N-terminus of MSL1 is critical for dosage compensation.

The male-specific lethal complex (MSL), which consists of five proteins and two non-coding roX RNAs, is involved in the transcriptional enhancement of X-linked genes to compensate for the sex chromosome monosomy in XY males compared with XX females. The MSL1 and MSL2 proteins form the heterotetrameric core of the MSL complex and are critical for the specific recruitment of the complex to the high-affinity 'entry' sites (HAS) on the X chromosome. In this study, we demonstrated that the N-terminal region of MSL1 is critical for stability and functions of MSL1.

Functional Role of C-terminal Domains in the MSL2 Protein of Drosophila melanogaster.

Dosage compensation complex (DCC), which consists of five proteins and two non-coding RNAs roX, specifically binds to the X chromosome in males, providing a higher level of gene expression necessary to compensate for the monosomy of the sex chromosome in male Drosophila compared to the two X chromosomes in females. The MSL2 protein contains the N-terminal RING domain, which acts as an E3 ligase in ubiquitination of proteins and is the only subunit of the complex expressed only in males. Functional role of the two C-terminal domains of the MSL2 protein, enriched with proline (P-domain) and basic amino acids (B-domain), was investigated.

New Drosophila promoter-associated architectural protein Mzfp1 interacts with CP190 and is required for housekeeping gene expression and insulator activity.

In Drosophila, a group of zinc finger architectural proteins recruits the CP190 protein to the chromatin, an interaction that is essential for the functional activity of promoters and insulators. In this study, we describe a new architectural C2H2 protein called Madf and Zinc-Finger Protein 1 (Mzfp1) that interacts with CP190. Mzfp1 has an unusual structure that includes six C2H2 domains organized in a C-terminal cluster and two tandem MADF domains.

The N-terminal dimerization domains of human and Drosophila CTCF have similar functionality.

Background: CTCF is highly likely to be the ancestor of proteins that contain large clusters of C2H2 zinc finger domains, and its conservation is observed across most bilaterian organisms. In mammals, CTCF is the primary architectural protein involved in organizing chromosome topology and mediating enhancer-promoter interactions over long distances. In Drosophila, CTCF (dCTCF) cooperates with other architectural proteins to establish long-range interactions and chromatin boundaries.

Interaction of MLE with CLAMP zinc finger is involved in proper MSL proteins binding to chromosomes in .

The male-specific lethal (MSL) complex binds to the male X chromosome to activate transcription. It comprises five proteins (MSL1, MSL2, MSL3, male absent on the first (MOF), and maleless (MLE)) and two long noncoding RNAs (lncRNAs; roX1 and roX2). The MLE helicase remodels the roX lncRNAs, enabling the lncRNA-mediated assembly of the dosage compensation complex.

Study of the Role of Long Noncoding roX RNA in Maintaining of the Dosage Compensation Complex in Drosophila melanogaster.

The proteins MSL1, MSL2, MSL3, MLE, and MOF and noncoding RNAs roX1 and roX2 form the Drosophila dosage compensation complex (DCC), which specifically binds to the X chromosome of males. It is known that noncoding RNA roX are primary component of the DCC in the process of assembly and spreading of the complex among the X chromosome of males. However, the role of this RNA in maintaining the structure of the already assembled complex remains unclear.

The N-Terminal Part of CP190 Is a Platform for Interaction with Multiple Architectural Proteins.

CP190 is a co-factor in many architectural proteins, being involved in the formation of active promoters and insulators. CP190 contains the N-terminal BTB/POZ (Broad-Complex, Tramtrack and Bric a brac/POxvirus and Zinc finger) domain and adjacent conserved regions involved in protein interactions. Here, we examined the functional roles of these domains of CP190 in vivo.

CRISPR/Cas9 Essential Gene Editing in Drosophila.

Since the addition of the CRISPR/Cas9 technology to the genetic engineering toolbox, the problems of low efficiency and off-target effects hamper its widespread use in all fields of life sciences. Furthermore, essential gene knockout usually results in failure and it is often not obvious whether the gene of interest is an essential one. Here, we report on a new strategy to improve the CRISPR/Cas9 genome editing, which is based on the idea that editing efficiency is tightly linked to how essential the gene to be modified is.

