In less than a decade, scientists located within 200 yards of one another identified the first cancer genes, discovered that a protein by itself can transmit an infectious disease, learned how to clone DNA, and founded the biotech company that created a new industry. The four discoverers, wild cards in the academic deck—Herbert Boyer, Michael Bishop, Harold Varmus, and Stanley Prusiner—came to San Francisco between 1966 and 1970, where they were joined by a cadre of face card leaders determined to transform a provincial campus into a major center for biomedical research. Their stories show that money and facilities are necessary but not sufficient for creative biomedical innovation. More crucially, the best discoverers respond to the lure of adventure and require freedom and time to tackle and solve hard problems in their own way. Rather than follow prescribed paths, real innovators need to roam free.

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Background

Clinical and genetic studies suggest circadian clock genes may contribute to biological mechanisms underlying alcohol use disorders (AUD). In particular, the Per2 gene regulates alcohol consumption in mutant animals, and in humans with AUD, the 10870 variant in PER2 has been associated with alcohol consumption. However, with respect to function, the molecular clock remains largely uncharacterized in AUD patients.

Methods

In skin fibroblast cultures from well-characterized human AUD patients (n = 19) and controls (n = 13), we used a bioluminescent reporter gene (Per2::luc) to measure circadian rhythms in gene expression at high sampling density for 5 days. Cells were genotyped for the PER2 10870 variant. The rhythm parameters period and amplitude were then analyzed using a case-control design and by genetic and clinical characteristics of the AUD subjects.

Results

There were no differences between AUD cases and controls in rhythm parameters. However, period was inversely correlated with illness severity (defined as the number of alcohol dependence criteria met). The PER2 variant 10870 was not associated with differences in rhythm parameters.

Conclusions

Our data suggest that differences in the cellular circadian clock are not pronounced in fibroblasts from AUD cases and controls. However, we found evidence that the circadian clock may be associated with an altered trajectory of AUD, possibly related to illness severity. Future work will be required to determine the mechanistic basis of this association.

We present a coarse-grained two dimensional mechanical model for the microtubule-tau bundles in neuronal axons in which we remove taus, as can happen in various neurodegenerative conditions such as Alzheimers disease, tauopathies, and chronic traumatic encephalopathy. Our simplified model includes (i) taus modeled as entropic springs between microtubules, (ii) removal of taus from the bundles due to phosphorylation, and (iii) a possible depletion force between microtubules due to these dissociated phosphorylated taus. We equilibrate upon tau removal using steepest descent relaxation. In the absence of the depletion force, the transverse rigidity to radial compression of the bundles falls to zero at about 60% tau occupancy, in agreement with standard percolation theory results. However, with the attractive depletion force, spring removal leads to a first order collapse of the bundles over a wide range of tau occupancies for physiologically realizable conditions. While our simplest calculations assume a constant concentration of microtubule intercalants to mediate the depletion force, including a dependence that is linear in the detached taus yields the same collapse. Applying percolation theory to removal of taus at microtubule tips, which are likely to be the protective sites against dynamic instability, we argue that the microtubule instability can only obtain at low tau occupancy, from 0.06-0.30 depending upon the tau coordination at the microtubule tips. Hence, the collapse we discover is likely to be more robust over a wide range of tau occupancies than the dynamic instability. We suggest in vitro tests of our predicted collapse.

Background

Lignocellulosic biomass is recognized as a promising renewable feedstock for the production of biofuels. However, current methods for converting biomass into fermentable sugars are considered too expensive and inefficient due to the recalcitrance of the secondary cell wall. Biomass composition can be modified to create varieties that are efficiently broken down to release cell wall sugars. This study focused on identifying the key biomass components influencing plant cell wall recalcitrance that can be targeted for selection in sugarcane, an important and abundant source of biomass.

