Regulation of RNA-binding proteins affinity to export receptors enables the nuclear basket proteins to distinguish and retain aberrant mRNAs.

Export of messenger ribonucleic acids (mRNAs) into the cytoplasm is a fundamental step in gene regulation processes, which is meticulously quality controlled by highly efficient mechanisms in eukaryotic cells. Yet, it remains unclear how the aberrant mRNAs are recognized and retained inside the nucleus. Using a new modelling approach for complex systems, namely the agent-based modelling (ABM) approach, we develop a minimal model of the mRNA quality control (QC) mechanism. Our results demonstrate that regulation of the affinity of RNA-binding proteins (RBPs) to export receptors along with the weak interaction between the nuclear basket protein (Mlp1 or Tpr) and RBPs are the minimum requirements to distinguish and retain aberrant mRNAs. Our results show that the affinity between Tpr and RBPs is optimized to maximize the retention of aberrant mRNAs. In addition, we demonstrate how the length of mRNA affects the QC process. Since longer mRNAs spend more time in the nuclear basket to form a compact conformation and initiate their export, nuclear basket proteins could more easily capture and retain them inside the nucleus.

Interferon Beta: From Molecular Level to Therapeutic Effects.

Interferon beta (IFNß) is a cytokine that is naturally produced by the immune system in response to biological and chemical stimuli. It signals by binding to the heterodimeric type I IFN receptor composed of the IFNAR1 and IFNAR2 chains, and regulates the expression of a plethora of genes by means of the classical JAK/STAT and other pathways. IFNß is pleiotropic in that it elicits antiviral, antiproliferative, and immunomodulatory activities on numerous cell types. The biological activities underpin the mechanisms by which the protein is used to treat various diseases such as hepatitis C infection and multiple sclerosis. Despite the success of IFNß therapy, the drug may evoke the production of antidrug antibodies that may reduce treatment efficiency. Immunogenicity is related to many factors: among them, structural properties, particularly aggregation, and T-cell and B-cell epitopes in the structure of IFNß, appear to be important. Knowledge of the structural properties of IFNß and its relation to immunogenicity may help scientists to develop safer and more effective forms. Several methods have been used to predict and reduce the immunogenicity of certain IFNß drug products. In this chapter, we review the current knowledge on IFNß from its structure, dynamic conformation, signaling pathway, and mechanism of action to its therapeutic effects. Immunogenicity and its relation to structural properties of IFNß are also discussed.

Rapid Brownian Motion Primes Ultrafast Reconstruction of Intrinsically Disordered Phe-Gly Repeats Inside the Nuclear Pore Complex.

Conformational behavior of intrinsically disordered proteins, such as Phe-Gly repeat domains, alters drastically when they are confined in, and tethered to, nan channels. This has challenged our understanding of how they serve to selectively facilitate translocation of nuclear transport receptor (NTR)-bearing macromolecules. Heterogeneous FG-repeats, tethered to the NPC interior, nonuniformly fill the channel in a diameter-dependent manner and adopt a rapid Brownian motion, thereby forming a porous and highly dynamic polymeric meshwork that percolates in radial and axial directions and features two distinguishable zones: a dense hydrophobic rod-like zone located in the center, and a peripheral low-density shell-like zone. The FG-meshwork is locally disrupted upon interacting with NTR-bearing macromolecules, but immediately reconstructs itself between 0.44 µs and 7.0 µs, depending on cargo size and shape. This confers a perpetually-sealed state to the NPC, and is solely due to rapid Brownian motion of FG-repeats, not FG-repeat hydrophobic bonds. Elongated-shaped macromolecules, both in the presence and absence of NTRs, penetrate more readily into the FG-meshwork compared to their globular counterparts of identical volume and surface chemistry, highlighting the importance of the shape effects in nucleocytoplasmic transport. These results can help our understanding of geometrical effects in, and the design of, intelligent and responsive biopolymer-based materials in nanofiltration and artificial nanopores.

Evolutionarily Conserved Sequence Features Regulate the Formation of the FG Network at the Center of the Nuclear Pore Complex.

The nuclear pore complex (NPC) is the portal for bidirectional transportation of cargos between the nucleus and the cytoplasm. While most of the structural elements of the NPC, i.e. nucleoporins (Nups), are well characterized, the exact transport mechanism is still under much debate. Many of the functional Nups are rich in phenylalanine-glycine (FG) repeats and are believed to play the key role in nucleocytoplasmic transport. We present a bioinformatics study conducted on more than a thousand FG Nups across 252 species. Our results reveal the regulatory role of polar residues and specific sequences of charged residues, named 'like charge regions' (LCRs), in the formation of the FG network at the center of the NPC. Positively charged LCRs prepare the environment for negatively charged cargo complexes and regulate the size of the FG network. The low number density of charged residues in these regions prevents FG domains from forming a relaxed coil structure. Our results highlight the significant role of polar interactions in FG network formation at the center of the NPC and demonstrate that the specific localization of LCRs, FG motifs, charged, and polar residues regulate the formation of the FG network at the center of the NPC.

