154Abstract Mendelian randomization (MR) is a widely-used method for causal inference using genetic data.Mendelian randomization studies of unrelated individuals may be susceptible to bias from family structure, for example, through dynastic effects which occur when parental genotypes directly affect offspring phenotypes. Here we describe methods for within-family Mendelian randomization and through simulations show that family-based methods can overcome bias due to dynastic effects. We illustrate these issues empirically using data from 61,008 siblings from the UK Biobank and Nord-Trøndelag Health Study. Both within-family and populationbased Mendelian randomization analyses reproduced established effects of lower BMI reducing risk of diabetes and high blood pressure. However, while MR estimates from population-based samples of unrelated individuals suggested that taller height and lower BMI increase educational attainment, these effects largely disappeared in within-family MR analyses. We found differences between population-based and within-family based estimates, indicating the importance of controlling for family effects and population structure in Mendelian randomization studies.

Summary Proteins possessing the SPX domain are found in several proteins involved in inorganic phosphate (Pi) transport and signalling in yeast and plants. Although the functions of several SPX‐domain protein subfamilies have recently been uncovered, the role of the SPX‐MFS subfamily is still unclear. Using quantitative RT‐PCR analysis, we studied the regulation of SPX‐MFS gene expression by the central regulator, OsPHR2 and Pi starvation. The function of OsSPX‐MFS1 in Pi homeostasis was analysed using an OsSPX‐MFS1 mutant (mfs1) and osa‐miR827 overexpression line (miR827‐Oe). Finally, heterologous complementation of a yeast mutant impaired in Pi transporter was used to assess the capacity of OsSPX‐MFS1 to transport Pi. Transcript analyses revealed that members of the SPX‐MFS family were mainly expressed in the shoots, with OsSPX‐MFS1 and OsSPX‐MFS3 being suppressed by Pi deficiency, while OsSPX‐MFS2 was induced. Mutation in OsSPX‐MFS1 (mfs1) and overexpression of the upstream miR827 (miR827‐Oe) plants impaired Pi homeostasis in the leaves. In addition, studies in yeast revealed that OsSPX‐MFS1 may be involved in Pi transport. The results suggest that OsSPX‐MFS1 is a key player in maintaining Pi homeostasis in the leaves, potentially acting as a Pi transporter.

As the prototypical member of the PTP family, protein tyrosine phosphatase 1B (PTP1B) is an attractive target for therapeutic interventions in type 2 diabetes. The extremely conserved catalytic site of PTP1B renders the design of selective PTP1B inhibitors intractable. Although discovered allosteric inhibitors containing a benzofuran sulfonamide scaffold offer fascinating opportunities to overcome selectivity issues, the allosteric inhibitory mechanism of PTP1B has remained elusive. Here, molecular dynamics (MD) simulations, coupled with a dynamic weighted community analysis, were performed to unveil the potential allosteric signal propagation pathway from the allosteric site to the catalytic site in PTP1B. This result revealed that the allosteric inhibitor compound-3 induces a conformational rearrangement in helix α7, disrupting the triangular interaction among helix α7, helix α3, and loop11. Helix α7 then produces a force, pulling helix α3 outward, and promotes Ser190 to interact with Tyr176. As a result, the deviation of Tyr176 abrogates the hydrophobic interactions with Trp179 and leads to the downward movement of the WPD loop, which forms an H-bond between Asp181 and Glu115. The formation of this H-bond constrains the WPD loop to its open conformation and thus inactivates PTP1B. The discovery of this allosteric mechanism provides an overall view of the regulation of PTP1B, which is an important insight for the design of potent allosteric PTP1B inhibitors.

In this work, a simple and highly selective colorimetric method has been developed for quantifying trace-level ATP using Fe3O4 nanoparticles (Fe3O4 NPs). It was discovered that Fe3O4 NPs could present the dramatically enhanced catalysis once anchored with ATP-specific aptamers (Apts), which is about 6-fold larger than that of bare Fe3O4 NPs. In the presence of ATP, however, the Apts would be desorbed from Fe3O4 NPs due to the Apts-target binding event, leading to the decrease of catalysis rationally depending on ATP concentrations. A colorimetric strategy was thereby developed to facilitate the highly selective detection of ATP, showing the linear concentrations ranging from 0.50 to 100 μM. Subsequently, the developed ATP sensor was employed for the evaluation of ATP in blood with the analysis performances comparably better than those of the documented detection methods, showing the potential applications in the clinical laboratory for the detective diagnosis of some ATP-indicative diseases. Importantly, such a catalysis-based detection strategy should be extended to other kinds of nanozymes with intrinsic catalysis properties (i.e., peroxidase and oxidase-like activities), promising as a universal candidate for monitoring various biological species simply by using target-specific recognition elements like Apts and antibodies.

