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1. Yeast surface display of full‐length human microtubule‐associated protein tau

   https://doi.org/10.1002/btpr.2920  

   Wang, Shiyao; Cho, Yong_Ku (October 2019, Biotechnology Progress)

   Abstract Microtubule‐associated protein tau is an intrinsically disordered, highly soluble protein found primarily in neurons. Under normal conditions, tau regulates the stability of axonal microtubules and intracellular vesicle transport. However, in patients of neurodegeneration such as Alzheimer's disease (AD), tau forms neurofibrillary deposits, which correlates well with the disease progression. Identifying molecular signatures in tau, such as posttranslational modification, truncation, and conformational change has great potential to detect earliest signs of neurodegeneration and develop therapeutic strategies. Here, we show that full‐length human tau, including the longest isoform found in the adult brain, can be robustly displayed on the surface of yeastSaccharomyces cerevisiae. Yeast‐displayed tau binds to anti‐tau antibodies that cover epitopes ranging from the N‐terminus to the 4R repeat region. Unlike tau expressed in the yeast cytosol, surface‐displayed tau was not phosphorylated at sites found in AD patients (probed by antibodies AT8, AT270, AT180, and PHF‐1). However, yeast‐displayed tau showed clear binding to paired helical filament (PHF) tau conformation‐specific antibodies Alz‐50, MC‐1, and Tau‐2. Although the tau possessed a conformation found in PHFs, oligomerization or aggregation into larger filaments was undetected. Taken together, yeast‐displayed tau enables robust measurement of protein interactions and is of particular interest for characterizing conformational change. 

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2. Investigation of the pharmacokinetic properties of synthetic heparan sulfate oligosaccharides

   https://doi.org/10.1093/glycob/cwac068  

   Arnold, Katelyn; Wang, Zhangjie; Lucas, Andrew; Zamboni, William; Xu, Yongmei; Liu, Jian (October 2022, Glycobiology)

   Abstract Heparan sulfate (HS) is a sulfated polysaccharide with a wide range of biological activities. There is an increasing interest in the development of structurally homogeneous HS oligosaccharides as therapeutics. However, the factors influencing the pharmacokinetic properties of HS-based therapeutics remain unknown. Here, we report the pharmacokinetic properties of a panel of dodecasaccharides (12-mers) with varying sulfation patterns in healthy mice and uncover the pharmacokinetic properties of an octadecasaccharide (18-mer) in acutely injured mice. In the 12-mer panel, 1 12-mer, known as dekaparin, is anticoagulant, and 3 12-mers are nonanticoagulant. The concentrations of 12-mers in plasma and urine were determined by the disaccharide analysis using liquid chromatography coupled with tandem mass spectrometry. We observed a striking difference between anticoagulant and nonanticoagulant oligosaccharides in the 12-mer panel, showing that anticoagulant dekaparin had a 4.6-fold to 8.6-fold slower clearance and 4.4-fold to 8-fold higher plasma exposure compared to nonanticoagulant 12-mers. We also observed that the clearance of HS oligosaccharides is impacted by disease. Using an antiinflammatory 18-mer, we discovered that the clearance of 18-mer is reduced 2.8-fold in a liver failure mouse model compared to healthy mice. Our results suggest that oligosaccharides are rapidly cleared renally if they have low interaction with circulating proteins. We observed that the clearance rate of oligosaccharides is inversely associated with the degree of binding to target proteins, which can vary in response to pathophysiological conditions. Our findings uncover a contributing factor for the plasma and renal clearance of oligosaccharides which will aid the development of HS-based therapeutics. 

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3. Tau regulates Arc stability in neuronal dendrites via a proteasome-sensitive but ubiquitin-independent pathway

   https://doi.org/10.1016/j.jbc.2024.107237  

   Yakout, Dina W; Shroff, Ankit; Wei, Wei; Thaker, Vishrut; Allen, Zachary D; Sajish, Mathew; Nazarko, Taras Y; Mabb, Angela M (May 2024, Journal of Biological Chemistry)

   DeMartino, George (Ed.)

