Nascent chains derived from a foldable protein sequence interact with specific ribosomal surface sites near the exit tunnel.

In order to become bioactive, proteins must be translated and protected from aggregation during biosynthesis. The ribosome and molecular chaperones play a key role in this process. Ribosome-bound nascent chains (RNCs) of intrinsically disordered proteins and RNCs bearing a signal/arrest sequence are known to interact with ribosomal proteins. However, in the case of RNCs bearing foldable protein sequences, not much information is available on these interactions. Here, via a combination of chemical crosslinking and time-resolved fluorescence-anisotropy, we find that nascent chains of the foldable globin apoHmp1-140 interact with ribosomal protein L23 and have a freely-tumbling non-interacting N-terminal compact region comprising 63-94 residues. Longer RNCs (apoHmp1-189) also interact with an additional yet unidentified ribosomal protein, as well as with chaperones. Surprisingly, the apparent strength of RNC/r-protein interactions does not depend on nascent-chain sequence. Overall, foldable nascent chains establish and expand interactions with selected ribosomal proteins and chaperones, as they get longer. These data are significant because they reveal the interplay between independent conformational sampling and nascent-protein interactions with the ribosomal surface.

Mapping Protein-Protein Interactions at Birth: Single-Particle Cryo-EM Analysis of a Ribosome-Nascent Globin Complex.

Interactions between ribosome-bound nascent chains (RNCs) and ribosomal components are critical to elucidate the mechanism of cotranslational protein folding. Nascent protein-ribosome contacts within the ribosomal exit tunnel were previously assessed mostly in the presence of C-terminal stalling sequences, yet little is known about contacts taking place in the absence of these strongly interacting motifs. Further, there is nearly no information about ribosomal proteins (r-proteins) interacting with nascent chains within the outer surface of the ribosome. Here, we combine chemical cross-linking, single-particle cryo-EM, and fluorescence anisotropy decays to determine the structural features of ribosome-bound apomyoglobin (apoMb). Within the ribosomal exit tunnel core, interactions are similar to those identified in previous reports. However, once the RNC enters the tunnel vestibule, it becomes more dynamic and interacts with ribosomal RNA (rRNA) and the L23 r-protein. Remarkably, on the outer surface of the ribosome, RNCs interact mainly with a highly conserved nonpolar patch of the L23 r-protein. RNCs also comprise a compact and dynamic N-terminal region lacking contact with the ribosome. In all, apoMb traverses the ribosome and interacts with it via its C-terminal region, while N-terminal residues sample conformational space and form a compact subdomain before the entire nascent protein sequence departs from the ribosome.

Is the answer to reducing early childhood caries in your pocket?

DATA SOURCES: Electronic scientific databases Embase, CINAHL, MEDLINE, PsycINFO and Web of Science were systematically searched and restricted to articles published from 1996 onwards and limited to articles published in English. This was carried out following an initial scoping search using keywords conducted in PubMed. STUDY SELECTION: Original studies investigating the use of mobile phone applications as a delivery method of healthcare interventions to parents and caregivers with children ≤6 years of age were included. As this was a mixed-methods systematic review, studies that have quantitative clinical outcomes and also qualitative outcomes of experiences, attitudes and beliefs of parents and caregivers were included. EndNote X8.2 and Rayyan.ai software was employed for title and abstract screening. DATA EXTRACTION AND SYNTHESIS: Three independent authors developed a combined data extraction tool to examine titles, abstracts and full texts of relevant articles against the inclusion criteria. The development of this tool was guided by the JBI reviewer's manual. Data extraction was completed by one reviewer, and verified by two further reviewers. Disagreements were resolved by discussion. Retrieved studies were assessed in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Data extracted included study reference information, study design, setting, sample sizes and intervention characteristics. A risk of bias assessment was undertaken using the Quality Appraisal for Diverse Studies tool, and a further risk assessment of quantitative and mixed methods studies. RESULTS: From 5953 studies initially identified, 5 studies were included in the review. One study identified using a gamification design within a mobile health app to promote oral health had statistically significant improvements in plaque and gingival indices compared to a control group at both a 6 and 12-week recall. Two studies reported a significant improvement in maternal knowledge of children's oral health as a result of using an oral health app. CONCLUSIONS: The delivery of oral health promotion through mobile health apps may be effective in reducing early childhood caries through improving health literacy in parents and caregivers, however key challenges in the app development process exist surrounding privacy issues and data protection.

Critical Beginnings: Selective Tuning of Solubility and Structural Accuracy of Newly Synthesized Proteins by the Hsp70 Chaperone System.

