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Low-density lipoprotein receptor-related protein 6 (LRP6) is a coreceptor of the β-catenin-dependent Wnt signaling pathway. The LRP6 ectodomain binds Wnt proteins, as well as Wnt inhibitors such as sclerostin (SOST), which negatively regulates Wnt signaling in osteocytes. Although LRP6 ectodomain 1 (E1) is known to interact with SOST, several unresolved questions remain, such as the reason why SOST binds to LRP6 E1E2 with higher affinity than to the E1 domain alone. Here, we present the crystal structure of the LRP6 E1E2–SOST complex with two interaction sites in tandem. The unexpected additional binding site was identified between the C-terminus of SOST and the LRP6 E2 domain. This interaction was confirmed by in vitro binding and cell-based signaling assays. Its functional significance was further demonstrated in vivo using Xenopus laevis embryos. Our results provide insights into the inhibitory mechanism of SOST on Wnt signaling.

Wnt signaling is initiated by Wnt ligand binding to the extracellular ligand binding domain, called the cysteine-rich domain (CRD), of a Frizzled (Fzd) receptor. Norrin, an atypical Fzd ligand, specifically interacts with Fzd4 to activate β-catenin-dependent canonical Wnt signaling. Much of the molecular basis that confers Norrin selectivity in binding to Fzd4 was revealed through the structural study of the Fzd4-Norrin complex. However, how the ligand interaction, seemingly localized at the CRD, is transmitted across full-length Fzd4 to the cytoplasm remains largely unknown. Here, we show that a flexible linker domain, which connects the CRD to the transmembrane domain, plays an important role in Norrin signaling. The linker domain directly contributes to the high-affinity interaction between Fzd4 and Norrin as shown by ∼10-fold higher binding affinity of Fzd4 to Norrin in the presence of the linker. Swapping the Fzd4 linker with the Fzd5 linker resulted in the loss of Norrin signaling, suggesting the importance of the linker in ligand-specific cellular response. In addition, structural dynamics of Fzd4 associated with Norrin binding investigated by hydrogen/deuterium exchange MS revealed Norrin-induced conformational changes on the linker domain and the intracellular loop 3 (ICL3) region of Fzd4. Cell-based functional assays showed that linker deletion, L430A and L433A mutations at ICL3, and C-terminal tail truncation displayed reduced β-catenin-dependent signaling activity, indicating the functional significance of these sites. Together, our results provide functional and biochemical dissection of Fzd4 in Norrin signaling.

Neuropeptide Y (NPY) is highly abundant in the brain and involved in various physiological processes related to food intake and anxiety, as well as human diseases such as obesity and cancer. However, the molecular details of the interactions between NPY and its receptors are poorly understood. Here, we report a cryo-electron microscopy structure of the NPY-bound neuropeptide Y1 receptor (Y1R) in complex with Gi1 protein. The NPY C-terminal segment forming the extended conformation binds deep into the Y1R transmembrane core, where the amidated C-terminal residue Y36 of NPY is located at the base of the ligand-binding pocket. Furthermore, the helical region and two N-terminal residues of NPY interact with Y1R extracellular loops, contributing to the high affinity of NPY for Y1R. The structural analysis of NPY-bound Y1R and mutagenesis studies provide molecular insights into the activation mechanism of Y1R upon NPY binding.

BACKGROUND CONTEXT. Proximal junctional kyphosis (PJK) following surgical treatment of lumbar degenerative kyphosis (LDK) is one of the critical complications leading to the failure of instrumentation and additional extensive surgery. However, most previous studies have focused on idiopathic scoliosis resulting from variable surgical techniques. LDK usually differ from other scoliotic deformities in terms of patient characteristics and disease mechanisms. PURPOSE. Identification of the prevalence of PJK after the surgical treatment of LDK and searching for the predictable value for the progression of PJK. Study design. Retrospective comparative study. Patient sample (must be included in clinical studies). Forty-seven consecutive patients who underwent surgical correction of a sagittal imbalance due to LDK, from January 2005 to December 2008 in a single spine clinic, were evaluated with a minimum 2 years follow-up (mean 3.8 years). METHODS. Patients were divided into 2 groups: with or without the occurrence of PJK, and three categorized factors according to patient characteristics, surgical variables, and the radiographic spinopelvic parameters were evaluated. RESULTS. PJK had occurred in 29 of 47 patients (61.7%). Among variable factors, old age, upper-instrumented vertebra below L2, lumbar lordosis to PI ratio, and the sum of lumbar lordosis, and the sacral slope related to PI were found to be statistically significant. CONCLUSIONS. The overall incidence of PJK following surgical treatment of LDK patients was higher than expected. Spinal biomechanics may be changed after long instrumented fusion surgery. Thorough consideration of these factors is needed in the treatment strategy of LDK patients. A long-term follow-up study should be conducted.

GPT-3 shows remarkable in-context learning ability of large-scale language models (LMs) trained on hundreds of billion scale data. Here we address some remaining issues less reported by the GPT-3 paper, such as a non-English LM, the performances of different sized models, and the effect of recently introduced prompt optimization on in-context learning. To achieve this, we introduce Hy-perCLOVA, a Korean variant of 82B GPT-3 trained on a Korean-centric corpus of 560B tokens. Enhanced by our Korean-specific tokenization, HyperCLOVA with our training configuration shows state-of-the-art in-context zero-shot and few-shot learning performances on various downstream tasks in Korean. Also, we show the performance benefits of promptbased learning and demonstrate how it can be integrated into the prompt engineering pipeline. Then we discuss the possibility of materializing the No Code AI paradigm by providing AI prototyping capabilities to nonexperts of ML by introducing HyperCLOVA studio, an interactive prompt engineering interface. Lastly, we demonstrate the potential of our methods with three successful in-house applications.

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