High resolution structure of Deinococcus bacteriophytochrome yields new insights into phytochrome architecture and evolution

Phytochromes are red/far red light photochromic photoreceptors that direct many photosensory behaviors in the bacterial, fungal, and plant kingdoms. They consist of an N-terminal domain that covalently binds a bilin chromophore and a C-terminal region that transmits the light signal, often through a histidine kinase relay. Using x-ray crystallography, we recently solved the first three-dimensional structure of a phytochrome, using the chromophore-binding domain of Deinococcus radiodurans bacterial phytochrome assembled with its chromophore, biliverdin IXalpha. Now, by engineering the crystallization interface, we have achieved a significantly higher resolution model. This 1.45A resolution structure helps identify an extensive buried surface between crystal symmetry mates that may promote dimerization in vivo. It also reveals that upon ligation of the C3(2) carbon of biliverdin to Cys(24), the chromophore A-ring assumes a chiral center at C2, thus becoming 2(R),3(E)-phytochromobilin, a chemistry more similar to that proposed for the attached chromophores of cyanobacterial and plant phytochromes than previously appreciated. The evolution of bacterial phytochromes to those found in cyanobacteria and higher plants must have involved greater fitness using more reduced bilins, such as phycocyanobilin, combined with a switch of the attachment site from a cysteine near the N terminus to one conserved within the cGMP phosphodiesterase/adenyl cyclase/FhlA domain. From analysis of site-directed mutants in the D. radiodurans phytochrome, we show that this bilin preference was partially driven by the change in binding site, which ultimately may have helped photosynthetic organisms optimize shade detection. Collectively, these three-dimensional structural results better clarify bilin/protein interactions and help explain how higher plant phytochromes evolved from prokaryotic progenitors.     

Community-level trachoma ecological associations and the use of geospatial analysis methods: A systematic review

BackgroundTrachoma is targeted for global elimination as a public health problem by 2030. Understanding individual, household, or community-associated factors that may lead to continued transmission or risk of recrudescence in areas where elimination has previously been achieved, is essential in reaching and maintaining trachoma elimination. We aimed to identify climatic, demographic, environmental, infrastructural, and socioeconomic factors associated in the literature with trachoma at community-level and assess the strength of their association with trachoma. Because of the potential power of geospatial analysis to delineate the variables most strongly associated with differences in trachoma prevalence, we then looked in detail at geospatial analysis methods used in previous trachoma studies.MethodsWe conducted a systematic literature review using five databases: Medline, Embase, Global Health, Dissertations & Theses Global, and Web of Science, including publications from January 1950 to January 2021. The review protocol was prospectively registered with PROSPERO (CRD42020191718).ResultsOf 35 eligible studies, 29 included 59 different trachoma-associated factors, with eight studies also including spatial analysis methods. Six studies included spatial analysis methods only. Higher trachomatous inflammation—follicular (TF) prevalence was associated with areas that: had lower mean annual precipitation, lower mean annual temperatures, and lower altitudes; were rural, were less accessible, had fewer medical services, had fewer schools; and had lower access to water and sanitation. Higher trachomatous trichiasis (TT) prevalence was associated with higher aridity index and increased distance to stable nightlights. Of the 14 studies that included spatial methods, 11 used exploratory spatial data analysis methods, three used interpolation methods, and seven used spatial modelling methods.ConclusionResearchers and decision-makers should consider the inclusion and potential influence of trachoma-associated factors as part of both research activities and programmatic priorities. The use of geospatial methods in trachoma studies remains limited but offers the potential to define disease hotspots and areas of potential recrudescence to inform local, national, and global programmatic needs.     

Phylogenomic analysis supports the monophyly of cryptophytes and haptophytes and the association of rhizaria with chromalveolates

Here we use phylogenomics with expressed sequence tag (EST) data from the ecologically important coccolithophore-forming alga Emiliania huxleyi and the plastid-lacking cryptophyte Goniomonas cf. pacifica to establish their phylogenetic positions in the eukaryotic tree. Haptophytes and cryptophytes are members of the putative eukaryotic supergroup Chromalveolata (chromists [cryptophytes, haptophytes, stramenopiles] and alveolates [apicomplexans, ciliates, and dinoflagellates]). The chromalveolates are postulated to be monophyletic on the basis of plastid pigmentation in photosynthetic members, plastid gene and genome relationships, nuclear "host" phylogenies of some chromalveolate lineages, unique gene duplication and replacements shared by these taxa, and the evolutionary history of components of the plastid import and translocation systems. However the phylogenetic position of cryptophytes and haptophytes and the monophyly of chromalveolates as a whole remain to be substantiated. Here we assess chromalveolate monophyly using a multigene dataset of nuclear genes that includes members of all 6 eukaryotic supergroups. An automated phylogenomics pipeline followed by targeted database searches was used to assemble a 16-protein dataset (6,735 aa) from 46 taxa for tree inference. Maximum likelihood and Bayesian analyses of these data support the monophyly of haptophytes and cryptophytes. This relationship is consistent with a gene replacement via horizontal gene transfer of plastid-encoded rpl36 that is uniquely shared by these taxa. The haptophytes + cryptophytes are sister to a clade that includes all other chromalveolates and, surprisingly, two members of the Rhizaria, Reticulomyxa filosa and Bigelowiella natans. The association of the two Rhizaria with chromalveolates is supported by the approximately unbiased (AU)-test and when the fastest evolving amino acid sites are removed from the 16-protein alignment.     

