Molecules that facilitate targeted protein degradation (TPD) offer great promise as novel therapeutics. Human hepatic lectin, asialoglycoprotein receptor (ASGR) is selectively expressed on hepatocytes. We have previously engineered an anti-ASGR1 antibody-mutant RSPO2 (RSPO2RA) fusion protein (called SWEETS) to drive tissue-specific degradation of ZNRF3/RNF43 E3-ubiquitin ligases, leading to hepatocyte specific enhanced Wnt signaling, proliferation, and restored liver function in mouse models. Such an antibody-RSPO2RA fusion molecule is currently in human clinical trials. In the current study, we identified two new ASGR1 and ASGR1/2 specific antibodies, 8M24 and 8G8. High-resolution crystal structures of ASGR1:8M24 and ASGR2:8G8 complexes revealed that these antibodies bind to distinct epitopes on the opposite sides of ASGR, away from the substrate binding site. Both antibodies enhanced Wnt-activity when assembled as SWEETS molecules with RSPO2RA through specific effects sequestering E3 ligases. In addition, 8M24-RSPO2RA and 8G8-RSPO2RA efficiently downregulated ASGR1 through TPD mechanisms. These results demonstrate the possibility of combining different therapeutic effects and different degradation mechanisms in a single molecule. less

Slow-controlled release fertilizers are experiencing a popularity in rice cultivation due to their effectiveness in yield and quality with low environmental costs. However, the underlying mechanism by which these fertilizers regulate grain quality remains inadequately understood. This study investigated the effects of five fertilizer management practices on rice quality in a two-year field experiment: CK, conventional fertilization, and four applications of slow-controlled release fertilizer (UF, urea formaldehyde; SCU, sulfur-coated urea; PCU, polymer-coated urea; BBF, controlled-release bulk blending fertilizer). In 2020 and 2021, the yields of UF and SCU groups showed significant decreases when compared to conventional fertilization, accompanied by a decline in nutritional quality. Additionally, PCU group exhibited poorer cooking and eating qualities. However, BBF group achieved increases in both yield (10.8 t hm-2 and 11.0 t hm-2) and grain quality reaching the level of CK group. The sufficient nitrogen supply in both the PCU and BBF groups during the grain-filling stage led to a greater capacity for the accumulation of proteins and amino acids in the PCU group compared to starch accumulation. Intriguingly, BBF group showed better carbon-nitrogen metabolism than that of PCU group. The optimal nitrogen supply present in BBF group suitable boosted the synthesis of amino acids involved in the glycolysis/ tricarboxylic acid cycle, thereby effectively coordinating carbon-nitrogen metabolism. The application of the new slow-controlled release fertilizer, BBF, is advantageous in regulating the carbon flow in the carbon-nitrogen metabolism to enhance rice quality. less

As in origami, morphogenesis in living systems heavily relies on tissue curving and folding, through the interplay between biochemical and biomechanical cues. In contrast, certain organs maintain their flat posture over several days. Here we identified a pathway, which is required for the maintenance of organ flatness, taking the sepal, the outermost floral organ, in Arabidopsis as a model system. Through genetic, cellular and mechanical approaches, our results demonstrate that global gene expression regulator VERNALIZATION INDEPENDENCE 4 (VIP4) fine-tunes the mechanical properties of sepal cell walls and maintains balanced growth on both sides of the sepals, mainly by orchestrating the distribution pattern of AUXIN RESPONSE FACTOR 3 (ARF3). vip4 mutation results in softer cell walls and faster cell growth on the adaxial sepal side, which eventually cause sepals to bend outward. Downstream of VIP4, ARF3 works through modulating auxin signaling to down-regulate pectin methylesterase VANGUARD1, resulting in decreased cell wall stiffness. Our work unravels a 3-component module, which relates hormonal patterns to organ curvature, and actively maintains sepal flatness during its growth. less

Cassava is a crucial staple crop for smallholder farmers in tropical Asia and Sub-Saharan Africa. Although high yield remains the top priority for farmers, the significance of nutritional values has increased in cassava breeding programs. A notable negative correlation between provitamin A and starch accumulation poses a significant challenge for breeding efforts. The negative correlation between starch and carotenoid levels in conventional and genetically modified cassava plants implies the absence of a direct genomic connection between the two traits. The competition among various carbon pathways seems to account for this relationship. In this study, we conducted a thorough analysis of 49 African cassava genotypes with varying levels of starch and provitamin A. Our goal was to identify factors contributing to differential starch accumulation. With the carotenoid levels of the varieties considered as a confounding effect on starch production, we found that yellow and white-fleshed storage roots did not differ significantly in most measured components of starch or de novo fatty acid biosynthesis. However, genes and metabolites associated with myo-inositol synthesis and cell wall component production were substantially enriched in high provitamin A genotypes. These results indicate that yellow-fleshed cultivars, in comparison to their white-fleshed counterparts, direct more carbon towards the synthesis of raffinose and cell wall components, a finding that is supported by a significant rise in the starch-free residue to total dry yield ratio in yellow storage roots versus white storage roots. Our findings enhance comprehension of the biosynthesis of starch and carotenoids in the storage roots of cassava. less