Biophysical coarse-grained modeling provides insights into transport through the nuclear pore complex.

The nuclear pore complex (NPC) is the gatekeeper of the nucleus, capable of actively discriminating between the active and inert cargo while accommodating a high rate of translocations. The biophysical mechanisms underlying transport, however, remain unclear due to the lack of information about biophysical factors playing role in transport. Based on published experimental data, we have established a coarse-grained model of an intact NPC structure to examine nucleocytoplasmic transport with refined spatial and temporal resolutions. Using our model, we estimate the transport time versus cargo sizes. Our findings suggest that the mean transport time of cargos smaller than 15 nm is independent of size, while beyond this size, there is a sharp increase in the mean transport time. The model confirms that kap-FG hydrophobicity is sufficient for active cargo transport. Moreover, our model predicts that during translocation, small and large cargo-complexes are hydrophobically attached to FG-repeat domains for 86 and 96% of their transport time, respectively. Inside the central channel FG-repeats form a thick layer on the wall leaving an open tube. The cargo-complex is almost always attached to this layer and diffuses back and forth, regardless of the cargo size. Finally, we propose a plausible model for transport in which the NPC can be viewed as a lubricated gate. This model incorporates basic assumptions underlying virtual-gate and reduction-of-dimensionality models with the addition of the FG-layer inside the central channel acting as a lubricant.

Nuclear pore complex: biochemistry and biophysics of nucleocytoplasmic transport in health and disease.

Nuclear pore complexes (NPCs) are the gateways connecting the nucleoplasm and cytoplasm. This structures are composed of over 30 different proteins and 60-125 MDa of mass depending on type of species. NPCs are bilateral pathways that selectively control the passage of macromolecules into and out of the nucleus. Molecules smaller than 40 kDa diffuse through the NPC passively while larger molecules require facilitated transport provided by their attachment to karyopherins. Kinetic studies have shown that approximately 1000 translocations occur per second per NPC. Maintaining its high selectivity while allowing for rapid translocation makes the NPC an efficient chemical nanomachine. In this review, we approach the NPC function via a structural viewpoint. Putting together different pieces of this puzzle, this chapter confers an overall insight into what molecular processes are engaged in import/export of active cargos across the NPC and how different transporters regulate nucleocytoplasmic transport. In the end, the correlation of several diseases and disorders with the NPC structural defects and dysfunctions is discussed.

Quantitative analysis of epithelial morphogenesis in Drosophila oogenesis: New insights based on morphometric analysis and mechanical modeling.

The process of epithelial morphogenesis is ubiquitous in animal development, but much remains to be learned about the mechanisms that shape epithelial tissues. The follicle cell (FC) epithelium encapsulating the growing germline of Drosophila is an excellent system to study fundamental elements of epithelial development. During stages 8 to 10 of oogenesis, the FC epithelium transitions between simple geometries-cuboidal, columnar and squamous-and redistributes cell populations in processes described as posterior migration, squamous cell flattening and main body cell columnarization. Here we have carried out a quantitative morphometric analysis of these poorly understood events in order to establish the parameters of and delimit the potential processes that regulate the transitions. Our results compel a striking revision of accepted views of these phenomena, by showing that posterior migration does not involve FC movements, that there is no role for columnar cell apical constriction in FC morphogenesis, and that squamous cell flattening may be a compliant response to germline growth. We utilize mechanical modeling involving finite element computational technologies to demonstrate that time-varying viscoelastic properties and growth are sufficient to account for the bulk of the FC morphogenetic changes.

Molecular dynamics study of talin-vinculin binding.

Cells can sense mechanical force in regulating focal adhesion assembly. One vivid example is the force-induced recruitment of vinculin to reinforce initial contacts between a cell and the extracellular matrix. Crystal structures of the unbound proteins and bound complex between the vinculin head subdomain (Vh1) and the talin vinculin binding site 1 (VBS1) indicate that vinculin undergoes a conformational change upon binding to talin. However, the molecular basis for this event and the precise nature of the binding pathway remain elusive. In this article, molecular dynamics is used to investigate the binding mechanism of Vh1 and VBS1 under minimal constraints to facilitate binding. One simulation demonstrates binding of the two molecules in the complete absence of external force. VBS1 makes early hydrophobic contact with Vh1 by positioning the critical hydrophobic residues (L608, L615, and L622) in the groove formed by helices 1 and 2 of Vh1. The solvent-exposed hydrophobic residues (V619 and L623) then gradually penetrate the hydrophobic core of Vh1, thus further separating helix 1 from helix 2. These critical residues are highly conserved as large hydrophobic side groups in other vinculin binding sites; studies also have demonstrated that these residues are essential in Vh1-VBS1 binding. Similar binding mechanisms are also demonstrated in separate molecular dynamics simulations of Vh1 binding to other vinculin binding sites both in talin and alpha-actinin.