Photosynthetic eukaryotes unite: endosymbiosis connects the dots

The photosynthetic organelle of algae and plants (the plastid) traces its origin to a primary endosymbiotic event in which a previously non-photosynthetic protist engulfed and enslaved a cyanobacterium. This eukaryote then gave rise to the red, green and glaucophyte algae. However, many algal lineages, such as the chlorophyll c-containing chromists, have a more complicated evolutionary history involving a secondary endosymbiotic event, in which a protist engulfed an existing eukaryotic alga (in this case, a red alga). Chromists such as diatoms and kelps then rose to great importance in aquatic habitats. Another algal group, the dinoflagellates, has undergone tertiary (engulfment of a secondary plastid) and even quaternary endosymbioses. In this review, we examine algal diversity and show endosymbiosis to be a major force in algal evolution. This area of research has advanced rapidly and long-standing issues such as the chromalveolate hypothesis and the extent of endosymbiotic gene transfer have recently been clarified.     

Conservation in the community: the Kilum-Ijim Forest Project,Cameroon

Thomas, D.H.L., Anders, S. & Penn, N.J. 2000. Conservation in the community: the Kilum-Ijim Forest Project, Cameroon. Ostrich 71 (1 & 2): 157–161.

BirdLife International has worked for ten years in Cameroon's North-West Province to assist the local people and government of Cameroon in achieving the conservation and sutainable management of the last significant remnant of a unique montane forest ecosystem. The Kilum/Ijim forests are almost certainly the last remaining habitat for the conservation of two endemic and threatened bird species, Bannerman's Turaco Tauraco bannermani and Banded Wattle-eye Platytiera laticincta. The forests have no legal designation as formal protected areas but have been conserved through local concern and knowledge of the multiple values of an intact forest ecosystem, backed up by the enforcement of traditional regulations, and support from the project to finding solutions to peoples' land use and natural resource management needs within and outside the forests. The project is now working under the umbrella of a national Biodiversity Conservation and Management Programme in Cameroon, funded by the Global Environment Facility, to establish the Kilum/Ijirn as the first community-managed forest in the country. A brief history and background to the project is presented, with emphasis on lessons learned by BiraLife and the success of current efforts to achieve the conservation of the forests through community management. The issues are discussed in the general context of an integrated conservation and development programme (ICDP).     

Association of monosodium glutamate intake with overweight in Chinese adults: the INTERMAP Study

Animal studies indicate that monosodium glutamate (MSG) can induce hypothalamic lesions and leptin resistance, possibly influencing energy balance, leading to overweight. This study examines the association between MSG intake and overweight in humans. We conducted a cross-sectional study involving 752 healthy Chinese (48.7% women), aged 40-59 years, randomly sampled from three rural villages in north and south China. The great majority of participants prepared their foods at home, without use of commercially processed foods. Diet was assessed with four in-depth multipass 24-h recalls. Participants were asked to demonstrate MSG amounts added in food preparation. Amounts shaken out were weighed by trained interviewers. Overweight was defined as BMI > or =25.0 or > or =23.0 kg/m(2)(based on World Health Organization recommendations for Asian populations). Eighty-two percent of participants were MSG users. Average intake was 0.33 g/day (s.d. = 0.40). With adjustment for potential confounders including physical activity and total energy intake, MSG intake was positively related to BMI. Prevalence of overweight was significantly higher in MSG users than nonusers. For users in the highest tertile of MSG intake compared to nonusers, the multivariable-adjusted odds ratios of overweight (BMI > or =23.0 and > or =25.0) were 2.10 (95% confidence interval, 1.13-3.90, P for trend across four MSG categories = 0.03) and 2.75 (95% confidence interval, 1.28-5.95, P = 0.04). This research provides data that MSG intake may be associated with increased risk of overweight independent of physical activity and total energy intake in humans.     