Stem growth habit is a key plant architecture trait determining yield potential in grain legumes, and the phenotypic change from the indeterminate stem growth habit of wild mungbeans (Vigna radiata) to the determinate stem growth habit of cultivated mungbeans is a critical domestication transition. Here we show that indeterminate stem growth in wild mungbean is modulated by a single gene, VrDet1, which encodes a signaling protein of shoot apical meristems. The transition from an indeterminate to a determinate stem growth habit was achieved by selection of two linked point mutations in two putative cis-regulatory elements, resulting in a significant reduction in gene expression. Both the wild-type nucleotides corresponding to the two point mutations were essential for VrDet1 function. In addition, two highly diverse haplotypes of Vrdet1 were found in cultivated mungbeans, suggesting dual domestication of Vrdet1. VrDet1 was orthologous to Dt1 in wild soybean and PvTFL1y in wild common bean, where multiple loss-of-function mutations altering the coding sequences of individual genes were selected to produce determinate stems in cultivated accessions. Interspecific comparison of these orthologs in the wild and cultivated accessions reveals the most conservative interspecific and intraspecific parallel domestication events with the broadest mutational spectrum of a domestication trait in leguminous crops. We also found that interspecifically and functionally conserved promoters possess cis-regulatory elements that are highly conserved in kind but greatly variable in number and order, demonstrating the evolutionary dynamics of regulatory sequences. This work provides insights into the origins of cultivated mungbean and exemplifies the conservativeness and plasticity of the domestication processes of related crops.

Heat shock transcription factors (Hsfs) are essential elements in plant signal transduction pathways that mediate gene expression in response to various abiotic stresses. Mungbean (Vigna radiata) is an important crop worldwide. The emergence of a genome database now allows for functional analysis of mungbean genes. In this study, we dissect the mungbean Hsfs using genome-wide identification and expression profiles. We characterized a total of 24 VrHsf genes and classified them into three groups (A, B, and C) based on their phylogeny and conserved domain structures. All VrHsf genes exhibit highly conserved exon-intron organization, with two exons and one intron. In addition, all VrHsf proteins contain 16 distinct motifs. Chromosome location analysis revealed that VrHsf genes are located on 8 of the 11 mungbean chromosomes, and that seven duplicated gene pairs had formed among them. Moreover, transcription patterns of VrHsf genes varied in different tissues, indicating their different roles in plant growth and development. We identified multiple stress related cis-elements in VrHsf promoter regions 2 kb upstream of the translation initiation codons, and the expression of most VrHsf genes was altered under different stress conditions, suggesting their potential functions in stress resistance pathways. These molecular characterization and expression profile analyses of VrHsf genes provide essential information for further function investigation.

Plants are often confronted to nutrient limiting conditions, such as inorganic phosphate (Pi) deficiency, resulting in a reduction in growth and yield. PHO2, encoding a ubiquitin-conjugating E2 enzyme, is a central component of the Pi-starvation response signalling pathway. A yeast-two-hybrid screen using Oryza sativa (rice) PHO2 as bait, revealed an interaction between OsPHO2 and OsGIGANTEA, a key regulator of flowering time, which was confirmed using bimolecular fluorescence complementation (BiFC). Characterization of rice Osgi and Ospho2 mutants revealed that they displayed several similar phenotypic features supporting a physiological role for this interaction. Reduced growth, leaf tip necrosis, delayed flowering and over-accumulation of Pi in leaves compared to wild type were shared features of Osgi and Ospho2 plants. Pi analysis of individual leaves demonstrated that Osgi, similar to Ospho2 mutants, were impaired in Pi remobilization from old to young leaves, albeit to a lesser extent. Transcriptome analyses revealed more than 55% of the genes differentially expressed in Osgi plants overlapped with the set of differentially expressed genes in Ospho2 plants. The interaction between OsPHO2 and OsGI links high-level regulators of Pi homeostasis and development in rice.

BackgroundPhosphatidyl ethanolamine-binding proteins (PEBPs) are involved in the regulation of plant architecture and flowering time. The functions of PEBP genes have been studied in many plant species. However, little is known about the characteristics and expression profiles of PEBP genes in wild peanut species, Arachis duranensis and Arachis ipaensis, the diploid ancestors of cultivated peanuts.ResultsIn this study, genome-wide identification methods were used to identify and characterize a total of 32 peanut PEBP genes, 16 from each of the two wild peanut species, A. duranensis and A. ipaensis. These PEBP genes were classified into 3 groups (TERMINAL FLOWER1-like, FLOWERING LOCUS T-like, and MOTHER OF FT AND TFL1-like) based on their phylogenetic relationships. The gene structures, motifs, and chromosomal locations for each of these PEBPs were analyzed. In addition, 4 interchromosomal duplications and 1 tandem duplication were identified in A. duranensis, and 2 interchromosomal paralogs and 1 tandem paralog were identified in A. ipaensis. Ninety-five different cis-acting elements were identified in the PEBP gene promoter regions and most genes had different numbers and types of cis-elements. As a result, the transcription patterns of these PEBP genes varied in different tissues and under long day and short day conditions during different growth phases, indicating the functional diversities of PEBPs in different tissues and their potential functions in plant photoperiod dependent developmental pathways. Moreover, our analysis revealed that AraduF950M/AraduWY2NX in A. duranensis, and Araip344D4/Araip4V81G in A. ipaensis are good candidates for regulating plant architecture, and that Aradu80YRY, AraduYY72S, and AraduEHZ9Y in A. duranensis and AraipVEP8T in A. ipaensis may be key factors regulating flowering time.ConclusionSixteen PEBP genes were identified and characterized from each of the two diploid wild peanut genomes, A. duranensis and A. ipaensis. Genetic characterization and spatio-temporal expression analysis support their importance in plant growth and development. These findings further our understanding of PEBP gene functions in plant species.