Stress-Induced Changes in Nucleocytoplasmic Localization of Crucial Factors in Gene Expression Regulation.

This study investigates the toxic effect of harmful materials, unfiltered by the placenta, on neonatal umbilical cord (UC) vessels, focusing on stress-induced adaptations in transcriptional and translational processes. It aims to analyze changes in pathways related to mRNA condensate formation, transcriptional regulation, and DNA damage response under maternal smoking-induced stress. UC vessels from neonates born to smoking (Sm) and nonsmoking mothers (Ctr) were examined.

1,6-Hexanediol Is Inducing Homologous Recombination by Releasing BLM from Assemblysomes in .

We recently demonstrated that 1,6-hexanediol inhibits the formation of assemblysomes. These membraneless cell organelles have important roles in co-translational protein complex assembly and also store halfway translated DNA damage response proteins for a timely stress response. Recognizing the therapeutic potential of 1,6-hexanediol in dismantling assemblysomes likely to be involved in chemo- or radiotherapy resistance of tumor cells, we initiated an investigation into the properties of 1,6-hexanediol.

Predictive Potential of RNA Polymerase B (II) Subunit 1 (RPB1) Cytoplasmic Aggregation for Neoadjuvant Chemotherapy Failure.

We aimed to investigate the contribution of co-translational protein aggregation to the chemotherapy resistance of tumor cells. Increased co-translational protein aggregation reflects altered translation regulation that may have the potential to buffer transcription under genotoxic stress. As an indicator for such an event, we followed the cytoplasmic aggregation of RPB1, the aggregation-prone largest subunit of RNA polymerase II, in biopsy samples taken from patients with invasive carcinoma of no special type.

Phase-separated ribosome-nascent chain complexes in genotoxic stress response.

Assemblysomes are EDTA- and RNase-resistant ribonucleoprotein (RNP) complexes of paused ribosomes with protruding nascent polypeptide chains. They have been described in yeast and human cells for the proteasome subunit Rpt1, and the disordered amino-terminal part of the nascent chain was found to be indispensable for the accumulation of the Rpt1-RNP into assemblysomes. Motivated by this, to find other assemblysome-associated RNPs we used bioinformatics to rank subunits of protein complexes according to their amino-terminal disorder propensity.

Not1 and Not4 inversely determine mRNA solubility that sets the dynamics of co-translational events.

Background: The Ccr4-Not complex is mostly known as the major eukaryotic deadenylase. However, several studies have uncovered roles of the complex, in particular of the Not subunits, unrelated to deadenylation and relevant for translation. In particular, the existence of Not condensates that regulate translation elongation dynamics has been reported.

Despite its sequence identity with canonical H4, Drosophila H4r product is enriched at specific chromatin regions.

Histone variants are different from their canonical counterparts in structure and are encoded by solitary genes with unique regulation to fulfill tissue or differentiation specific functions. A single H4 variant gene (His4r or H4r) that is located outside of the histone cluster and gives rise to a polyA tailed messenger RNA via replication-independent expression is preserved in Drosophila strains despite that its protein product is identical with canonical H4. In order to reveal information on the possible role of this alternative H4 we epitope tagged endogenous H4r and studied its spatial and temporal expression, and revealed its genome-wide localization to chromatin at the nucleosomal level.

Not4 and Not5 modulate translation elongation by Rps7A ubiquitination, Rli1 moonlighting, and condensates that exclude eIF5A.

In this work, we show that Not4 and Not5 from the Ccr4-Not complex modulate translation elongation dynamics and change ribosome A-site dwelling occupancy in a codon-dependent fashion. These codon-specific changes in not5Δ cells are very robust and independent of codon position within the mRNA, the overall mRNA codon composition, or changes of mRNA expression levels. They inversely correlate with codon-specific changes in cells depleted for eIF5A and positively correlate with those in cells depleted for ribosome-recycling factor Rli1.

FKBP10 Regulates Protein Translation to Sustain Lung Cancer Growth.

Cancer therapy is limited, in part, by lack of specificity. Thus, identifying molecules that are selectively expressed by, and relevant for, cancer cells is of paramount medical importance. Here, we show that peptidyl-prolyl-cis-trans-isomerase (PPIase) FK506-binding protein 10 (FKBP10)-positive cells are present in cancer lesions but absent in the healthy parenchyma of human lung.

Co-translational assembly of proteasome subunits in NOT1-containing assemblysomes.

The assembly of large multimeric complexes in the crowded cytoplasm is challenging. Here we reveal a mechanism that ensures accurate production of the yeast proteasome, involving ribosome pausing and co-translational assembly of Rpt1 and Rpt2. Interaction of nascent Rpt1 and Rpt2 then lifts ribosome pausing.

Mutations in the Genes or in the Translation Machinery Similarly Display Increased Resistance to Histidine Starvation.

The genes encode subunits of the conserved Ccr4-Not complex, a global regulator of gene expression, and in particular of mRNA metabolism. They were originally identified in a selection for increased resistance to histidine starvation in the yeast . Recent work indicated that the Not5 subunit, ortholog of mammalian CNOT3, determines global translation levels by defining binding of the Ccr4-Not scaffold protein Not1 to ribosomal mRNAs during transcription.

