Expression of breast cancer resistance protein (BCRP/ABCG2) in human placenta throughout gestation and at term before and after labor

Breast cancer resistance protein, BCRP, is a multidrug resistance protein that is highly expressed in the human placenta. In cancer tissues, this protein actively extrudes a wide variety of chemically and structurally unrelated chemotherapeutic drugs and other compounds. Studies in mice have shown that in the absence of BCRP activity in the placenta, there is a 2-fold increase in the uptake in BCRP substrates into fetus. This suggests that in the placenta, BCRP extrudes compounds that would otherwise cross the syncytiotrophoblast cells into fetal circulation. The purpose of this study was to examine the expression and localization of BCRP in the human placenta throughout gestation. Tissues from 6-13, 16-19, 24-29, 32-35, and 38-41 weeks of gestation were used. Real time RT-PCR analysis demonstrated that the mRNA levels of BCRP in the placenta do not change significantly as gestation progressed. However, Western blot analysis revealed that the protein levels increased towards the end of gestation. We demonstrated that BCRP is localized to the syncytiotrophoblast of the placenta and in some fetal blood vessels within the placenta. Tissues from the early stages of pregnancy (6-13 weeks) showed fewer BCRP positive blood vessels than term tissues (38-41 weeks).     

Structure of a self-splicing group II intron catalytic effector domain 5: parallels with spliceosomal U6 RNA

Domain 5 (D5) is absolutely required for all catalytic functions of group II introns. Here we describe the solution NMR structure, electrostatic calculations, and detailed magnesium ion-binding surface of D5 RNA from the Pylaiella littoralis large ribosomal RNA intron (D5-PL). The overall structure consists of a hairpin capped by a GNRA tetraloop. The stem is divided into lower and upper helices of 8 and 5 bp, respectively, separated by an internal bulge. The D5-PL internal bulge nucleotides stack into the helical junction, resulting in a coupling between the bulge A25 and the closing base pair (G8-C27) of the lower helix. Comparison of the D5-PL structure to previously reported related structures indicates that our structure is most similar, in the helical regions, to the crystal structure of D5 from yeast Ai5gamma (D5-Ai5gamma) and the NMR structure of the U6 snRNA stem-loop region. Our structure differs in many respects from both the NMR and X-ray structures of D5-Ai5gamma in the bulge region. Electrostatic calculations and NMR chemical shift perturbation analyses reveal magnesium ion-binding sites in the tetraloop, internal bulge, and the AGC triad in the lower stem. Our results suggest that the structure, electrostatic environment, and the magnesium ion-binding sites within the tetraloop, bulge, and triad regions are conserved features of the splicing machinery of both the group II introns and the spliceosome that are likely key for catalytic function.     

Dynamic flexibility of double-stranded RNA activated PKR in solution

PKR, an interferon-induced double-stranded RNA activated serine-threonine kinase, is a component of signal transduction pathways mediating cell growth control and responses to stress and viral infection. Analysis of separate PKR functional domains by NMR and X-ray crystallography has revealed details of PKR RNA binding domains and kinase domain, respectively. Here, we report the structural characteristics, calculated from biochemical and neutron scattering data, of a native PKR fraction with a high level of autophosphorylation and constitutive kinase activity. The experiments reveal association of the protein monomer into dimers and tetramers, in the absence of double-stranded RNA or other activators. Low-resolution structures of the association states were obtained from the large angle neutron scattering data and reveal the relative orientation of all protein domains in the activated kinase dimer. Low-resolution structures were also obtained for a PKR tetramer-monoclonal antibody complex. Taken together, this information leads to a new model for the structure of the functioning unit of the enzyme, highlights the flexibility of PKR and sheds light on the mechanism of PKR activation. The results of this study emphasize the usefulness of low-resolution structural studies in solution on large flexible multiple domain proteins.     

Genomic and epidemiological characteristics of SARS-CoV-2 in Africa

Since late 2019, the coronavirus disease 2019 (COVID-19) outbreak, caused by SARS-CoV-2, has rapidly evolved to become a global pandemic. Each country was affected but with a varying number of infected cases and mortality rates. Africa was hit late by the pandemic but the number of cases rose sharply. In this study, we investigated 224 SARS-CoV-2 genome sequences from the Global Initiative on Sharing Avian Influenza Data (GISAID) in the early part of the outbreak, of which 69 were from Africa. We analyzed a total of 550 mutations by comparing them with the reference SARS-CoV-2 sequence from Wuhan. We classified the mutations observed based on country and region, and afterwards analyzed common and unique mutations on the African continent as a whole. Correlation analyses showed that the duo variants ORF1ab/RdRp 4715L and S protein 614G variants, which are strongly linked to fatality rate, were not significantly and positively correlated with fatality rates (r = -0.03757, P = 0.5331 and r = -0.2876, P = 0.6389, respectively), although increased number of cases correlated with number of deaths (r = 0.997, P = 0.0002). Furthermore, most cases in Africa were mainly imported from American and European countries, except one isolate with no mutation and was similar to the original isolate from Wuhan. Moreover, unique mutations specific to countries were identified in the early phase of the outbreak but these mutations were not regional-specific. There were common mutations in all isolates across the continent as well as similar isolate-specific mutations in different regions. Our findings suggest that mutation is rapid in SARS-CoV-2 in Africa and although these mutations spread across the continent, the duo variants could not possibly be the sole cause of COVID-19 deaths in Africa in the early phase of the outbreak.     

