forgetting has been localized to a large population of KCs (6), including the γ-subset, which is also critical for initial memory formation (21, 27). The site of action of DAMB for forgetting has yet to be established; however, the subgroups of PPL1-DANs implicated in forgetting are the same as those that signal aversive reinforcement and are required for learning (28–30). It leaves open the question of whether the brain circuitry underlying forgetting and learning is dissociable, or whether forgetting and learning share the same circuit but are driven by distinct activity patterns and molecular machinery (26). Second, shock reinforcement elicits multiple memory traces through at least three dopamine pathways to different subdomains in the MB lobes (28, 29). Functional imaging studies have also revealed Ca2+-based memory traces in different KC populations (31). It is poorly understood how forgetting of these memory traces differs, and it remains unknown whether there are multiple regulatory neural pathways. Notably, when PPL1-DANs are inactivated, forgetting still occurs, albeit at a lower rate (7). This incomplete block suggests the existence of an additional pathway(s) that conveys forgetting signals to the MB. Third, other than memory decay over time, forgetting is also observed through interference (32, 33), when new learning or reversal learning is introduced after training (6, 34, 35). Time-based and interference-based forgetting shares a similar dependence on Rac and DAMB (6, 7). However, it is not known whether distinct circuits underlie forgetting in these different contexts.In the current study, we focus on the diverse set of MB extrinsic neurons (MBENs) that interconnect the MB lobes with other brain regions, which include 34 MB output neurons (MBONs) of 21 types and ∼130 dopaminergic neurons of 20 types in the PPL1 and protocerebral anterior medial (PAM) clusters (36, 37). These neurons have been intensively studied in olfactory memory formation, consolidation, and retrieval in recent years (e.g., 24, 28–30, 38–48); however, their roles in forgetting have not been characterized except for the aforementioned PPL1-DANs. In a functional screen, we unexpectedly found that several Gal4 driver lines of MBENs showed significantly better 3-h memory retention when the Gal4-expressing cells were inactivated. The screen has thus led us to identify two types of MBENs that are not involved in initial learning but play important and additive roles in mediating memory decay. Furthermore, neither of these MBEN types is required for reversal learning, supporting the notion that there is a diversity of neural circuits that drive different forms of forgetting. 相似文献
18.
Xingjie Ren Jin Sun Benjamin E. Housden Yanhui Hu Charles Roesel Shuailiang Lin Lu-Ping Liu Zhihao Yang Decai Mao Lingzhu Sun Qujie Wu Jun-Yuan Ji Jianzhong Xi Stephanie E. Mohr Jiang Xu Norbert Perrimon Jian-Quan Ni 《Proceedings of the National Academy of Sciences of the United States of America》2013,110(47):19012-19017
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Activation of Big Grain1 significantly improves grain size by regulating auxin transport in rice 总被引:3,自引:0,他引:3
Linchuan Liu Hongning Tong Yunhua Xiao Ronghui Che Fan Xu Bin Hu Chengzhen Liang Jinfang Chu Jiayang Li Chengcai Chu 《Proceedings of the National Academy of Sciences of the United States of America》2015,112(35):11102-11107
Grain size is one of the key factors determining grain yield. However, it remains largely unknown how grain size is regulated by developmental signals. Here, we report the identification and characterization of a dominant mutant big grain1 (Bg1-D) that shows an extra-large grain phenotype from our rice T-DNA insertion population. Overexpression of BG1 leads to significantly increased grain size, and the severe lines exhibit obviously perturbed gravitropism. In addition, the mutant has increased sensitivities to both auxin and N-1-naphthylphthalamic acid, an auxin transport inhibitor, whereas knockdown of BG1 results in decreased sensitivities and smaller grains. Moreover, BG1 is specifically induced by auxin treatment, preferentially expresses in the vascular tissue of culms and young panicles, and encodes a novel membrane-localized protein, strongly suggesting its role in regulating auxin transport. Consistent with this finding, the mutant has increased auxin basipetal transport and altered auxin distribution, whereas the knockdown plants have decreased auxin transport. Manipulation of BG1 in both rice and Arabidopsis can enhance plant biomass, seed weight, and yield. Taking these data together, we identify a novel positive regulator of auxin response and transport in a crop plant and demonstrate its role in regulating grain size, thus illuminating a new strategy to improve plant productivity.Because it is one of the most important staple food crops cultivated worldwide, improvement of grain yield is a major focus of rice-breeding programs (1). Grain size is one of the determining factors of grain yield (2, 3). A number of quantitative trait loci (QTLs) controlling rice grain size have been identified in recent years (4–11). However, functional mechanisms of these genes remain largely unknown. Because QTLs usually have important functions in determining grain size, many of them have been widely selected in breeding processes or existed in modern elite varieties, and a certain QTL could be only applicable in certain varieties (12). Thus, exploration of new grain size-associated genes and elucidation of their functional mechanisms have great significance for further improvement of rice yield (12).Seed size, as well as other organ size, is controlled by various plant hormones, such as auxin, brassinosteroid, and cytokinin (10, 13, 14). A number of studies have demonstrated that auxin plays a vital role in organ size determination by affecting cell division, cell expansion, and differentiation (15–17). Auxin exists predominantly as indole-3-acetic acid (IAA) in plants, and genetic studies of its biosynthetic genes in Arabidopsis have demonstrated that IAA regulates many aspects of plant growth and development, including stem elongation, lateral branching, vascular development, and tropic growth responses (18, 19). Combined with biochemical studies, the tryptophan (Trp)-dependent IAA biosynthesis pathway has been clearly established involving the YUCCA family flavin monooxgenases (20). Importantly, the two-step pathway is highly conserved throughout the plant kingdom (21). Until very recently, the Trp-independent auxin biosynthetic pathway was elucidated as contributing to early embryogenesis in Arabidopsis (22). Primary auxin signaling is a rapid process initiated from the hormone perception by receptor TIR1, an F-box protein, followed by degradation of the negative regulator AUX/IAA proteins, and further release the downstream auxin response factors (ARFs) (23–26). However, how the ARFs work in plants remains elusive. Auxin transport, generally referring to the cell-to-cell transportation of the hormone directed basipetally from shoots to roots in vascular tissues, plays a critical role in auxin response (18). The transport involves a number of membrane-associated proteins, such as PINs (protein inhibitor of nNOS), AUX1 (AUXIN TRANSPORTER PROTEIN 1), and ABCBs (ATP-BINDING CASSETTE, SUB-FAMILY B PROTEINS) as efflux or influx carriers (27–30). Disruption of auxin transport induced by either gene mutations or chemical inhibitor treatment will lead to diverse development defects, such as decreased lateral organ initiation and defective tropic growth responses (27, 31–34).In this study, we identify a rice mutant, named big grain1-D (Bg1-D) because it is a dominant mutant having extralarge grain size. BG1 encodes a novel plasma membrane-associated protein, and is specifically induced by auxin treatment. We show that BG1 is a new positive regulator of auxin response involved in auxin transport, and demonstrate that manipulation of BG1 expression can greatly improve grain size and plant productivity. 相似文献