In cellular regulatory networks, genetic activity is controlled by molecular signals that determine when and how often a given gene is transcribed. In genetically controlled pathways, the protein product encoded by one gene often regulates expression of other genes. The time delay, after activation of the first promoter, to reach an effective level to control the next promoter depends on the rate of protein accumulation. We have analyzed the chemical reactions controlling transcript initiation and translation termination in a single such “genetically coupled” link as a precursor to modeling networks constructed from many such links. Simulation of the processes of gene expression shows that proteins are produced from an activated promoter in short bursts of variable numbers of proteins that occur at random time intervals. As a result, there can be large differences in the time between successive events in regulatory cascades across a cell population. In addition, the random pattern of expression of competitive effectors can produce probabilistic outcomes in switching mechanisms that select between alternative regulatory paths. The result can be a partitioning of the cell population into different phenotypes as the cells follow different paths. There are numerous unexplained examples of phenotypic variations in isogenic populations of both prokaryotic and eukaryotic cells that may be the result of these stochastic gene expression mechanisms.

https://www.pnas.org/content/pnas/94/3/814.full.pdf

Stochastic mechanisms in gene expression

https://faculty.buffalostate.edu/wadswogj/courses/450/RNAi%20Feeding%20Timmons.pdf

Ingestion of bacterially expressed dsRNAs can produce specific and potent genetic interference in Caenorhabditis elegans

Fluctuations in rates of gene expression can produce highly erratic time patterns of protein production in individual cells and wide diversity in instantaneous protein concentrations across cell populations. When two independently produced regulatory proteins acting at low cellular concentrations competitively control a switch point in a pathway, stochastic variations in their concentrations can produce probabilistic pathway selection, so that an initially homogeneous cell population partitions into distinct phenotypic subpopula- tions. Many pathogenic organisms, for example, use this mechanism to randomly switch surface features to evade host responses. This coupling between molecular-level fluctuations and macroscopic phenotype selection is analyzed using the phage l lysis-lysogeny decision circuit as a model system. The fraction of infected cells selecting the lysogenic pathway at different phage:cell ratios, predicted using a molecular- level stochastic kinetic model of the genetic regulatory circuit, is consistent with experimental observations. The kinetic model of the decision circuit uses the stochastic formulation of chemical kinetics, stochastic mechanisms of gene expression, and a statistical-thermodynamic model of promoter regulation. Conven- tional deterministic kinetics cannot be used to predict statistics of regulatory systems that produce probabilis- tic outcomes. Rather, a stochastic kinetic analysis must be used to predict statistics of regulatory outcomes for such stochastically regulated systems.

/pdf/stochastic-kinetic-analysis-of-developmental-pathway-545z0j2lmj.pdf

Stochastic kinetic analysis of developmental pathway bifurcation in phage lambda -infected escherichia coli cells

Cells are intrinsically noisy biochemical reactors: low reactant numbers can lead to significant statistical fluctuations in molecule numbers and reaction rates Here we use an analytic model to investigate the emergent noise properties of genetic systems We find for a single gene that noise is essentially determined at the translational level, and that the mean and variance of protein concentration can be independently controlled The noise strength immediately following single gene induction is almost twice the final steady-state value We find that fluctuations in the concentrations of a regulatory protein can propagate through a genetic cascade; translational noise control could explain the inefficient translation rates observed for genes encoding such regulatory proteins For an autoregulatory protein, we demonstrate that negative feedback efficiently decreases system noise The model can be used to predict the noise characteristics of networks of arbitrary connectivity The general procedure is further illustrated for an autocatalytic protein and a bistable genetic switch The analysis of intrinsic noise reveals biological roles of gene network structures and can lead to a deeper understanding of their evolutionary origin

/pdf/intrinsic-noise-in-gene-regulatory-networks-474991bvgn.pdf

Intrinsic noise in gene regulatory networks

Real-Time Kinetics of Gene Activity in Individual Bacteria

Transcription and translation initiation frequencies of the Escherichia coli lac operon.

Publisher Summary This chapter discusses that nucleic acid hybrid formation has been used as a research tool for approximately one decade. It has been a decade of remarkable progress toward understanding mechanisms of gene expression. Probably no other single procedure has contributed more to this success than has hybridization. However, the procedure is still usually used as a crude assay for genetic relatedness, used less often for precise quantitative estimations. Its value as a technique has become even greater with studies that are undertaken to determine the variables that contribute to final yield of hybrid. Hybridization has continued to be of great value for some time. The additional complexity of the genomes of higher organisms gives added complexity to interpretations of hybridizations with their nucleic acids. An indirect approach that is provided if small segments of eukaryotic chromosomes can be pursed, this is now possible for bacterial genes. The chapter concludes that the problems will have to be solved by more rigorous analyses and by refinements in the techniques. This will be necessary unless a chemical method more useful than hybridization is developed to study nucleic acid specificity.

Principles and practices of nucleic acid hybridization.

Specific endonucleolytic cleavage sites for decay of Escherichia coli mRNA.

Degradation of messenger RNA from the lactose operon (lac mRNA) was measured during the inhibition of protein synthesis by chloramphenicol (CM) or of translation-initiation by kasugamycin (KAS) With increasing CM concentration mRNA decay becomes slower, but there is no direct proportionality between rates of chemical decay and polypeptide synthesis During exponential growth lac mRNA is cleaved endonucleolytically (Blundell and Kennell, 1974) At a CM concentration which completely inhibits all polypeptide synthesis this cleavage is blocked In contrast, if only the initiation of translation is blocked by addition of KAS, the cleavage rate as well as the rate of chemical decay are increased significantly without delay These faster rates do not result from immediate degradation of the lengthening stretch of ribosome-free proximal message, since the full-length size is present and the same discrete message sizes are generated during inhibition

Translation and mRNA Decay

The endoribonuclease, RNase I, was purified from the periplasm of Escherichia coli. Based on PAGE, it has molecular mass of approximately 27 kDa with a migration rate indistinguishable from that of the recently reported RNase M from E. coli. The amino acid sequence of the two enzymes must be very similar based on two-dimensional mapping of their tryptic peptides and suggests either a post-transcriptional modification to yield different proteins from the same gene or evolution of two genes by gene duplication. However, while RNase I could degrade each of the four ribonucleotide homopolymers, only poly(U) or poly(C) were good substrates for RNase M with possibly some hydrolysis of poly(A). The reaction rate for poly(C) hydrolysis with RNase M was about ten times faster than for poly(U), while for RNase I the rates were about equal. Besides differences in specificity, RNase M was only located in the spheroplasts while RNase I found in the periplasm of growing cells. In terms of function, RNase I is known to cause degradation of rRNA during periods of stress or non-growth, whereas it has been proposed that RNase M is the endonuclease for mRNA degradation in growing cells.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1990.tb15336.x

David Kennell

Papers

Transcription and translation initiation frequencies of the Escherichia coli lac operon.

Principles and practices of nucleic acid hybridization.

Specific endonucleolytic cleavage sites for decay of Escherichia coli mRNA.

Translation and mRNA Decay

Purification and characterization of Escherichia coli RNase I. Comparisons with RNase M.