Serotype-Specific Structural Differences in the Protease-Cofactor Complexes of the Dengue Virus Family

With an estimated 40% of the world population at risk, dengue poses a significant threat to human health, especially in tropical and subtropical regions. Preventative and curative efforts, such as vaccine development and drug discovery, face additional challenges due to the occurrence of four antigenically distinct serotypes of the causative dengue virus (DEN1 to -4). Complex immune responses resulting from repeat assaults by the different serotypes necessitate simultaneous targeting of all forms of the virus. One of the promising targets for drug development is the highly conserved two-component viral protease NS2B-NS3, which plays an essential role in viral replication by processing the viral precursor polyprotein into functional proteins. In this paper, we report the 2.1-Å crystal structure of the DEN1 NS2B hydrophilic core (residues 49 to 95) in complex with the NS3 protease domain (residues 1 to 186) carrying an internal deletion in the N terminus (residues 11 to 20). While the overall folds within the protease core are similar to those of DEN2 and DEN4 proteases, the conformation of the cofactor NS2B is dramatically different from those of other flaviviral apoprotease structures. The differences are especially apparent within its C-terminal region, implicated in substrate binding. The structure reveals for the first time serotype-specific structural elements in the dengue virus family, with the reported alternate conformation resulting from a unique metal-binding site within the DEN1 sequence. We also report the identification of a 10-residue stretch within NS3pro that separates the substrate-binding function from the catalytic turnover rate of the enzyme. Implications for broad-spectrum drug discovery are discussed.Dengue fever (DF) affects tens of millions of people each year, with an average mortality rate of 5%, comprising mostly young children. The increasing spread and frequency of global epidemics of this disease have heightened the urgency for developing effective strategies for prevention, diagnosis, and treatment. Though several groups are involved in vaccine development (17, 23, 25), dengue presents a unique, complex challenge. Four antigenically distinct serotypes of the causative dengue virus (DENV) (DEN1 to -4) occur in nature, with differing pathogenicities in partially overlapping geographic locations. In individuals previously exposed to a certain serotype, repeat assault by a different serotype can lead to potentially fatal complications of the disease, dengue hemorrhagic fever and dengue shock syndrome. The presence of subneutralizing levels of antibodies against the first serotype, resulting in antibody-dependent enhancement (ADE), is believed to be a causative mechanism underlying this complication. Forty percent of the world population lives in dengue risk areas, mainly in tropical and subtropical regions, which necessitates the development of vaccines and therapeutics that can simultaneously protect against all four serotypes (24).DEN1 to -4 belong to the family Flaviviridae (genus Flavivirus) of positive-stranded RNA viruses transmitted by Aedes aegypti mosquitoes. Upon infection, the genomic RNA is translated by the host cell machinery into a 370-kDa polyprotein, which is subsequently cleaved and processed into 10 distinct structural (C, E, and prM) and nonstructural (NS1, 2A, 2B, 3, 4A, 4B, and 5) proteins. A majority of this processing (junctions of NS2A/2B, 2B/3, 3/4A, and 4B/5, as well as internal sites within C, 2A, 3, and 4A) is carried out by a virus-encoded two-component protease, NS2B-NS3. Additional processing, especially of sites toward the N terminus (junctions of C/prM, prM/E, E/NS1, NS1/NS2A, and NS4A/4B), employs cellular proteases, such as signalase. The NS2B-NS3-mediated cleavage forms an essential step in the viral replicative cycle, as evidenced by the lack of production of infectious virions in mutants carrying inactivating substitutions in the protease. Of the duo, the N terminus of NS3 encodes the enzymatic core, while a hydrophilic core within NS2B provides an essential cofactor function (9).Owing to its essential nature in viral replication and the promise and success of drugs targeting the proteases in the treatment of human immunodeficiency virus (HIV) and hepatitis C virus (HCV) infections (4), several recent studies have concentrated on identifying inhibitors of the flaviviral protease (5, 10, 21, 26-28). An added advantage of targeting the protease is that it is highly conserved between the serotypes (63 to 74%). For such structure-based drug design efforts, the availability of three-dimensional (3D) structures is an essential prerequisite, and given the differing pathogenicities and immune complexity generated by the multiple serotypes, it is a prudent first step to determine the structures of all four DENV proteases. The crystal structures of the DEN2 and DEN4 proteases in complex with various lengths of NS2B are available (7, 15). In this paper, we present the structures of an optimal construct of DEN1pro in complex with the essential hydrophilic core of NS2B (NS2Bc), as well as its active-site mutant. These structures reveal a novel and unexpected serotype-specific structural element embedded in the DENV genome, with implications for the discovery of broad-spectrum drugs. We also report the identification of a 10-residue stretch within the NS3 sequence that allows the separation of the substrate-binding function of the enzyme from its catalytic efficiency.