Proteins are particularly prone to aggregation immediately after release from the ribosome, and it is therefore important to elucidate the role of chaperones during these key steps of protein life. The Hsp70 and trigger factor (TF) chaperone systems interact with nascent proteins during biogenesis and immediately post-translationally. It is unclear, however, whether these chaperones can prevent formation of soluble and insoluble aggregates. Here, we address this question by monitoring the solubility and structural accuracy of globin proteins biosynthesized in an Escherichia coli cell-free system containing different concentrations of the bacterial Hsp70 and TF chaperones. We find that Hsp70 concentrations required to grant solubility to newly synthesized proteins are extremely sensitive to client-protein sequence. Importantly, Hsp70 concentrations yielding soluble client proteins are insufficient to prevent formation of soluble aggregates. In fact, for some aggregation-prone protein variants, avoidance of soluble-aggregate formation demands Hsp70 concentrations that exceed cellular levels in E. coli. In all, our data highlight the prominent role of soluble aggregates upon nascent-protein release from the ribosome and show the limitations of the Hsp70 chaperone system in the case of highly aggregation-prone proteins. These results demonstrate the need to devise better strategies to prevent soluble-aggregate formation upon release from the ribosome.

Distribution and solvent exposure of Hsp70 chaperone binding sites across the Escherichia coli proteome.

Many proteins must interact with molecular chaperones to achieve their native state in the cell. Yet, how chaperone binding-site characteristics affect the folding process is poorly understood. The ubiquitous Hsp70 chaperone system prevents client-protein aggregation by holding unfolded conformations and by unfolding misfolded states. Hsp70 binding sites of client proteins comprise a nonpolar core surrounded by positively charged residues. However, a detailed analysis of Hsp70 binding sites on a proteome-wide scale is still lacking. Further, it is not known whether proteins undergo some degree of folding while chaperone bound. Here, we begin to address the above questions by identifying Hsp70 binding sites in 2258 Escherichia coli (E. coli) proteins. We find that most proteins bear at least one Hsp70 binding site and that the number of Hsp70 binding sites is directly proportional to protein size. Aggregation propensity upon release from the ribosome correlates with number of Hsp70 binding sites only in the case of large proteins. Interestingly, Hsp70 binding sites are more solvent-exposed than other nonpolar sites, in protein native states. Our findings show that the majority of E. coli proteins are systematically enabled to interact with Hsp70 even if this interaction only takes place during a fraction of the protein lifetime. In addition, our data suggest that some conformational sampling may take place within Hsp70-bound states, due to the solvent exposure of some chaperone binding sites in native proteins. In all, we propose that Hsp70-chaperone-binding traits have evolved to favor Hsp70-assisted protein folding devoid of aggregation.

Protein folding in vitro and in the cell: From a solitary journey to a team effort.

Correct protein folding is essential for the health and function of living organisms. Yet, it is not well understood how unfolded proteins reach their native state and avoid aggregation, especially within the cellular milieu. Some proteins, especially small, single-domain and apparent two-state folders, successfully attain their native state upon dilution from denaturant. Yet, many more proteins undergo misfolding and aggregation during this process, in a concentration-dependent fashion. Once formed, native and aggregated states are often kinetically trapped relative to each other. Hence, the early stages of protein life are absolutely critical for proper kinetic channeling to the folded state and for long-term solubility and function. This review summarizes current knowledge on protein folding/aggregation mechanisms in buffered solution and within the bacterial cell, highlighting early stages. Remarkably, teamwork between nascent chain, ribosome, trigger factor and Hsp70 molecular chaperones enables all proteins to overcome aggregation propensities and reach a long-lived bioactive state.

Fluorescence Anisotropy Decays and Microscale-Volume Viscometry Reveal the Compaction of Ribosome-Bound Nascent Proteins.

This work introduces a technology that combines fluorescence anisotropy decay with microscale-volume viscometry to investigate the compaction and dynamics of ribosome-bound nascent proteins. Protein folding in the cell, especially when nascent chains emerge from the ribosomal tunnel, is poorly understood. Previous investigations based on fluorescence anisotropy decay determined that a portion of the ribosome-bound nascent protein apomyoglobin (apoMb) forms a compact structure. This work, however, could not assess the size of the compact region. The combination of fluorescence anisotropy with microscale-volume viscometry, presented here, enables identifying the size of compact nascent-chain subdomains using a single fluorophore label. Our results demonstrate that the compact region of nascent apoMb contains 57-83 amino acids and lacks residues corresponding to the two native C-terminal helices. These amino acids are necessary for fully burying the nonpolar residues in the native structure, yet they are not available for folding before ribosome release. Therefore, apoMb requires a significant degree of post-translational folding for the generation of its native structure. In summary, the combination of fluorescence anisotropy decay and microscale-volume viscometry is a powerful approach to determine the size of independently tumbling compact regions of biomolecules. This technology is of general applicability to compact macromolecules linked to larger frameworks.

Remembering cultural experiences: lifespan distributions, richness and content of autobiographical memories of museum visits.

