Current opinion in structural biology最新文献

Splicing is a critical processing step during pre-mRNA maturation in eukaryotes. The correct selection of splice sites during the early steps of spliceosome assembly is highly important and crucial for the regulation of alternative splicing. Splice site recognition and alternative splicing depend on cis-regulatory sequence elements in the RNA and trans-acting splicing factors that recognize these elements and crosstalk with the canonical splicing machinery. Structural mechanisms involving early spliceosome complexes are governed by dynamic RNA structures, protein-RNA interactions and conformational flexibility of multidomain RNA binding proteins. Here, we highlight structural studies and integrative structural biology approaches, which provide complementary information from cryo-EM, NMR, small angle scattering, and X-ray crystallography to elucidate mechanisms in the regulation of early spliceosome assembly and quality control, highlighting the role of conformational dynamics.

RNA, either from invading pathogens or within the hosts, is one of the principal PAMPs (pathogen-associated molecular patterns). Toll-like receptors (TLRs) and other receptors of the innate immune system exist that detect immunostimulatory RNA including double and single stranded RNA, and then induce cytokine-mediated antiviral and proinflammatory responses. Recent years have seen remarkable progress in biochemical, immunological, and structural biological studies on TLRs, opening new avenues for TLR signaling. In this review, we highlight our current understanding of RNA- sensing TLRs and discuss the regulatory mechanisms that normally prevent inappropriate responses to self.

Molecular simulations of biological systems tend to be significantly more compute-intensive than those in materials science and astrophysics, due to important contributions of long-range electrostatic forces and large numbers of time steps (>1E9) required. Simulations of biomolecular complexes of microseconds to milliseconds are considered state-of-the-art today. However, these time scales are miniscule in comparison to physiological time scales relevant to molecular machine activity, drug action, and elongation cycles for protein synthesis, RNA synthesis, and DNA synthesis (seconds to days). While an exascale supercomputer has simulated an entire virus for nanoseconds, this supercomputer would need to be 10 billion times faster to simulate that virus for 3 hours of physiological time, demonstrating the insatiable need for computing power. With growing interest in computational drug design from the pharmaceutical sector, the biological sciences are positioned to be an industry driver in computing.

DNA confined to nanofluidic channels with a cross-section from tens to hundreds of nm wide and hundreds of microns long stretches in an equilibrium process free of flow or end tethering. Because DNA is free to move along the channel axis, its extension is exquisitely sensitive to DNA–DNA interactions and the DNA persistence length, as well as the contour length. We discuss how this sensitivity has been used to probe DNA-protein interactions at physiological concentrations of both DNA and proteins.

HIV-1, the causative agent of AIDS, is a retrovirus that packages two copies of unspliced viral RNA as a dimer into newly budding virions. The unspliced viral RNA also serves as an mRNA template for translation of two polyproteins. Recent studies suggest that the fate of the viral RNA (genome or mRNA) is determined at the level of transcription. RNA polymerase II uses heterogeneous transcription start sites to generate major transcripts that differ in only two guanosines at the 5ʹ end. Remarkably, this two-nucleotide difference is sufficient to alter the structure of the 5ʹ-untranslated region and generate two pools of RNA with distinct functions. The presence of both RNA species is needed for optimal viral replication and fitness.

RNA's ability to form and interconvert between multiple secondary and tertiary structures is critical to its functional versatility and the traditional view of RNA structures as static entities has shifted towards understanding them as dynamic conformational ensembles. In this review we discuss RNA structural ensembles and their dynamics, highlighting the concept of conformational energy landscapes as a unifying framework for understanding RNA processes such as folding, misfolding, conformational changes, and complex formation. Ongoing advancements in cryo-electron microscopy and chemical probing techniques are significantly enhancing our ability to investigate multiple structures adopted by conformationally dynamic RNAs, while traditional methods such as nuclear magnetic resonance spectroscopy continue to play a crucial role in providing high-resolution, quantitative spatial and temporal information. We discuss how these methods, when used synergistically, can provide a comprehensive understanding of RNA conformational ensembles, offering new insights into their regulatory functions.

While the structure/function paradigm for folded domains was established decades ago, our understanding of how intrinsically disordered regions (IDRs) contribute to biological function is still evolving. IDRs exist as conformational ensembles that can range from highly compact to highly extended depending on their sequence composition. IDR sequences are less conserved than those of folded domains, but often display short, conserved segments termed short linear motifs (SLiMs), that often mediate protein–protein interactions and are often regulated by posttranslational modifications, giving rise to complex functionality when multiple, differently regulated SLiMs are combined. This combinatorial functionality was associated with signaling and regulation soon after IDRs were first recognized as functional elements within proteins. Here, we discuss roles for disorder in proteins that regulate cyclin-dependent kinases, the master timekeepers of the eukaryotic cell cycle. We illustrate the importance of intrinsic flexibility in the transmission of regulatory signals by these entirely disordered proteins.

Evolution has fostered robust DNA damage response (DDR) mechanisms to combat DNA lesions. However, disruptions in this intricate machinery can render cells overly reliant on the remaining functional but often less accurate DNA repair pathways. This increased dependence on error-prone pathways may result in improper repair and the accumulation of mutations, fostering genomic instability and facilitating the uncontrolled cell proliferation characteristic of cancer initiation and progression. Strategies based on the concept of synthetic lethality (SL) leverage the inherent genomic instability of cancer cells by targeting alternative pathways, thereby inducing selective death of cancer cells. This review emphasizes recent advancements in structural investigations of pivotal SL targets. The significant contribution of structure-based methodologies to SL research underscores their potential impact in characterizing the growing number of SL targets, largely due to advances in next-generation sequencing. Harnessing these approaches is essential for advancing the development of precise and personalized SL therapeutic strategies.

RNAs are remarkably versatile molecules that can fold into intricate three-dimensional (3D) structures to perform diverse cellular and viral functions. Despite their biological importance, relatively few RNA 3D structures have been solved, and our understanding of RNA structure–function relationships remains in its infancy. This limitation partly arises from challenges posed by RNA's complex conformational landscape, characterized by structural flexibility, formation of multiple states, and a propensity to misfold. Recently, cryo-electron microscopy (cryo-EM) has emerged as a powerful tool for the visualization of conformationally dynamic RNA-only 3D structures. However, RNA's characteristics continue to pose challenges. We discuss experimental methods developed to overcome these hurdles, including the engineering of modular modifications that facilitate the visualization of small RNAs, improve particle alignment, and validate structural models.