Promoting tau secretion and propagation by hyperactive p300/CBP via autophagy-lysosomal pathway in tauopathy.

BackgroundThe trans-neuronal propagation of tau has been implicated in the progression of tau-mediated neurodegeneration. There is critical knowledge gap in understanding how tau is released and transmitted, and how that is dysregulated in diseases. Previously, we reported that lysine acetyltransferase p300/CBP acetylates tau and regulates its degradation and toxicity. However, whether p300/CBP is involved in regulation of tau secretion and propagation is unknown.MethodWe investigated the relationship between p300/CBP activity, the autophagy-lysosomal pathway (ALP) and tau secretion in mouse models of tauopathy and in cultured rodent and human neurons. Through a high-through-put compound screen, we identified a new p300 inhibitor that promotes autophagic flux and reduces tau secretion. Using fibril-induced tau spreading models in vitro and in vivo, we examined how p300/CBP regulates tau propagation.ResultsIncreased p300/CBP activity was associated with aberrant accumulation of ALP markers in a tau transgenic mouse model. p300/CBP hyperactivation blocked autophagic flux and increased tau secretion in neurons. Conversely, inhibiting p300/CBP promoted autophagic flux, reduced tau secretion, and reduced tau propagation in fibril-induced tau spreading models in vitro and in vivo.ConclusionsWe report that p300/CBP, a lysine acetyltransferase aberrantly activated in tauopathies, causes impairment in ALP, leading to excess tau secretion. This effect, together with increased intracellular tau accumulation, contributes to enhanced spreading of tau. Our findings suggest that inhibition of p300/CBP as a novel approach to correct ALP dysfunction and block disease progression in tauopathy

Small-Molecule Inhibitors of Protein-Protein Interactions: Progressing toward the Reality

The past 20 years have seen many advances in our understanding of protein-protein interactions (PPIs) and how to target them with small-molecule therapeutics. In 2004, we reviewed some early successes; since then, potent inhibitors have been developed for diverse protein complexes, and compounds are now in clinical trials for six targets. Surprisingly, many of these PPI clinical candidates have efficiency metrics typical of "lead-like" or "drug-like" molecules and are orally available. Successful discovery efforts have integrated multiple disciplines and make use of all the modern tools of target-based discovery-structure, computation, screening, and biomarkers. PPIs become progressively more challenging as the interfaces become more complex, i.e., as binding epitopes are displayed on primary, secondary, or tertiary structures. Here, we review the last 10 years of progress, focusing on the properties of PPI inhibitors that have advanced to clinical trials and prospects for the future of PPI drug discovery

LC3B Binds to the Autophagy Protease ATG4b with High Affinity Using a Bipartite Interface

Autophagy is a catabolic cellular process in which unwanted proteins and organelles are degraded by lysosomes. It is characterized by the formation of the double-membrane autophagosome decorated with LC3B, a protein that mediates autophagosomal fusion with lysosomes. The cysteine protease ATG4b acts at two stages in the life cycle of LC3B. We set out to characterize the protein-protein interaction between LC3B and ATG4b. Through biochemical and biophysical studies, we show that the ubiquitin-like core of LC3B (residues 1-115; "LC3B-115"), which lacks the C-terminal cleavage site (between residue 120 and 121), binds to full-length ATG4b with a surprisingly tight dissociation constant (KD) in the low nanomolar range; 10-30-fold tighter than that of the substrate pro-LC3B (residues 1-125) or the product LC3B-I (residues 1-120). Consequently, LC3B-115 is a potent inhibitor of the ATG4b-mediated cleavage of pro-LC3B (IC50 = 15 nM). Binding of the LC3B-115 has no effect on the conformation of the active site of ATG4b, as judged by the turnover of a peptide substrate ("substrate-33"), derived from LC3B-I residues 116-120. Conversely, truncations of ATG4b show that binding and proteolysis of LC3B critically depend on the C-terminal tail of ATG4b, whereas proteolysis of the peptide substrate-33 does not require the C-terminal tail of ATG4b. These results support a bipartite model for LC3B-ATG4b binding in which the core of LC3B binds to ATG4b and the C-terminal tail of pro-LC3B organizes the ATG4b active site; additionally, the C-terminal tail of ATG4b contributes at least 1000-fold higher binding affinity to the LC3B-ATG4b interaction and likely wraps around the LC3B-ubiquitin core. PPIs are often described as containing an energetic "hot spot" for binding; in the case of LC3B-ATG4b, however, the substrate-enzyme complex contains multiple, energetically relevant domains that differentially affect binding affinity and catalytic efficiency

