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1. T7Max transcription system

   https://doi.org/10.1186/s13036-023-00323-1  

   Deich, Christopher; Cash, Brock; Sato, Wakana; Sharon, Judee; Aufdembrink, Lauren; Gaut, Nathaniel J.; Heili, Joseph; Stokes, Kaitlin; Engelhart, Aaron E.; Adamala, Katarzyna P. (December 2023, Journal of Biological Engineering)

   Abstract BackgroundEfficient cell-free protein expression from linear DNA templates has remained a challenge primarily due to template degradation. In addition, the yields of transcription in cell-free systems lag behind transcriptional efficiency of live cells. Most commonly used in vitro translation systems utilize T7 RNA polymerase, which is also the enzyme included in many commercial kits. ResultsHere we present characterization of a variant of T7 RNA polymerase promoter that acts to significantly increase the yields of gene expression withinin vitrosystems. We have demonstrated that T7Max increases the yield of translation in many types of commonly used in vitro protein expression systems. We also demonstrated increased protein expression yields from linear templates, allowing the use of T7Max driven expression from linear templates. ConclusionsThe modified promoter, termed T7Max, recruits standard T7 RNA polymerase, so no protein engineering is needed to take advantage of this method. This technique could be used with any T7 RNA polymerase- basedin vitroprotein expression system. 

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2. Phosphorylation of mitochondrial transcription factor B2 controls mitochondrial DNA binding and transcription

   https://doi.org/10.1016/j.bbrc.2020.05.141  

   Bostwick, Alicia M.; Moya, Gonzalo E.; Senti, Mackenna L.; Basu, Urmimala; Shen, Jiayu; Patel, Smita S.; Dittenhafer-Reed, Kristin E. (July 2020, Biochemical and Biophysical Research Communications)
3. Quantitative dissection of transcription in development yields evidence for transcription-factor-driven chromatin accessibility

   https://doi.org/10.7554/eLife.56429  

   Eck, Elizabeth; Liu, Jonathan; Kazemzadeh-Atoufi, Maryam; Ghoreishi, Sydney; Blythe, Shelby A; Garcia, Hernan G (October 2020, eLife)

   null (Ed.)

   Cells in the brain, liver and skin, as well as many other organs, all contain the same DNA, yet behave in very different ways. This is because before a gene can produce its corresponding protein, it must first be transcribed into messenger RNA. As an organism grows, the transcription of certain genes is switched on or off by regulatory molecules called transcription factors, which guide cells towards a specific ‘fate’. These molecules bind to specific locations within the regulatory regions of DNA, and for decades biologist have tried to use the arrangement of these sites to predict which proteins a cell will make. Theoretical models known as thermodynamic models have been able to successfully predict transcription in bacteria. However, this has proved more challenging to do in eukaryotes, such as yeast, fruit flies and humans. One of the key differences is that DNA in eukaryotes is typically tightly wound into bundles called nucleosomes, which must be disentangled in order for transcription factors to access the DNA. Previous thermodynamic models have suggested that DNA in eukaryotes randomly switches between being in a wound and unwound state. The models assume that once unwound, regulatory proteins stabilize the DNA in this form, making it easier for other transcription factors to bind to the DNA. Now, Eck, Liu et al. have tested some of these models by studying the transcription of a gene involved in the development of fruit flies. The experiments showed that no thermodynamic model could accurately mimic how this gene is regulated in the embryos of fruit flies. This led Eck, Liu et al. to identify a model that is better at predicting the activation pattern of this developmental gene. In this model, instead of just ‘locking’ DNA into an unwound shape, transcription factors can also actively speed up the unwinding of DNA. This improved understanding builds towards the goal of predicting gene regulation, where DNA sequences can be used to tell where and when cell decisions will be made. In the future, this could allow the development of new types of therapies that can regulate transcription in different diseases. 

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4. An AT-hook transcription factor promotes transcription of histone, spliced-leader, and piRNA clusters

   https://doi.org/10.1093/nar/gkaf079  

   Wang, Yi-Hui; Hertz, Hannah_L; Pastore, Benjamin; Tang, Wen (February 2025, Nucleic Acids Research)

   Abstract In all three domains of life, genes with related functions can be organized into specific genomic regions known as gene clusters. In eukaryotes, histone, piRNA (Piwi-interacting RNA), and rDNA (ribosomal DNA) clusters are among the most notable clusters which play fundamental roles in chromatin formation, genome integrity, and translation, respectively. These clusters have long been thought to be regulated by distinct transcriptional mechanisms. In this study, using Caenorhabditis elegans as a model system we identify ATTF-6, a member of the AT-hook family, as a key factor for the expression of histone, piRNA, and 5S rDNA-SL1 (spliced leader 1) clusters. ATTF-6 is essential for C. elegans viability. It forms distinct nuclear foci at both piRNA and 5S rDNA-SL1 clusters. Loss of ATTF-6 leads to a depletion of histone mRNAs, SL1 transcripts, and piRNAs. Additionally, we demonstrate that ATTF-6 is required for the recruitment of USTC (Upstream Sequence Transcription Complex) to piRNA clusters, which is necessary for piRNA production. Collectively, our findings reveal a unifying role for an AT-hook transcription factor in promoting the expression of fundamental gene clusters. 

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5. Mechanisms and Functions of the RNA Polymerase II General Transcription Machinery during the Transcription Cycle

   https://doi.org/10.3390/biom14020176  

   Archuleta, Stephen R.; Goodrich, James A.; Kugel, Jennifer F. (February 2024, Biomolecules)

   Central to the development and survival of all organisms is the regulation of gene expression, which begins with the process of transcription catalyzed by RNA polymerases. During transcription of protein-coding genes, the general transcription factors (GTFs) work alongside RNA polymerase II (Pol II) to assemble the preinitiation complex at the transcription start site, open the promoter DNA, initiate synthesis of the nascent messenger RNA, transition to productive elongation, and ultimately terminate transcription. Through these different stages of transcription, Pol II is dynamically phosphorylated at the C-terminal tail of its largest subunit, serving as a control mechanism for Pol II elongation and a signaling/binding platform for co-transcriptional factors. The large number of core protein factors participating in the fundamental steps of transcription add dense layers of regulation that contribute to the complexity of temporal and spatial control of gene expression within any given cell type. The Pol II transcription system is highly conserved across different levels of eukaryotes; however, most of the information here will focus on the human Pol II system. This review walks through various stages of transcription, from preinitiation complex assembly to termination, highlighting the functions and mechanisms of the core machinery that participates in each stage. 

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