Haplotype-based breeding approaches hold promise for enhancing crop improvement strategies, allowing for targeted selection of superior genetic combinations to develop high-yielding and resilient varieties. The current study aimed at identification of NRT1.1 nitrate transporter haplotype that could serve as \"donors\" in haplotype-based breeding. We phenotyped 272 rice accessions in hydroponics with sufficient and low nitrogen (N) for nitrate uptake efficiency. By employing principal component and hierarchical cluster analysis, the accessions were grouped into N efficient, intermediate, and inefficient clusters. Haplotype analysis unveiled the presence of two haplogroups for OsNRT1.1A, three for OsNRT1.1B, and five for OsNRT1.1C. Through haplo-pheno association, the comparison of mean trait values revealed H2 and H3 as the superior haplotypes (SH) for OsNRT1.1A and OsNRT1.1B, respectively. In the case of OsNRT1.1C, H3 and H1 emerged as SH within the N-efficient cluster. Conversely, the inferior haplotypes (IH) consisted of H1 in OsNRT1.1A, H3 in OsNRT1.1B, and H3 and H2 in OsNRT1.1C within the N-inefficient cluster. However, relative expression of OsNRT1.1 (with specific paralogs) in contrasting rice accessions revealed that a few of the inferior accession exhibited higher expression levels in the root but lower in the shoot, which might have contributed to their N-inefficiency. Furthermore, amino acid change at position 403 (Isoleucine to Valine) in inferior accessions influences the active site OsNRT1.1C protein causing N-inefficiency. Ours is the first report on haplotype analysis of NRT1.1 gene demonstrating its genetic diversity, as well as its association with phenotype will have potential implications for improving nitrate uptake efficiency. less

The importance of auxin in plant organ development including root nodule formation is well established. Using auxin reporter constructs the spatiotemporal auxin distribution pattern during nodule development has previously been illustrated. However, our understanding of how this pattern is built-up and maintained still remains elusive. To this end, we studied how the auxin gradient visualized by DR5 expression patterns at different stages of nodule development in Medicago truncatula (Medicago), is correlated with the spatiotemporal expression patterns of known auxin biosynthesis and auxin transport genes. In addition, we record the MtPIN10-GFP expression pattern and polar positioning on the cell plasma membranes during nodule primordium development to investigate the auxin flux. RNA interference and the application of auxin synthesis blockers were used to demonstrate the relevance of biosynthesis and transport at the initial stages of the nodulation process. Our results show that upon rhizobium inoculation, preceding the first mitotic activity, a specific set of MtYUCs and MtPINs as well as MtLAX2 are expressed in the pericycle contributing to the creation of an auxin maximum. Overall, we demonstrate that dynamic spatiotemporal expression of both, MtYUCs and MtPINs, result in specific auxin outputs in subsequent stages of nodule primordia and nodule meristem formation. less

Calorie restriction (CR) is a dietary intervention to promote health and longevity. CR causes various metabolic changes in both the production and circulation of metabolites; however, it remains unclear which of the changed metabolite(s) can account for the physiological benefits of CR. Through metabolomic analysis of metabolites undergoing abundance changes during CR and subsequent functional validation, we found that lithocholic acid (LCA) is the only metabolite that alone can recapitulate the effects of CR, including activation of AMPK and the rejuvenating effects of muscle regeneration, grip strength and running capacity in mice. Interestingly, LCA also activates AMPK and exerts life- and health-extending effects in Caenorhabditis elegans and Drosophila melanogaster, indicating that these animal models are able to transmit the signalling of LCA once administered. Knockout of AMPK abrogates LCA-induced phenotypes, in nematodes and flies, as well as in mice. Together, we have identified that administration of the CR-upregulated metabolite LCA alone can confer anti-ageing benefits to metazoans, in an AMPK-dependent manner. less