Haematological and biochemical reference values for healthy adults in the middle belt of Ghana

Background

Reference values are very important in clinical management of patients, screening participants for enrolment into clinical trials and for monitoring the onset of adverse events during these trials. The aim of this was to establish gender-specific haematological and biochemical reference values for healthy adults in the central part of Ghana.

Methods

A total of 691 adults between 18 and 59 years resident in the Kintampo North Municipality and South District in the central part of Ghana were randomly selected using the Kintampo Health and Demographic Surveillance System and enrolled in this cross-sectional survey. Out of these, 625 adults made up of 316 males and 309 females were assessed by a clinician to be healthy. Median values and nonparametric 95% reference values for 16 haematology and 22 biochemistry parameters were determined for this population based on the Clinical Laboratory and Standards Institute guidelines. Values established in this study were compared with the Caucasian values being used currently by our laboratory as reference values and also with data from other African and western countries.

Results

Reference values established include: haemoglobin 113–164 g/L for males and 88–144 g/L for females; total white blood cell count 3.4–9.2×10⁹/L; platelet count 88–352×10⁹/L for males and 89–403×10⁹/L for females; alanine aminotransferase 8–54 U/L for males and 6–51 U/L for females; creatinine 56–119 µmol/L for males and 53–106 µmol/L for females. Using the haematological reference values based on the package inserts would have screened out up to 53% of potential trial participants and up to 25% of the population using the biochemical parameters.

Conclusion

We have established a panel of locally relevant reference parameters for commonly used haematological and biochemical tests. This is important as it will help in the interpretation of laboratory results both for clinical management of patients and safety monitoring during a trial.     

Asymmetry of 13C labeled 3-pyruvate affords improved site specific labeling of RNA for NMR spectroscopy

Selective isotopic labeling provides an unparalleled window within which to study the structure and dynamics of RNAs by high resolution NMR spectroscopy. Unlike commonly used carbon sources, the asymmetry of ¹³C-labeled pyruvate provides selective labeling in both the ribose and base moieties of nucleotides using Escherichia coli variants, that until now were not feasible. Here we show that an E. coli mutant strain that lacks succinate and malate dehydrogenases (DL323) and grown on [3-¹³C]-pyruvate affords ribonucleotides with site specific labeling at C5′ (~95%) and C1′ (~42%) and minimal enrichment elsewhere in the ribose ring. Enrichment is also achieved at purine C2 and C8 (~95%) and pyrimidine C5 (~100%) positions with minimal labeling at pyrimidine C6 and purine C5 positions. These labeling patterns contrast with those obtained with DL323 E. coli grown on [1, 3-¹³C]-glycerol for which the ribose ring is labeled in all but the C4′ carbon position, leading to multiplet splitting of the C1′, C2′ and C3′ carbon atoms. The usefulness of these labeling patterns is demonstrated with a 27-nt RNA fragment derived from the 30S ribosomal subunit. Removal of the strong magnetic coupling within the ribose and base leads to increased sensitivity, substantial simplification of NMR spectra, and more precise and accurate dynamic parameters derived from NMR relaxation measurements. Thus these new labels offer valuable probes for characterizing the structure and dynamics of RNA that were previously limited by the constraint of uniformly labeled nucleotides.     

Differential binding of 2'-biotinylated analogs of c-di-GMP with c-di-GMP riboswitches and binding proteins

C-di-GMP has emerged as a signalling molecule that regulates a variety of processes in several bacteria; therefore there is interest in the development of biotinylated analogs for the identification of binding partners. No detailed study has been done to evaluate if biotinylated analogs of c-di-GMP are capable of binding to c-di-GMP receptors. Herein, we evaluate the binding of commercially available 2'-biotinylated c-di-GMP and phosphorothioate 2'-biotinylated c-di-GMP, prepared via a facile solid-phase synthesis, to several c-di-GMP receptors. Docking, using Autodock vina software, as well as experimental studies of these analogs, with c-di-GMP class I and II riboswitches and binding proteins, reveal that some, but not all, c-di-GMP receptors can tolerate the 2'-modification of c-di-GMP with biotin.     

Psychological health and religious coping of Ghanaian women with infertility

Background

Infertility has been shown to have considerable psychological effects on the well-being of couples, especially women. Religion has been found as a resource used by infertile women to cope with their distress. Little research has examined the influence of religious coping on psychological distress among infertile women in Ghana. This study examines the relationship between positive and negative religious coping and psychological health for women with infertility problems in Ghana.

Methods

One hundred and fifty married women who were receiving assisted reproduction care in two specialized clinics were recruited for this study. Participants were administered with the Brief Symptom Inventory and Brief Religious Coping Scale to assess psychological health associated with infertility and religious coping respectively. A hierarchical regression was performed to examine the relative contribution of the domains of psychological health (i.e. somatization, anxiety and depression) in predicting negative religious coping and positive religious.

Results

The results showed that negative religious coping was significant and positively correlated with somatization, depression and anxiety. Furthermore, a positive relationship also existed between positive religious coping and somatization and anxiety but not depression. After controlling for age and duration of infertility, somatization and anxiety predicted positive religious coping whilst all the domains of psychological health (somatization, anxiety and depression) precited negative religious coping.

Conclusions

This study expanded on the existing literature by examining positive and negative religious coping with psychological distress associated with infertility for women. These results underscore the need for health professionals providing therapies for women with infertility to acknowledge and consider their religious beliefs as this influences their mental health.