Transcription factors form a ternary complex with NIPBL/MAU2 to localize cohesin at enhancers.

While the cohesin complex is a key player in genome architecture, how it localizes to specific chromatin sites is not understood. Recently, we and others have proposed that direct interactions with transcription factors lead to the localization of the cohesin-loader complex (NIPBL/MAU2) within enhancers. Here, we identify two clusters of LxxLL motifs within the NIPBL sequence that regulate NIPBL dynamics, interactome, and NIPBL-dependent transcriptional programs.

β-TrCP-Mediated Proteolysis of Mis18β Prevents Mislocalization of CENP-A and Chromosomal Instability.

Restricting the localization of evolutionarily conserved histone H3 variant CENP-A to the centromere is essential to prevent chromosomal instability (CIN), an important hallmark of cancers. Overexpressed CENP-A mislocalizes to non-centromeric regions and contributes to CIN in yeast, flies, and human cells. Centromeric localization of CENP-A is facilitated by the interaction of Mis18β with CENP-A specific chaperone HJURP.

Crossing boundaries of light microscopy resolution discerns novel assemblies in the nucleolus.

The nucleolus is the largest membraneless organelle and nuclear body in mammalian cells. It is primarily involved in the biogenesis of ribosomes, essential macromolecular machines responsible for synthesizing all proteins required by the cell. The assembly of ribosomes is evolutionarily conserved and accounts for the most energy-consuming cellular process needed for cell growth, proliferation, and homeostasis.

DNAJC9 prevents CENP-A mislocalization and chromosomal instability by maintaining the fidelity of histone supply chains.

The centromeric histone H3 variant CENP-A is overexpressed in many cancers. The mislocalization of CENP-A to noncentromeric regions contributes to chromosomal instability (CIN), a hallmark of cancer. However, pathways that promote or prevent CENP-A mislocalization remain poorly defined.

ST37 Distribution in the Moscow Region and Properties of Clinical and Foodborne Isolates.

of the phylogenetic lineage II (PLII) are common in the European environment and are hypovirulent. Despite this, they caused more than a third of the sporadic cases of listeriosis and multi-country foodborne outbreaks. ST37 is one of them.

Bayesian analysis dissects kinetic modulation during non-stationary gene expression.

Labelling of nascent stem loops with fluorescent proteins has fostered the visualization of transcription in living cells. Quantitative analysis of recorded fluorescence traces can shed light on kinetic transcription parameters and regulatory mechanisms. However, existing methods typically focus on steady state dynamics.

Architectural basis for cylindrical self-assembly governing Plk4-mediated centriole duplication in human cells.

Proper organization of intracellular assemblies is fundamental for efficient promotion of biochemical processes and optimal assembly functionality. Although advances in imaging technologies have shed light on how the centrosome is organized, how its constituent proteins are coherently architected to elicit downstream events remains poorly understood. Using multidisciplinary approaches, we showed that two long coiled-coil proteins, Cep63 and Cep152, form a heterotetrameric building block that undergoes a stepwise formation into higher molecular weight complexes, ultimately generating a cylindrical architecture around a centriole.

Impact of on the Field of Single-Molecule Biophysics.

Cellular functions depend on the dynamic assembly of protein regulator complexes at specific cellular locations. Single Molecule Tracking (SMT) is a method of choice for the biochemical characterization of protein dynamics in vitro and in vivo. SMT follows individual molecules in live cells and provides direct information about their behavior.

MYC amplifies gene expression through global changes in transcription factor dynamics.

The MYC oncogene has been studied for decades, yet there is still intense debate over how this transcription factor controls gene expression. Here, we seek to answer these questions with an in vivo readout of discrete events of gene expression in single cells. We engineered an optogenetic variant of MYC (Pi-MYC) and combined this tool with single-molecule RNA and protein imaging techniques to investigate the role of MYC in modulating transcriptional bursting and transcription factor binding dynamics in human cells.

The role of protease-activated receptor 1 signaling in CD8 T cell effector functions.

CD8 T cells are essential for adaptive immunity against viral infections. Protease activated receptor 1 (PAR1) is expressed by CD8 T cells; however, its role in T cell effector function is not well defined. Here we show that in human CD8 T cells, PAR1 stimulation accelerates calcium mobilization.

Towards a 'Spot On' Understanding of Transcription in the Nucleus.

Regulation of transcription by RNA Polymerase II (RNAPII) is a rapidly evolving area of research. Technological developments in microscopy have revealed insight into the dynamics, structure, and localization of transcription components within single cells. A frequent observation in many studies is the appearance of 'spots' in cell nuclei associated with the transcription process.

CENP-A overexpression promotes aneuploidy with karyotypic heterogeneity.

Chromosomal instability (CIN) is a hallmark of many cancers. Restricting the localization of centromeric histone H3 variant CENP-A to centromeres prevents CIN. CENP-A overexpression (OE) and mislocalization have been observed in cancers and correlate with poor prognosis; however, the molecular consequences of CENP-A OE on CIN and aneuploidy have not been defined.

Aggregation and Prion-Inducing Properties of the G-Protein Gamma Subunit Ste18 are Regulated by Membrane Association.

