Hundreds of myosin 10s are pushed to the tips of filopodia and could cause traffic jams on actin

Myosin 10 (Myo10) is a motor protein known for its role in filopodia formation. Although Myo10-driven filopodial dynamics have been characterized, there is no information about the absolute number of Myo10 molecules during the filopodial lifecycle. To better understand molecular stoichiometries and packing restraints in filopodia, we measured Myo10 abundance in these structures. We combined SDS-PAGE densitometry with epifluorescence microscopy to quantitate HaloTag-labeled Myo10 in U2OS cells. About 6% of total intracellular Myo10 localizes to filopodia, where it enriches at opposite cellular ends. Hundreds of Myo10s are in a typical filopodium, and their distribution across filopodia is log-normal. Some filopodial tips even contain more Myo10 than accessible binding sites on the actin filament bundle. Live-cell movies reveal a dense cluster of over a hundred Myo10 molecules that initiates filopodial elongation. Hundreds of Myo10 molecules continue to accumulate during filopodial growth, but accumulation ceases when retraction begins. Rates of filopodial elongation, second-phase elongation, and retraction are inversely related to Myo10 quantities. Our estimates of Myo10 molecules in filopodia provide insight into the physics of packing Myo10, its cargo, and other filopodia-associated proteins in narrow membrane compartments. Our protocol provides a framework for future work analyzing Myo10 abundance and distribution upon perturbation

A Method for Photoinitating Protein Folding in a Nondenaturing Environment

The early kinetic events of protein folding are an important part of the folding pathway, yet our understanding towards the process is limited. Information from the study of these early events can allow us to distinguish between the various models that have been proposed to describe the folding of a protein in real time. Unlike “typical” chemical kinetics with well-defined initial and final states, the initial state of a denatured protein is relatively ill-defined. This uncertainty introduces ambiguity in the interpretation of the experimental data on the early events in protein folding. Toward developing a unified theory of protein folding, it is necessary to begin the observation of the refolding process from a well-defined initial state, trigger folding as rapidly as possible, and to follow the protein in real time as it samples its conformational space over its highly complex free-energy landscape

Snapshots of a molecular swivel in action

Members of the serine family of site-specific recombinases exchange DNA strands via 180° rotation about a central protein-protein interface. Modeling of this process has been hampered by the lack of structures in more than one rotational state for any individual serine recombinase. Here we report crystal structures of the catalytic domains of four constitutively active mutants of the serine recombinase Sin, providing snapshots of rotational states not previously visualized for Sin, including two seen in the same crystal. Normal mode analysis predicted that each tetramer's lowest frequency mode (i.e. most accessible large-scale motion) mimics rotation: two protomers rotate as a pair with respect to the other two. Our analyses also suggest that rotation is not a rigid body movement around a single symmetry axis but instead uses multiple pivot points and entails internal motions within each subunit

Insights from quantitative and mathematical modelling on the proposed 2030 goal for gambiense human African trypanosomiasis (gHAT)

Gambiense human African trypanosomiasis (gHAT) is a parasitic, vector-borne neglected tropical disease that has historically affected populations across West and Central Africa and can result in death if untreated. Following from the success of recent intervention programmes against gHAT, the World Health Organization (WHO) has defined a 2030 goal of global elimination of transmission (EOT). The key proposed indicator to measure achievement of the goal is to have zero reported cases. Results of previous mathematical modelling and quantitative analyses are brought together to explore both the implications of the proposed indicator and the feasibility of achieving the WHO goal. Whilst the indicator of zero case reporting is clear and measurable, it is an imperfect proxy for EOT and could arise either before or after EOT is achieved. Lagging reporting of infection and imperfect diagnostic specificity could result in case reporting after EOT, whereas the converse could be true due to underreporting, lack of coverage, and cryptic human and animal reservoirs. At the village-scale, the WHO recommendation of continuing active screening until there are three years of zero cases yields a high probability of local EOT, but extrapolating this result to larger spatial scales is complex. Predictive modelling of gHAT has consistently found that EOT by 2030 is unlikely across key endemic regions if current medical-only strategies are not bolstered by improved coverage, reduced time to detection and/or complementary vector control. Unfortunately, projected costs for strategies expected to meet EOT are high in the short term and strategies that are cost-effective in reducing burden are unlikely to result in EOT by 2030. Future modelling work should aim to provide predictions while taking into account uncertainties in stochastic dynamics and infection reservoirs, as well as assessment of multiple spatial scales, reactive strategies, and measurable proxies of EOT