Comparison of Compartmental and Non-Compartmental Analysis to Detect Biopharmaceutical Similarity of Intravenous Nanomaterial-Based Rifabutin Formulations.

Pharmacometric analysis is often used to quantify the differences and similarities between formulation prototypes. In the regulatory framework, it plays a significant role in the evaluation of bioequivalence. While non-compartmental analysis provides an unbiased data evaluation, mechanistic compartmental models such as the physiologically-based nanocarrier biopharmaceutics model promise improved sensitivity and resolution for the underlying causes of inequivalence.

The Essential Role of Prolines and Their Conformation in Allosteric Regulation of Kaiso Zinc Finger DNA-Binding Activity by the Adjacent C-Terminal Loop.

Kaiso is a methyl-DNA-binding protein containing three C2H2 zinc fingers with a C-terminal extension that participates in DNA binding. The linker between the last zinc finger and the DNA-binding portion of the extension contains two prolines that are highly conserved in vertebrates and in cognate ZBTB4 and ZBTB38 proteins. Prolines provide chain rigidity and can exist in cis and trans conformations that can be switched by proline isomerases, affecting protein function.

Structural basis for interaction between CLAMP and MSL2 proteins involved in the specific recruitment of the dosage compensation complex in Drosophila.

Transcriptional regulators select their targets from a large pool of similar genomic sites. The binding of the Drosophila dosage compensation complex (DCC) exclusively to the male X chromosome provides insight into binding site selectivity rules. Previous studies showed that the male-specific organizer of the complex, MSL2, and ubiquitous DNA-binding protein CLAMP directly interact and play an important role in the specificity of X chromosome binding.

Modulation of Aptamer-Ligand-Binding by Complementary Oligonucleotides: A G-Quadruplex Anti-Ochratoxin A Aptamer Case Study.

Short oligonucleotides are widely used for the construction of aptamer-based sensors and logical bioelements to modulate aptamer-ligand binding. However, relationships between the parameters (length, location of the complementary region) of oligonucleotides and their influence on aptamer-ligand interactions remain unclear. Here, we addressed this task by comparing the effects of short complementary oligonucleotides (ssDNAs) on the structure and ligand-binding ability of an aptamer and identifying ssDNAs' features that determine these effects.

Dimerization Activity of a Disordered N-Terminal Domain from CLAMP Protein.

In , CLAMP is an essential zinc-finger transcription factor that is involved in chromosome architecture and functions as an adaptor for the dosage compensation complex. Most of the known architectural proteins have structural N-terminal homodimerization domains that facilitate distance interactions. Because CLAMP performs architectural functions, we tested its N-terminal region for the presence of a homodimerization domain.

Mechanisms of CP190 Interaction with Architectural Proteins in Drosophila Melanogaster.

Most of the known Drosophila architectural proteins interact with an important cofactor, CP190, that contains three domains (BTB, M, and D) that are involved in protein-protein interactions. The highly conserved N-terminal CP190 BTB domain forms a stable homodimer that interacts with unstructured regions in the three best-characterized architectural proteins: dCTCF, Su(Hw), and Pita. Here, we identified two new CP190 partners, CG4730 and CG31365, that interact with the BTB domain.

Fluorescently Labeled PLGA Nanoparticles for Visualization In Vitro and In Vivo: The Importance of Dye Properties.

Fluorescently labeled nanoparticles are widely used for evaluating their distribution in the biological environment. However, dye leakage can lead to misinterpretations of the nanoparticles' biodistribution. To better understand the interactions of dyes and nanoparticles and their biological environment, we explored PLGA nanoparticles labeled with four widely used dyes encapsulated (coumarin 6, rhodamine 123, DiI) or bound covalently to the polymer (Cy5.

Mutations of Phosphorylation Sites in MSL1 Protein Do Not Affect Dosage Compensation in Drosophila melanogaster.