The 2.7-Å crystal structure of a 194-kDa homodimeric fragment of the 6-deoxyerythronolide B synthase

The x-ray crystal structure of a 194-kDa fragment from module 5 of the 6-deoxyerythronolide B synthase has been solved at 2.7 Å resolution. Each subunit of the homodimeric protein contains a full-length ketosynthase (KS) and acyl transferase (AT) domain as well as three flanking “linkers.” The linkers are structurally well defined and contribute extensively to intersubunit or interdomain interactions, frequently by means of multiple highly conserved residues. The crystal structure also reveals that the active site residue Cys-199 of the KS domain is separated from the active site residue Ser-642 of the AT domain by ≈80 Å. This distance is too large to be covered simply by alternative positioning of a statically anchored, fully extended phosphopantetheine arm of the acyl carrier protein domain from module 5. Thus, substantial domain reorganization appears necessary for the acyl carrier protein to interact successively with both the AT and the KS domains of this prototypical polyketide synthase module. The 2.7-Å KS–AT structure is fully consistent with a recently reported lower resolution, 4.5-Å model of fatty acid synthase stucture, and emphasizes the close biochemical and structural similarity between polyketide synthase and fatty acid synthase enzymology

Lesion based diagnostic performance of dual phase 99mTc-MIBI SPECT/CT imaging and ultrasonography in patients with secondary hyperparathyroidism

Abstract Background We aimed to evaluate the diagnostic performance of 99mTc-MIBI SPECT/CT and ultrasonography in patients with secondary hyperparathyroidism (SHPT), and explored the factors that affect the diagnostic performance. Methods 99mTc-MIBI SPECT/CT and ultrasonography were performed in 50 patients with SHPT within 1 month before they underwent surgery. Imaging results were confirmed by the pathology. Pearson correlation analysis was used to determine the correlation of PTH level with clinical data. The optimal cutoff value for predicting positive 99mTc-MIBI results was evaluated by ROC analysis in lesions diameter. Results Forty-nine patients had a positive 99mTc-MIBI imaging results and 39 patients had positive ultrasonography results. The sensitivities of 99mTc-MIBI and ultrasonography were 98.00% and 78.00%, respectively. A total of 199 lesions were resected in 50 patients. Among them, 183 lesions were proved to be parathyroid hyperplasia. On per-lesion basis analysis, the sensitivity and specificity of 99mTc-MIBI and ultrasonography were 59.34% and 75.00% vs 46.24% and 80.00%, respectively. The Pearson correlation analysis showed that the serum AKP and PTH level had a significant linear association (r = 0.699, P < 0.001). The lesion diameter was a statistically significant predictive factor in predicting positive 99mTc-MIBI SPECT/CT. The optimal cutoff value for predicting positive 99mTc-MIBI results evaluated by ROC analysis in lesions diameter was 8.05 mm. Conclusion Dual phase 99mTc-MIBI SPECT/CT imaging had a higher sensitivity in patients with SHPT than ultrasonography. Therefore, using 99mTc-MIBI positioning the lesion could be an effective method pre-surgical in patients with SHPT

Contralesional Cortical and Network Features Associated with Preoperative Language Deficit in Glioma Patients

Lower-grade Gliomas anchored in eloquent areas cause varying degrees of language impairment. Except for a tumor&rsquo;s features, contralesional compensation may explain these differences. Therefore, studying changes in the contralateral hemisphere can provide insights into the underlying mechanisms of language function compensation in patients with gliomas. This study included 60 patients with eloquent-area or near-eloquent-area gliomas. The participants were grouped according to the degree of language defect. T1 and diffusion tensor imaging were obtained. The contralesional cortical volume and the subcortical network were compared between groups. Patients with unimpaired language function showed elevated cortical volume in the midline areas of the frontal and temporal lobes. In subcortical networks, the group also had the highest global efficiency and shortest global path length. Ten nodes had intergroup differences in nodal efficiency, among which four nodes were in the motor area and four nodes were in the language area. Linear correlation was observed between the efficiency of the two nodes and the patient&rsquo;s language function score. Functional compensation in the contralesional hemisphere may alleviate language deficits in patients with gliomas. Structural compensation mainly occurs in the contralesional midline area in the frontal and temporal lobes, and manifests as an increase in cortical volume and subcortical network efficiency