Epigenetic Profiling Reveals a Developmental Decrease in Promoter Accessibility During Cortical Maturation in vivo

Axon regeneration in adult central nervous system (CNS) is limited in part by a developmental decline in the ability of injured neurons to re-express needed regeneration associated genes (RAGs). Adult CNS neurons may lack appropriate pro-regenerative transcription factors, or may display chromatin structure that restricts transcriptional access to RAGs. Here we performed epigenetic profiling around the promoter regions of key RAGs, and found progressive restriction across a time course of cortical maturation. These data identify a potential intrinsic constraint to axon growth in adult CNS neurons. Neurite outgrowth from cultured postnatal cortical neurons, however, proved insensitive to treatments that improve axon growth in other cell types, including combinatorial overexpression of AP1 factors, overexpression of histone acetyltransferases, and pharmacological inhibitors of histone deacetylases. This insensitivity could be due to intermediate chromatin closure at the time of culture, and highlights important differences in cell culture models used to test potential pro-regenerative interventions

Population Dynamics with Threshold Effects Give Rise to a Diverse Family of Allee Effects

The Allee effect describes populations that deviate from logistic growth models and arises in applications including ecology and cell biology. A common justification for incorporating Allee effects into population models is that the population in question has altered growth mechanisms at some critical density, often referred to as a threshold effect. Despite the ubiquitous nature of threshold effects arising in various biological applications, the explicit link between local threshold effects and global Allee effects has not been considered. In this work, we examine a continuum population model that incorporates threshold effects in the local growth mechanisms. We show that this model gives rise to a diverse family of Allee effects and we provide a comprehensiv

Rethinking the Political Future: An Alternative to the Ethno-Sectarian Division of Iraq

In the coming year, the political leadership in Iraq will need to make a final determination as to whether they are going to structure the state of Iraq as a federal state with ethnically heterogeneous provinces, a loose federal state with ethnically defined provinces or regions, or whether they are going to divide the state into three new states based on ethno-sectarian lines. A number of prominent American law makers and foreign policy shapers have strongly advocated for the soft, and sometimes hard, partition of Iraq — either through the creation of a loose federal structure based on ethno-sectarian lines, or through its outright partition. These commentators have prophesized that the ethno-sectarian division of Iraq “may soon be all we have left. In fact, the ethno-sectarian division of Iraq is fraught with logistical infeasibilities and dangers that threaten to compound the issues facing the people of Iraq instead of solving them. The political solution rests not on a return to failed approaches of division and entrenched conflict, but rather on the construction of a viable modern federal state that promotes unity, political compromise, and consensus building. To address the question of whether the future of Iraq rests with ethno-sectarian division or with multi-ethnic federalism, this Article first addresses the ideas behind ethno-sectarian division and describes the most prominent plans for the division of Iraq along ethno-sectarian lines. This Article then critiques such a division of Iraq by: (1) identifying the overwhelming lack of popular support for such a division; (2) exposing the practical and political difficulties of dividing a state as diverse and heterogeneous as Iraq; (3) discussing the likelihood that ethno-sectarian division will increase violent conflict; (4) highlighting the lessons of prior ethno-sectarian divisionist attempts; (5) noting insurmountable constitutional hurdles; and (6) setting out the significant signs of recent progress and cooperation in the Iraqi political framework

Exact sharp-fronted solutions for nonlinear diffusion on evolving domains

Models of diffusive processes that occur on evolving domains are frequently employed to describe biological and physical phenomena, such as diffusion within expanding tissues or substrates. Previous investigations into these models either report numerical solutions or require an assumption of linear diffusion to determine exact solutions. Unfortunately, numerical solutions do not reveal the relationship between the model parameters and the solution features. Additionally, experimental observations typically report the presence of sharp fronts, which are not captured by linear diffusion. Here we address both limitations by presenting exact sharp-fronted solutions to a model of degenerate nonlinear diffusion on a growing domain. We obtain the solution by identifying a series of transformations that converts the model of nonlinear diffusion on a growing domain to the porous medium equation on a fixed domain, which admits known exact solutions. We determine expressions for critical time scales and domain growth rates such that the diffusive population never reaches the domain boundaries and hence the solution remains valid

Exact solutions for diffusive transport on heterogeneous growing domains

From the smallest biological systems to the largest cosmological structures, spatial domains undergo expansion and contraction. Within these growing domains, diffusive transport is a common phenomenon. Mathematical models have been widely employed to investigate diffusive processes on growing domains. However, a standard assumption is that the domain growth is spatially uniform. There are many relevant examples where this is not the case, such as the colonisation of growing gut tissue by neural crest cells. As such, it is not straightforward to disentangle the individual roles of heterogeneous growth and diffusive transport. Here we present exact solutions to models of diffusive transport on domains undergoing spatially non-uniform growth. The exact solutions are obtained via a combination of transformation, convolution and superposition techniques. We verify the accuracy of these solutions via comparison with simulations of a corresponding lattice-based random walk. We explore various domain growth functions, including linear growth, exponential growth and contraction, and oscillatory growth. Provided the domain size remains positive, we find that the derived solutions are valid. The exact solutions reveal the relationship between model parameters, such as the diffusivity and the type and rate of domain growth, and key statistics, such as the survival and splitting probabilities

The Application of CRISPR Technology to High Content Screening in Primary Neurons

Axon growth is coordinated by multiple interacting proteins that remain incompletely characterized. High content screening (HCS), in which manipulation of candidate genes is combined with rapid image analysis of phenotypic effects, has emerged as a powerful technique to identify key regulators of axon outgrowth. Here we explore the utility of a genome editingapproach referred to as CRISPR (Clustered Regularly Interspersed Palindromic Repeats) for knockout screening in primary neurons. In the CRISPR approach a DNA-cleaving Cas enzyme is guided to genomic target sequences by user-created guide RNA (sgRNA), where it initiates a double-stranded break that ultimately results in frameshift mutation and loss of protein production. Using electroporation of plasmid DNA that co-expresses Cas9enzyme and sgRNA, we first verified the ability of CRISPR targeting to achieve protein-level knockdown in cultured postnatal cortical neurons. Targeted proteins included NeuN (RbFox3) and PTEN, a well-studied regulator of axon growth. Effective knockdown lagged at least four days behind transfection, but targeted proteins were eventually undetectable by immunohistochemistry in \u3e 80% of transfected cells. Consistent with this, anti-PTEN sgRNA produced no changes in neurite outgrowth when assessed three days post-transfection. When week-long cultures were replated, however, PTEN knockdown consistently increased neurite lengths. These CRISPR-mediated PTEN effects were achieved using multi-well transfection and automated phenotypic analysis, indicating the suitability of PTEN as a positive control for future CRISPR-based screening efforts. Combined, these data establish an example of CRISPR-mediated protein knockdown in primary cortical neurons and its compatibility with HCS workflows