Search

Reports a study that shows ASH1 messenger RNA (mRNA) preferentially accumulates in daughter cells by a process that is dependent on actin and myosin. The study results suggesting that localization of mRNA may have been an early property of the eukaryotic lineage; References and notes.

Discusses research which studied triplet-repeat diseases. What such diseases are; Aspects of each generation of a repeated sequence of three nucleotides; Gauging the severity of such diseases; Cause of such diseases related to genetic defect; Triplet repeats outside the coding region; Models to explain such diseases; Example of myotonic dystrophy; Repeats that result in a gain of function; Cases of two myotonic dystrophy patients.

The physical manifestation of learning and memory formation in the brain can be expressed by strengthening or weakening of synaptic connections through morphological changes. Local actin remodeling underlies some forms of plasticity and may be facilitated by local β-actin synthesis, but dynamic information is lacking. In this work, we use single-molecule in situ hybridization to demonstrate that dendritic β-actin messenger RNA (mRNA) and ribosomes are in a masked, neuron-specific form. Chemically induced long-term potentiation prompts transient mRNA unmasking, which depends on factors active during synaptic activity. Ribosomes and single β-actin mRNA motility increase after stimulation, indicative of release from complexes. Hence, the single-molecule assays we developed allow for the quantification of activity-induced unmasking and availability for active translation. Further, our work demonstrates that β-actin mRNA and ribosomes are in a masked state that is alleviated by stimulation. [ABSTRACT FROM AUTHOR]

Translation is the fundamental biological process converting mRNA information into proteins. Single-molecule imaging in live cells has illuminated the dynamics of RNA transcription; however, it is not yet applicable to translation. Here, we report single-molecule imaging of nascent peptides (SINAPS) to assess translation in live cells.The approach provides direct readout of initiation, elongation, and location of translation.We show that mRNAs coding for endoplasmic reticulum (ER) proteins are translated when they encounter the ER membrane. Single-molecule fluorescence recovery after photobleaching provides direct measurement of elongation speed (5 amino acids per second). In primary neurons, mRNAs are translated in proximal dendrites but repressed in distal dendrites and display "bursting" translation.This technology provides a tool with which to address the spatiotemporal translation mechanism of single mRNAs in living cells. [ABSTRACT FROM AUTHOR]

The article focuses on the development of a fluorescence microscopy method that reports on first translation events of individual messenger RNA molecules. It states the biosensor will allow scientists to examine spatiotemporal regulation of translation during normal growth and stress, as well as during oocyte development in Drosophila. It reports the study found messenger RNA wasn't translated in the cell nucleus and that translation is regulated by sequestration within P-bodies.

Cellular messenger RNA levels are achieved by the combinatorial complexity of factors controlling transcription, yet the small number of molecules involved in these pathways fluctuates stochastically. It has not yet been experimentally possible to observe the activity of single polymerases on an endogenous gene to elucidate how these events occur in vivo. Here, we describe a method of fluctuation analysis of fluorescently labeled RNA to measure dynamics of nascent RNA--including initiation, elongation, and termination--at an active yeast locus. We find no transcriptional memory between initiation events, and elongation speed can vary by threefold throughout the cell cycle. By measuring the abundance and intranuclear mobility of an upstream transcription factor, we observe that the gene firing rate is directly determined by trans-activating factor search times. [ABSTRACT FROM AUTHOR]

The article presents the discussion on DNA surveillance pathway using the biomechanical link between supercoiling and transcription for homeostatically amplifying transcriptional fluctuations including differentiation of embryonic stem cells into neural ectodermal cells.

Understanding gene expression requires the ability to follow the fate of individual molecules. Here we use a cellular system for monitoring messenger RNA (mRNA) expression to characterize the movement in real time of single mRNA-protein complexes (mRNPs) in the nucleus of living mammalian cells. This mobility was not directed but was governed by simple diffusion. Some mRNPs were partially corralled throughout the nonhomogenous nuclear environment, but no accumulation at subnuclear domains was observed. Following energy deprivation, energy-independent motion of mRNPs was observed in a highly ATP-dependent nuclear environment; movements were constrained to chromatinpoor domains and excluded by newly formed chromatin barriers. This observation resolves a controversy, showing that the energetic requirements of nuclear mRNP trafficking are consistent with a diffusional model. [ABSTRACT FROM AUTHOR]

A key goal of biology is to relate the expression of specific genes to a particular cellular phenotype. However, current assays for gene expression destroy the structural context. By combining advances in computational fluorescence microscopy with multiplex probe design, we devised technology in which the expression of many genes can be visualized simultaneously inside single cells with high spatial and temporal resolution. Analysis of 11 genes in serum-stimulated cultured cells revealed unique patterns of gene expression within individual cells. Using the nucleus as the substrate for parallel gene analysis, we provide a platform for the fusion of genomics and cell biology: “cellular genomics.” [ABSTRACT FROM AUTHOR]

Reports on experiments in which fluorescence in situ hybridization (FISH) and digital imaging microscopy were modified to allow detection of single RNA molecules. Use of synthesized oligodeoxynucleotide probes; Detection of RNA transcription; Implications of findings.

The transcription and transport of messenger RNA (mRNA) are critical steps in regulating the spatial and temporal components of gene expression, but it has not been possible to observe the dynamics of endogenous mRNA in primary mammalian tissues. We have developed a transgenic mouse in which all β-actin mRNA is fluorescently labeled. We found that β-actin mRNA in primary fibroblasts localizes predominantly by diffusion and trapping as single mRNAs. In cultured neurons and acute brain slices, we found that multiple β-actin mRNAs can assemble together, travel by active transport, and disassemble upon depolarization by potassium chloride. Imaging of brain slices revealed immediate early induction of β-actin transcription after depolarization. Studying endogenous mRNA in live mouse tissues provides insight into its dynamic regulation within the context of the cellular and tissue microenvironment. [ABSTRACT FROM AUTHOR]