Proteins MSL1 and MSL2 form the core of the Drosophila dosage compensation complex, which specifically binds to the X chromosome of males. Phosphorylation of certain amino acid residues was previously shown to regulate MSL1 activity. In the present work, transgenic lines of Drosophila expressing mutant variants of the MSL1 protein were obtained, in which amino acids undergoing phosphorylation were replaced.

Drosophila architectural protein CTCF is not essential for fly survival and is able to function independently of CP190.

CTCF is the most likely ancestor of proteins that contain large clusters of C2H2 zinc finger domains (C2H2) and is conserved among most bilateral organisms. In mammals, CTCF functions as the main architectural protein involved in the organization of topology-associated domains (TADs). In vertebrates and Drosophila, CTCF is involved in the regulation of homeotic genes.

TTK Isoforms Interact with Two Regions of the Mep-1 Protein of Drosophila melanogaster.

The Drosophila TTK protein is involved in the processes of cell differentiation and is represented by two isoforms, TTK69 and TTK88, which have a common N-terminal BTB domain and different C-terminal sequences. Earlier, it was shown that TTK69 represses the activity of enhancers and promoters by recruiting a conserved among higher eukaryotes NURD complex to chromatin. The Mep-1 protein was found in the NURD-complex of Drosophila, and this protein can interact with the C-terminal region of TTK69.

Exploring the systemic delivery of a poorly water-soluble model drug to the retina using PLGA nanoparticles.

During the drug development process, many pharmacologically active compounds are discarded because of poor water solubility, but nanoparticle-based formulations are increasingly proposed as a solution for this problem. We therefore studied the distribution of nanoparticulate carriers and the delivery of their poorly water-soluble cargo to a structure of the central nervous system, the retina, under naive and pathological conditions. The lipophilic fluorescent dye coumarin 6 (Cou6) was encapsulated into poly(lactic-co-glycolic acid) PLGA nanoparticles (NPs).

CTCF As an Example of DNA-Binding Transcription Factors Containing Clusters of C2H2-Type Zinc Fingers.

In mammals, most of the boundaries of topologically associating domains and all well-studied insulators are rich in binding sites for the CTCF protein. According to existing experimental data, CTCF is a key factor in the organization of the architecture of mammalian chromosomes. A characteristic feature of the CTCF is that the central part of the protein contains a cluster consisting of eleven domains of C2H2-type zinc fingers, five of which specifically bind to a long DNA sequence conserved in most animals.

Mechanism and functional role of the interaction between CP190 and the architectural protein Pita in Drosophila melanogaster.

Background: Pita is required for Drosophila development and binds specifically to a long motif in active promoters and insulators. Pita belongs to the Drosophila family of zinc-finger architectural proteins, which also includes Su(Hw) and the conserved among higher eukaryotes CTCF. The architectural proteins maintain the active state of regulatory elements and the long-distance interactions between them.

A Non-stop identity complex (NIC) supervises enterocyte identity and protects from premature aging.

A hallmark of aging is loss of differentiated cell identity. Aged midgut differentiated enterocytes (ECs) lose their identity, impairing tissue homeostasis. To discover identity regulators, we performed an RNAi screen targeting ubiquitin-related genes in ECs.

Exploring the Interplay between Drug Release and Targeting of Lipid-Like Polymer Nanoparticles Loaded with Doxorubicin.

Targeted delivery of doxorubicin still poses a challenge with regards to the quantities reaching the target site as well as the specificity of the uptake. In the present approach, two colloidal nanocarrier systems, NanoCore-6.4 and NanoCore-7.

The insulator functions of the polydactyl C2H2 zinc finger protein CTCF: Necessity versus sufficiency.

In mammals, a C2H2 zinc finger (C2H2) protein, CTCF, acts as the master regulator of chromosomal architecture and of the expression of Hox gene clusters. Like mammalian CTCF, the homolog, dCTCF, localizes to boundaries in the bithorax complex (BX-C). Here, we have determined the minimal requirements for the assembly of a functional boundary by dCTCF and two other C2H2 zinc finger proteins, Pita and Su(Hw).