Results

Biomass composition and the amount of glucan converted into glucose after saccharification were measured in leaf and culm tissues from seven sugarcane genotypes varying in fiber composition after no pretreatment and dilute acid, hydrothermal and ionic liquid pretreatments. In extractives-free sugarcane leaf and culm tissue, glucan, xylan, acid-insoluble lignin (AIL) and acid-soluble lignin (ASL) ranged from 20 to 32%, 15% to 21%, 14% to 20% and 2% to 4%, respectively. The ratio of syringyl (S) to guaiacyl (G) content in the lignin ranged from 1.5 to 2.2 in the culm and from 0.65 to 1.1 in the leaf. Hydrothermal and dilute acid pretreatments predominantly reduced xylan content, while the ionic liquid (IL) pretreatment targeted AIL reduction. The amount of glucan converted into glucose after 26 h of pre-saccharification was highest after IL pretreatment (42% in culm and 63.5% in leaf) compared to the other pretreatments. Additionally, glucan conversion in leaf tissues was approximately 1.5-fold of that in culm tissues. Percent glucan conversion varied between genotypes but there was no genotype that was superior to all others across the pretreatment groups. Path analysis revealed that S/G ratio, AIL and xylan had the strongest negative associations with percent glucan conversion, while ASL and glucan content had strong positive influences.

Conclusion

To improve saccharification efficiency of lignocellulosic biomass, breeders should focus on reducing S/G ratio, xylan and AIL content and increasing ASL and glucan content. This will be key for the development of sugarcane varieties for bioenergy uses.

Epithiodiketopiperazines (ETPs) are a structurally complex class of fungal natural products with potent anticancer activity. In ETPs, the diketopiperazine ring is spanned by a disulfide bond that is constrained in a high-energy eclipsed conformation. We employed computational, synthetic, and spectroscopic methods to investigate the physicochemical attributes of this atypical disulfide bond. We find that the disulfide bond is stabilized by two n→π* interactions, each with large energies (3-5 kcal/mol). The n→π* interactions in ETPs make disulfide reduction much more difficult, endowing stability in physiological environments in a manner that could impact their biological activity. These data reveal a previously unappreciated means to stabilize a disulfide bond and highlight the utility of the n→π* interaction in molecular design.

To fight COVID-19, much effort has been directed toward in vitro drug repurposing. Here, we investigate the impact of colloidal aggregation, a common screening artifact, in these repurposing campaigns. We tested 56 drugs reported as active in biochemical assays for aggregation by dynamic light scattering and by detergent-based enzyme counter screening; 19 formed colloids at concentrations similar to their literature IC₅₀'s, and another 14 were problematic. From a common repurposing library, we further selected another 15 drugs that had physical properties resembling known aggregators, finding that six aggregated at micromolar concentrations. This study suggests not only that many of the drugs repurposed for SARS-CoV-2 in biochemical assays are artifacts but that, more generally, at screening-relevant concentrations, even drugs can act artifactually via colloidal aggregation. Rapid detection of these artifacts will allow the community to focus on those molecules that genuinely have potential for treating COVID-19.

Like blood neutrophils, dHL60 cells respond to a uniform concentration of attractant by polarizing in apparently random directions. How each cell chooses its own direction is unknown. We now find that an arrow drawn from the center of the nucleus of an unpolarized cell to its centrosome strongly predicts the subsequent direction of attractant-induced polarity: Of 60 cells that polarized in response to uniform f-Met-Leu-Phe (fMLP), 42 polarized to the left of this arrow, 6 polarized to the right, and 12 polarized directly toward or away from the centrosome. To investigate this directional bias we perturbed a regulatory pathway, downstream of Cdc42 and partitioning-defective 6 (Par6), which controls centrosome orientation relative to polarity of other cells. Dominant negative Par6 mutants block polarity altogether, as previously shown for disrupting Cdc42 activity. Cells remain able to polarize, but without directional bias, if their microtubules are disrupted with nocodazole, or they express mutant proteins that interfere with activities of PKCzeta or dynein. Expressing constitutively active glycogen synthase kinase 3beta (GSK3beta) causes cells to polarize preferentially to the right. Distributions of most of these polarity regulators localize to the centrosome but show no left-right asymmetry before polarization. Together, these findings suggest that an intrinsically chiral structure, perhaps the centrosome, serves as a template for directing polarity in the absence of spatial cues. Such a template could help to determine left-right asymmetry and planar polarity in development.