   Tauopathies are neurodegenerative disorders characterized by the deposition of aggregates of the microtubule-associated protein tau, a main component of neurofibrillary tangles. Alzheimer’s disease (AD) is the most common type of tauopathy and dementia, with amyloid-beta pathology as an additional hallmark feature of the disease. Besides its role in stabilizing microtubules, tau is localized at postsynaptic sites and can regulate synaptic plasticity. The activity-regulated cytoskeleton-associated protein (Arc) is an immediate early gene that plays a key role in synaptic plasticity, learning, and memory. Arc has been implicated in AD pathogenesis and regulates the release of amyloid-beta. We found that decreased Arc levels correlate with AD status and disease severity. Importantly, Arc protein was upregulated in the hippocampus of Tau KO mice and dendrites of Tau KO primary hippocampal neurons. Overexpression of tau decreased Arc stability in an activity-dependent manner, exclusively in neuronal dendrites, which was coupled to an increase in the expression of dendritic and somatic surface GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors. The tau-dependent decrease in Arc was found to be proteasome-sensitive, yet independent of Arc ubiquitination and required the endophilin-binding domain of Arc. Importantly, these effects on Arc stability and GluA1 localization were not observed in the commonly studied tau mutant, P301L. These observations provide a potential molecular basis for synaptic dysfunction mediated through the accumulation of tau in dendrites. Our findings confirm that Arc is misregulated in AD and further show a physiological role for tau in regulating Arc stability and AMPA receptor targeting. 

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4. Activity of bacteria isolated from bats against Pseudogymnoascus destructans in China

   https://doi.org/10.1111/1751-7915.13765  

   Li, Zhongle; Li, Aoqiang; Hoyt, Joseph R.; Dai, Wentao; Leng, Haixia; Li, Yanfei; Li, Wei; Liu, Sen; Jin, Longru; Sun, Keping; et al (February 2021, Microbial Biotechnology)

   Summary White‐nose syndrome, a disease that is caused by the psychrophilic fungusPseudogymnoascus destructans, has threatened several North America bat species with extinction. Recent studies have shown that East Asian bats are infected withP. destructansbut show greatly reduced infections. While several factors have been found to contribute to these reduced infections, the role of specific microbes in limitingP. destructansgrowth remains unexplored. We isolated three bacterial strains with the ability to inhibitP. destructans, namely,Pseudomonas yamanorumGZD14026,Pseudomonas brenneriXRD11711 andPseudomonas fragiGZD14479, from bats in China.Pseudomonas yamanorum, with the highest inhibition score, was selected to extract antifungal active substance. Combining mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy analyses, we identified the active compound inhibitingP. destructansas phenazine‐1‐carboxylic acid (PCA), and the minimal inhibitory concentration (MIC) was 50.12 μg ml⁻¹. Whole genome sequencing also revealed the existence of PCA biosynthesis gene clusters. Gas chromatography‐mass spectrometry (GC‐MS) analysis identified volatile organic compounds. The results indicated that 10 ppm octanoic acid, 100 ppm 3‐tert‐butyl‐4‐hydroxyanisole (isoprenol) and 100 ppm 3‐methyl‐3‐buten‐1‐ol (BHA) inhibited the growth ofP. destructans. These results support that bacteria may play a role in limiting the growth ofP. destructanson bats. 

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5. Basement Membrane Mimetic Hydrogel Cooperates with Rho‐Associated Protein Kinase Inhibitor to Promote the Development of Acini‐Like Salivary Gland Spheroids

   https://doi.org/10.1002/anbr.202300088  

   Fowler, Eric W.; Witt, Robert L.; Jia, Xinqiao (November 2023, Advanced NanoBiomed Research)

   Successful engineering of functional salivary glands necessitates the creation of cell‐instructive environments for ex vivo expansion and lineage specification of primary human salivary gland stem cells (hS/PCs). Herein, basement membrane mimetic hydrogels are prepared using hyaluronic acid, cell adhesive peptides, and hyperbranched polyglycerol (HPG), with or without sulfate groups, to produce “hyperGel+” or “hyperGel”, respectively. Differential scanning fluorescence experiments confirm the ability of the sulfated HPG precursor to stabilize fibroblast growth factor 10. The hydrogels are nanoporous, cytocompatible, and cell‐permissive, enabling the development of multicellular hS/PC spheroids in 14 days. The incorporation of sulfated HPG species in the hydrogel enhances cell proliferation. Culture of hS/PCs in hyperGel+ in the presence of a Rho kinase inhibitor Y‐27632 (Y‐27) leads to the development of spheroids with a central lumen, increases the expression of acinar marker aquaporin‐3 at the transcript level (AQP3), and decreases the expression of ductal marker keratin 7 at both the transcript (KRT7) and the protein levels (K7). Reduced expression of transforming growth factor beta (TGF‐β) targets SMAD2/3 is also observed in Y27‐treated cultures, suggesting attenuation of TGF‐β signaling. Thus, hyperGel+ cooperates with the Rho‐associated protein kinase inhibitor to promote the development of lumened spheroids with enhanced expression of acinar markers. 

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