Plants have evolved to anticipate and adjust their growth and development in response to environmental changes. To mitigate the negative influence of global climate change on crop production, understanding the key regulators of plant performance is imperative. EARLY FLOWERING 3 (ELF3) is such a regulator involved in the circadian clock and thermomorphogenesis. Arabidopsis thaliana ELF3 contains a prion-like domain (PrD) that functions as a thermosensor, enabling its liquid-liquid phase separation at high ambient temperatures. To understand the conservation of this function across the plant kingdom, we traced the evolutionary emergence of ELF3 with a focus on the PrD, which confers liquid-liquid phase separation. We observed that the presence of the domain within ELF3, mainly contributed by the length of polyglutamine (polyQ) repeats, is largely restricted to Brassicales. This suggests that thermosensory function of ELF3 is a rather recent and secondary acquirement that was added to its main function. By analyzing 319 natural Arabidopsis thaliana accessions, we detected a wide range of polyQ length variation in ELF3. However, polyQ length is only weakly associated with geographic origin, climate conditions, and classic temperature-responsive phenotypes. Consequently, we conclude that although the emergence of PrD is not likely to be a key driver of environmental adaptation, it adds an extra layer to the role of ELF3 in thermomorphogenesis. less

UV RESISTANCE LOCUS 8 (UVR8) has been identified in Arabidopsis thaliana as the receptor for UV-B radiation mediating photomorphogenic responses and acclimation to UV-B radiation. However, UVR8-mediated UV-B signaling pathways in rice, that has two proteins (UVR8a and UVR8b) with homology to AtUVR8, remain largely unknown. In this study, UVR8a and UVR8b were found to be expressed mainly in rice leaves and leaf sheaths, while the level of UVR8b was higher than that of UVR8a. In agreement with prior studies on AtUVR8, uvr8b and uvr8a uvr8b rice mutants exposed to UV-B showed reduced UVB-induced growth inhibition and upregulation of CHS and HY5 transcripts along with acclimation to UV-B, overexpressing UVR8a or UVR8b enhanced UV-B-induced growth inhibition and acclimation to UV-B, compared to wild-type plants. UV-B was able to enhance the interaction between CONSTITUTIVE PHOTOMORPHOGENESIS1 (COP1) with UVR8a/UVR8b, whereas the interaction intensity of REPRESSOR OF UV-B PHOTOMORPHOGENESIS2 (RUP2) with UVR8a was significantly higher than that with UVR8b. In addition, UVR8a and UVR8b were also found in the nucleus and cytoplasm, but OsUVR8 proteins were localized in nucleus in the absence of UV-B. The level of OsUVR8 monomer showed an invisible change in the leaves of rice seedlings transferred from white light to white light supplemented with UV-B?even UV-B can weaken the interactions of UVR8a or/and UVR8b. Therefore, both UVR8a and UVR8b, that have different location and response modes with Arabidopsis UVR8, function in the response of rice to UV-B radiation, whereas UVR8b plays a predominant role in this process. less

Evolutionary physiologists have long been interested in physiological mechanisms underpinning variation in life-history performance. Recent efforts to elucidate these mechanisms focused on bioenergetics and oxidative stress. One underappreciated area that could play a role in mediating variation in performance is the unfolded protein response (UPR), a cellular stress response that reduces secretory protein load, enhances endoplasmic reticulum (ER) protein folding and clearance capacity during stress and during its adaptive phase. Given that the ER and mitochondria interact to regulate cellular homeostasis, it seems intuitive that UPR phenotype would correlate strongly with mitochondrial physiology, which in turn would contribute to variations in whole-organism metabolism. One way researchers have been studying cellular controls of life-history traits is by assessing stress resistance and bioenergetic properties of primary dermal fibroblasts. However, it is unclear if findings from dermal fibroblasts can be generalized to other cell and tissue types, and if fibroblasts\' phenotypes are repeatable across different life-history stages. This study aimed to explore the relationships between UPR profile, cellular respiration, and stress resistance using primary dermal fibroblasts isolated at puberty and primary lung fibroblasts isolated at adulthood. Specifically, we tested if 1) UPR profile of dermal fibroblasts isolated at puberty corresponds to UPR profile of lung fibroblasts isolated at adulthood, 2) UPR profile of dermal fibroblasts isolated at puberty and lung fibroblasts isolated at adulthood correspond to cellular bioenergetics of lung fibroblasts isolated at adulthood, and 3) UPR profile of dermal fibroblasts isolated at puberty corresponds to multiplex stress resistance (ER stress, oxidative stress, DNA damage) of lung fibroblasts isolated at adulthood. We found that only tunicamycin induced BiP expression was repeatable in skin and lung fibroblasts. Tunicamycin induced expressions of BiP, GRP94, and CNX in skin fibroblasts predicted resistance of lung fibroblasts to tunicamycin, (but not thapsigargin and other inducers of lethal stress), which is indicative for the pro-survival role of UPR during stress. Tunicamycin induced BiP expression in skin and lung fibroblasts also predicted multiple cellular bioenergetics parameters in lung fibroblasts. less