Little is known about our autobiographical memories for cultural events. This represents an opportunity for cultural institutions such as museums, as examination of visitor memories is one way in which they can seek to understand the long-term impact they may have on their visitors. This research applied a coding model developed from autobiographical memory theory to analysis of participants' memories for museum visits, considering the distribution of memories across the life span, types of memories and content. Differences between visitor groups (age, visit frequency) were also considered. Findings showed a strong recency effect in the life-span distribution, suggesting the importance of social sharing in memories of cultural experience. Analysis of content showed a hierarchy of information that was present in museum memories. Knowledge acquired during the event of the visit was important, as was contextualising information whereby visitors situated the memory within their autobiographical knowledge and chronology. Emotions and thoughts were also salient. Visitor differences had minimal impact on content, with the exception of some effects that were consistent with the literature on memory and ageing. This research develops understanding of autobiographical memories for cultural experiences and provides insight to museums, with practical implications in terms of understanding visitors' experiences.

Multicentre study of physical abuse and limb fractures in young children in the East Anglia Region, UK.

OBJECTIVE: To determine if the detection of physical abuse in young children with fractures is of uniform high standard in the East Anglia Region of the UK, and whether we can identify areas for improvement in our detection of high-risk groups. DESIGN: Multicentre retrospective 4-year study. SETTING: 7 hospitals across the East Anglia Region of Britain (East Anglia Paediatric Physical Abuse and Fractures study). PARTICIPANTS: Age groups and fractures indicated as being at higher risk for physical abuse (all children under 12 months of age, and fractures of humerus and femur in children under 36 months of age). OUTCOME MEASURES: Our criterion for physical abuse was the decision of a multiagency child protection case conference (CPCC). RESULTS: Probability of CPCC decision of physical abuse was highest in infants, ranging from 50% of fractures sustained in the first month of life (excluding obstetric injuries) to 10% at 12 months of age. Only 46%-86% of infants (under 12 months) with a fracture were assessed by a paediatrician for physical abuse after their fracture. Significant variation in the use of skeletal surveys and in CPCC decision of physical abuse was noted in children attending different hospitals. CONCLUSIONS: It is a concern that significant variation between hospitals was found in the investigation and detection of physical abuse as confirmed by CPCC decisions. To minimise failure to detect true cases of physical abuse, we recommend that all high-risk children should be assessed by a paediatrician prior to discharge from the emergency department. Our proposed criteria for assessment (where we found probability of CPCC decision of physical abuse was at least 10%) are any child under the age of 12 months with any fracture, under 18 months of age with femur fracture and under 24 months with humeral shaft fracture (not supracondylar).

Building flux capacity: Citizen scientists increase resolution of soil greenhouse gas fluxes.

Though citizen science programs have been broadly successful in diverse scientific fields, their adoption has lagged in some disciplines, including soil science and ecosystem ecology. Collaborations with citizen scientists may be viewed as a conundrum in these disciplines, which often require substantial labor and technical experience; citizen scientists could improve sampling capacity but may reduce sample quality or require training and oversight prior to and while performing specialized tasks. To demonstrate the feasibility of incorporating citizen scientists into soil biogeochemistry research, we conducted a proof-of-concept study in high-elevation meadows of the Sierra Nevada in California. A collaboration between university researchers and citizen scientists allowed us to assess spatial and diel patterns of soil greenhouse gas (GHG) fluxes with an intensity and frequency that would otherwise be beyond the capacity of a typical research laboratory. This collaboration with citizen scientists increased our sampling intensity by over 700% while only doubling the sampling error relative to that of full-time researchers. With training and support from project scientists, citizen scientists collected data that demonstrate spatial independence of carbon dioxide, methane, and nitrous oxide at scales between 1 m and 175 m. Additionally, we found a lack of temporal variation over a 24-h period for all three GHGs. Citizen scientists participating in this one-day event reported levels of satisfaction commensurate with longer-term, immersive campaigns. The place-based event also proved an effective tool for teaching intangible concepts of soil biogeochemistry and promoting local conservation. Despite perceived barriers to entry, this study demonstrates the mutual benefits of citizen science collaborations in soil science and ecosystem ecology, encouraging adoption by disciplines that have been slow to take advantage of such collaborations. Short-term, local citizen science events can provide meaningful experiences for area residents and teach global biogeochemical cycles in a place-based context.

Barcoding in trypanosomes.

Trypanosomes (genus Trypanosoma) are parasites of humans, and wild and domestic mammals, in which they cause several economically and socially important diseases, including sleeping sickness in Africa and Chagas disease in the Americas. Despite the development of numerous molecular diagnostics and increasing awareness of the importance of these neglected parasites, there is currently no universal genetic barcoding marker available for trypanosomes. In this review we provide an overview of the methods used for trypanosome detection and identification, discuss the potential application of different barcoding techniques and examine the requirements of the 'ideal' trypanosome genetic barcode. In addition, we explore potential alternative genetic markers for barcoding Trypanosoma species, including an analysis of phylogenetically informative nucleotide changes along the length of the 18S rRNA gene.