Not5-dependent co-translational assembly of Ada2 and Spt20 is essential for functional integrity of SAGA.

Acetylation of histones regulates gene expression in eukaryotes. In the yeast Saccharomyces cerevisiae it depends mainly upon the ADA and SAGA histone acetyltransferase complexes for which Gcn5 is the catalytic subunit. Previous screens have determined that global acetylation is reduced in cells lacking subunits of the Ccr4–Not complex, a global regulator of eukaryotic gene expression.

Building on the Ccr4-Not architecture.

In a recent issue of Nature Communications Ukleja and co-workers reported a cryo-EM 3D reconstruction of the Ccr4-Not complex from Schizosaccharomyces pombe with an immunolocalization of the different subunits. The newly gained architectural knowledge provides cues to apprehend the functional diversity of this major eukaryotic regulator. Indeed, in the cytoplasm alone, Ccr4-Not regulates translational repression, decapping and deadenylation, and the Not module additionally plays a positive role in translation.

Translational Capacity of a Cell Is Determined during Transcription Elongation via the Ccr4-Not Complex.

The current understanding of gene expression considers transcription and translation to be independent processes. Challenging this notion, we found that translation efficiency is determined during transcription elongation through the imprinting of mRNAs with Not1, the central scaffold of the Ccr4-Not complex. We determined that another subunit of the complex, Not5, defines Not1 binding to specific mRNAs, particularly those produced from ribosomal protein genes.

Ccr4-Not is at the core of the eukaryotic gene expression circuitry.

In this mini-review, we summarize our current knowledge about the cross-talk between the different levels of gene expression. We introduce the Ccr4 (carbon catabolite repressed 4)-Not (negative on TATA-less) complex as a candidate to be a master regulator that orchestrates between the different levels of gene expression. An integrated view of the findings about the Ccr4-Not complex suggests that it is involved in gene expression co-ordination.

The Not5 subunit of the ccr4-not complex connects transcription and translation.

Recent studies have suggested that a sub-complex of RNA polymerase II composed of Rpb4 and Rpb7 couples the nuclear and cytoplasmic stages of gene expression by associating with newly made mRNAs in the nucleus, and contributing to their translation and degradation in the cytoplasm. Here we show by yeast two hybrid and co-immunoprecipitation experiments, followed by ribosome fractionation and fluorescent microscopy, that a subunit of the Ccr4-Not complex, Not5, is essential in the nucleus for the cytoplasmic functions of Rpb4. Not5 interacts with Rpb4; it is required for the presence of Rpb4 in polysomes, for interaction of Rpb4 with the translation initiation factor eIF3 and for association of Rpb4 with mRNAs.

High Fcp1 phosphatase activity contributes to setting an intense transcription rate required in Drosophila nurse and follicular cells for egg production.

During transcription cycles serine side chains in the carboxyl terminal domain (CTD) of the largest subunit of RNA polymerase II undergo dynamic phosphorylation-de-phosphorylation changes, and the modification status of the CTD serves as a signal for proteins involved in transcription and RNA maturation. We show here that the major CTD de-phosphorylating enzyme Fcp1 is expressed at high levels in germline cells of Drosophila. We used transgene constructs to modify the Fcp1 phosphatase level in Drosophila ovaries and found that high levels of Fcp1 are required for intensive gene expression in nurse cells.

The involvement of Importin-β and peroxiredoxin-6005 in mitochondrial biogenesis.

Importin-β is encoded by the Ketel gene in Drosophila. Upon running out of the maternal Importin-β dowry larvae without the Ketel gene slow down and before dying possess symptoms characteristic for mitochondrial cytopathies. Death of the larvae is almost certainly the consequence of ceasing import of proteins, including some of the transcription factors, into the nuclei.

The DRE motif is a key component in the expression regulation of the importin-beta encoding Ketel gene in Drosophila.

Importin-beta, encoded by the Ketel gene in Drosophila, is a key component of nuclear protein import, the formation of the spindle microtubules and the assembly of the nuclear envelope. The Drosophila embryos rely on the maternal importin-beta dowry at the beginning of their life. Expression of the zygotic Ketel gene commences during gastrulation in every cell and while the expression is maintained in the mitotically active diploid cells it ceases in the non-dividing larval cells in which nuclear protein import is assured by the long persisting importin-beta molecules.

Plaque-based competitive hybridization.

The authors have developed a simple, cost-saving experimental design, plaque-based competitive hybridization (PBCH), for genome-wide identification of genes differentially expressed in different tissues. PBCH offers advantages in comparison with other methods used in comparative genomics by combining the principles of differential hybridization with the subtractive hybridization. PBCH is particularly advantageous when libraries with few differences are to be analyzed.

Long persistence of importin-beta explains extended survival of cells and zygotes that lack the encoding gene.

Importin-beta is an essential component of nuclear protein import, spindle formation and nuclear envelope assembly. Formerly, the function of the Drosophila Ketel gene, which encodes importin-beta and is essential for the survival to adulthood, seemed to be required only in the mitotically active cells. We report here that importin-beta function is required in every cell and that this protein possesses an exceptionally long life span.