Yeast prions and mnemons are respectively transmissible and non-transmissible self-perpetuating protein assemblies, frequently based on cross-β ordered detergent-resistant aggregates (amyloids). Prions cause devastating diseases in mammals and control heritable traits in yeast. It was shown that the de novo formation of the prion form [] of yeast release factor Sup35 is facilitated by aggregates of other proteins.

Blockade of Toll-like receptor 4 (TLR4) reduces oxidative stress and restores phospho-ERK1/2 levels in Leydig cells exposed to high glucose.

Aims: Hyperglycemia in combination with oxidative stress plays a significant pathophysiological role in diabetic testicular dysfunction, often leading to infertility. Activation of Toll-like receptor 4 (TLR4) has been reported to mediate oxidative stress during diabetes. However, engagement of the TLR4 signaling pathway in diabetic testicular dysfunction has not been previously explored.

Toll-like receptor 4 (TLR4) as a possible pathological mechanism in hyperglycemia-associated testicular dysfunction.

Hyperglycemia is a chief factor in diabetes, a complex disease associated with reproductive disorders, mainly testicular dysfunction, which contributes to male infertility. Leydig cells are the predominant cell population in the testis interstitium and, when stimulated, they are capable of initiating immune responses playing crucial roles in the mechanisms related to testis' homeostasis. These cells express TLR4, an innate immune receptor, which is known to be modulated by hyperglycemia in other cell populations and tissue types.

Glucose Signaling Is Connected to Chromosome Segregation Through Protein Kinase A Phosphorylation of the Dam1 Kinetochore Subunit in .

The Dam1 complex is an essential component of the outer kinetochore that mediates attachments between spindle microtubules and chromosomes. Dam1p, a subunit of the Dam1 complex, binds to microtubules and is regulated by Aurora B/Ipl1p phosphorylation. We find that overexpression of cAMP-dependent protein kinase (PKA) catalytic subunits (, , , ) is lethal in mutants and increases the rate of chromosome loss in wild-type cells.

Single-Molecule Analysis Reveals Linked Cycles of RSC Chromatin Remodeling and Ace1p Transcription Factor Binding in Yeast.

It is unknown how the dynamic binding of transcription factors (TFs) is molecularly linked to chromatin remodeling and transcription. Using single-molecule tracking (SMT), we show that the chromatin remodeler RSC speeds up the search process of the TF Ace1p for its response elements (REs) at the CUP1 promoter. We quantified smFISH mRNA data using a gene bursting model and demonstrated that RSC regulates transcription bursts of CUP1 only by modulating TF occupancy but does not affect initiation and elongation rates.

Single molecule analysis of lamin dynamics.

The nuclear envelope (NE) is an essential cellular structure that contributes to nuclear stability, organization, and function. Mutations in NE-associated proteins result in a myriad of pathologies with widely diverse clinical manifestations, ages of onsets, and affected tissues. Notably, several hundred disease-causing mutations have been mapped to the LMNA gene, which encodes the intermediate filament proteins lamin A and C, two of the major architectural components of the nuclear envelope.

Associations of Simian Immunodeficiency Virus (SIV)-Specific Follicular CD8 T Cells with Other Follicular T Cells Suggest Complex Contributions to SIV Viremia Control.

Follicular CD8 T (fCD8) cells reside within B cell follicles and are thought to be immune-privileged sites of HIV/SIV infection. We have observed comparable levels of fCD8 cells between chronically SIV-infected rhesus macaques with low viral loads (LVL) and high viral loads (HVL), raising the question concerning their contribution to viremia control. In this study, we sought to clarify the role of SIV-specific fCD8 cells in lymph nodes during the course of SIV infection in rhesus macaques.

Single-molecule analysis of steroid receptor and cofactor action in living cells.

Population-based assays have been employed extensively to investigate the interactions of transcription factors (TFs) with chromatin and are often interpreted in terms of static and sequential binding. However, fluorescence microscopy techniques reveal a more dynamic binding behaviour of TFs in live cells. Here we analyse the strengths and limitations of in vivo single-molecule tracking and performed a comprehensive analysis on the intranuclear dwell times of four steroid receptors and a number of known cofactors.

Mislocalization of centromeric histone H3 variant CENP-A contributes to chromosomal instability (CIN) in human cells.

Chromosomal instability (CIN) is a hallmark of many cancers and a major contributor to tumorigenesis. Centromere and kinetochore associated proteins such as the evolutionarily conserved centromeric histone H3 variant CENP-A, associate with centromeric DNA for centromere function and chromosomal stability. Stringent regulation of cellular CENP-A levels prevents its mislocalization in yeast and flies to maintain genome stability.

Labeling Extracellular Vesicles for Nanoscale Flow Cytometry.

Extracellular vesicles (EVs), including exosomes and microvesicles, are 30-800 nm vesicles that are released by most cell types, as biological packages for intercellular communication. Their importance in cancer and inflammation makes EVs and their cargo promising biomarkers of disease and cell-free therapeutic agents. Emerging high-resolution cytometric methods have created a pressing need for efficient fluorescent labeling procedures to visualize and detect EVs.

Transcription-dependent radial distribution of TCF7L2 regulated genes in chromosome territories.

Human chromosomes occupy distinct territories in the interphase nucleus. Such chromosome territories (CTs) are positioned according to gene density. Gene-rich CTs are generally located in the center of the nucleus, while gene-poor CTs are positioned more towards the nuclear periphery.