Dance, sports or academics: which monopolize students emotional quotient

Background: Emotional Intelligence (EI), often measured as an Emotional Intelligence Quotient (EQ), refers to skills that provide the ability to balance emotions and reason, which provides long-term happiness. Focus on Intelligence Quotient (IQ) in the current era has to be changed for the betterment of society. Objectives was to assess and compare Emotional Quotient (EQ) among students who excel in dance, sports, and academics.Methods: This was a cross-sectional study conducted among three groups of minimum 30 students who are excellent in dance, academics or sports. EQ was assessed using a 22 item EQ test which measures the psychological dimensions such as emotional sensitivity, emotional maturity and emotional competency and also total EQ.Results: Out of 91 students, maximum students were having extremely high EQ in the domains of emotional sensitivity 36 (39.6%), emotional competency 88 (96.7%) and total EQ 85 (93.4%). Dancers showed significantly high EQ in the domains of emotional maturity, emotional competency, and overall EQ compared to sportspersons and academicians. Increased age, agriculture and business occupation of mother and presence of another sibling were significantly associated with emotional intelligence of students.Conclusions: This study showed that inclusion of extracurricular activities like dance and sports along with academics may improve the self-confidence and increased awareness about one’s own emotions, leading to raised EQ

Fast Benchtop Fabrication of Laminar Flow Chambers for Advanced Microscopy Techniques

Background: Fluid handling technology is acquiring an ever more prominent place in laboratory science whether it is in simple buffer exchange systems, perfusion chambers, or advanced microfluidic devices. Many of these applications remain the providence of laboratories at large institutions with a great deal of expertise and specialized equipment. Even with the expansion of these techniques, limitations remain that frequently prevent the coupling of controlled fluid flow with other technologies, such as coupling microfluidics and high-resolution position and force measurements by optical trapping microscopy. Method: Here we present a method for fabrication of multiple-input laminar flow devices that are optically clear [glass] on each face, chemically inert, reusable, inexpensive, and can be fabricated on the benchtop in approximately one hour. Further these devices are designed to allow flow regulation by a simple gravity method thus requiring no specialized equipment to drive flow. Here we use these devices to perform total internal reflection fluorescence microscopy measurements as well as position sensitive optical trapping experiments. Significance: Flow chamber technology needs to be more accessible to the general scientific community. The method presented here is versatile and robust. These devices use standard slides and coverslips making them compatible with nearly all types and models of light microscopes. These devices meet the needs of groups doing advanced optical trapping experiments, but could also be adapted by nearly any lab that has a function for solution flow coupled with microscopy

Brief of Corporate Law Professors as Amici Curie in Support of Respondents

The Supreme Court has looked to the rights of corporate shareholders in determining the rights of union members and non-members to control political spending, and vice versa. The Court sometimes assumes that if shareholders disapprove of corporate political expression, they can easily sell their shares or exercise control over corporate spending. This assumption is mistaken. Because of how capital is saved and invested, most individual shareholders cannot obtain full information about corporate political activities, even after the fact, nor can they prevent their savings from being used to speak in ways with which they disagree. Individual shareholders have no “opt out” rights or practical ability to avoid subsidizing corporate political expression with which they disagree. Nor do individuals have the practical option to refrain from putting their savings into equity investments, as doing so would impose damaging economic penalties and ignore conventional financial guidance for individual investors