Microarchitecture of the tsetse fly proboscis.

BACKGROUND: Tsetse flies (genus Glossina) are large blood-sucking dipteran flies that are important as vectors of human and animal trypanosomiasis in sub-Saharan Africa. Tsetse anatomy has been well described, including detailed accounts of the functional anatomy of the proboscis for piercing host skin and sucking up blood. The proboscis also serves as the developmental site for the infective metacyclic stages of several species of pathogenic livestock trypanosomes that are inoculated into the host with fly saliva. To understand the physical environment in which these trypanosomes develop, we have re-examined the microarchitecture of the tsetse proboscis. RESULTS: We examined proboscises from male and female flies of Glossina pallidipes using light microscopy and scanning electron microscopy (SEM). Each proboscis was removed from the fly head and either examined intact or dissected into the three constituent components: Labrum, labium and hypopharynx. Our light and SEM images reaffirm earlier observations that the tsetse proboscis is a formidably armed weapon, well-adapted for piercing skin, and provide comparative data for G. pallidipes. In addition, the images reveal that the hypopharynx, the narrow tube that delivers saliva to the wound site, ends in a remarkably ornate and complex structure with around ten finger-like projections, each adorned with sucker-like protrusions, contradicting previous descriptions that show a simple, bevelled end like a hypodermic needle. The function of the finger-like projections is speculative; they appear to be flexible and may serve to protect the hypopharynx from influx of blood or microorganisms, or control the flow of saliva. Proboscises were examined after colonisation by Trypanosoma congolense savannah. Consistent with the idea that colonisation commences in the region nearest the foregut, the highest densities of trypanosomes were found in the region of the labrum proximal to the bulb, although high densities were also found in other regions of the labrum. Trypanosomes were visible through the thin wall of the hypopharynx by both light microscopy and SEM. CONCLUSIONS: We highlight the remarkable architecture of the tsetse proboscis, in particular the intricate structure of the distal end of the hypopharynx. Further work is needed to elucidate the function of this intriguing structure.

Rediscovery of Trypanosoma (Pycnomonas) suis, a tsetse-transmitted trypanosome closely related to T. brucei.

The African tsetse-transmitted trypanosomes are considered to be a well-known group of parasitic protozoa, but in 2008 a novel and distinctive trypanosome related to Trypanosoma brucei was discovered among tsetse isolates from Msubugwe in Tanzania. The host range, distribution and potential pathogenicity of this new trypanosome remain to be elucidated; such studies would be facilitated by a sensitive and specific identification method. Here, we identified two highly repetitive elements in the genome of the new trypanosome: a 177 bp repeat, which was located predominantly on the highly abundant minichromosomes, and a 138 bp repeat, which was widely dispersed in the genome. A PCR test based on each repeat was specific for the new trypanosome and sensitive to <0.1 trypanosome equivalent. These PCR tests were used to identify trypanosomes in archival pig blood smears from the 1950's, confirming the identity of the Msubugwe trypanosome as Trypanosoma (Pycnomonas) suis. We also present data on the molecular karyotype and spliced leader (SL, miniexon) repeat of the new trypanosome, both of which distinguish T. suis from other, better-known African tsetse-transmitted trypanosomes. The rediscovery of T. suis opens new lines of research into the evolution and biology of the African trypanosomes.

Active haemorrhage of a renal allograft detected on portable ultrasound.

Function of a renal allograft relies on the integrity of its vascular anatomy. Renal biochemistry, ultrasound and percutaneous biopsy are used in combination to determine allograft function. Biopsy is not without risk, and in this case study we demonstrate a rare but a potentially life-threatening complication of renal allograft biopsy.

Perceived calorie intake and state body-image satisfaction in women attempting weight loss: a preliminary investigation.

State body-image satisfaction levels (BIS) can be predicted by appearance concerns, eating attitudes and body mass index (BMI). Determinants of state BIS and its variability were examined in women attempting weight loss. Little is known about contextual cues that influence state BIS; therefore the effect of eating on BIS was examined. Forty-six females attending a commercial weight loss group completed measures of shape and weight concerns, appearance beliefs and dietary restraint, followed by completion of a diary that assessed state BIS, mood and perceived calorie intake immediately following evening meals. Regression analysis indicated that after controlling for mood, state BIS was associated with higher BMI and dietary restraint. Greater variability in state BIS was associated with greater variability in perceived calorie intake. This relationship was fully mediated by greater variability in mood. The findings imply that state BIS warrants investigation as a process that may influence weight loss management.