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The authors review the publications on RNA polymerase II transcription machinery. The topics of interest include general transcription factors, Mediator complex transmitting regulatory information to enzyme, site-specific protein-DNA crosslinking studies and molecular architecture of this machinery.

Research has been conducted on the yeast RNA polymerase II mutant. The mechanisms of activation in eukaryotes and bacteria are discussed.

Survival of mycobacteria, both free-living and host-dependent pathogenic species, is dependent on their ability to evade being killed by the stresses they routinely encounter. Toxin-antitoxin (TA) systems are unique to bacteria and archaea and are thought to function as stress survival proteins. Here, we study the activity of the endoribonuclease toxin derived from the MazEF TA system in Mycobacterium smegmatis , designated MazEF-ms. We first enlisted a specialized RNA-seq method, 5' RNA-seq, to identify the primary RNA target(s) of the MazF-ms toxin. Just two tRNA species, tRNALys-UUU and tRNALys-CUU, were targeted for cleavage by MazF-ms at a single site within their anticodon sequence (UU↓U and CU↓U) to render these tRNAs nonfunctional for protein synthesis. The 5' RNA-seq dataset also revealed hallmarks of ribosome stalling predominantly at Lys AAA codons even though both Lys tRNAs were cleaved by MazF-ms. Stalled ribosomes were then cleaved on their 5' side by one or more RNases, resulting in very selective degradation of only those mRNAs harboring ribosomes stalled at Lys codons. This highly surgical, codon-dependent degradation of mRNA transcripts was validated using quantitative mass spectrometry of proteins that were newly synthesized during MazF-ms expression. The M. smegmatis proteome was altered as predicted, Lys AAA codon-rich proteins was downregulated while Lys AAA codon deficient proteins were upregulated. Analysis of specific subsets of proteins that were upregulated or downregulated was consistent with the growth-arrested phenotype of MazF-ms expressing cells. Curiously, the tRNA target and mechanism of action of MazF-ms paralleled that of one atypical MazF toxin in M. tuberculosis , suggesting manipulation of the levels of lysine tRNAs as the preferred conduit for reprogramming the proteomes via ribosome stalling at rare AAA codons in these GC-rich mycobacteria. [ABSTRACT FROM AUTHOR]

analysis of the halobacterial S9 operon reveals that it consists of two genes that encode protein subunits similar to the RPB10 and RPB6 subunits of the eukaryotic RNA polymerases I, II and III of Saccharomyces cerevisiae. The amino acid identity of archaebacterial and eukaryotic proteins was more than 60%. The amino acid similarity indicates that the protein subunits are highly conserved and mediate transcription.

The coordination of interrelated biochemical cell processes must be done efficiently. This is probably carried out at the level of transcription. An important part of this regulation is the multisubunit enzyme RNA polymerase II (Pol II). Research shows that the transcription of genes which encode the subunits of Pol II follows a divergent or head-to-head process which reflects the efficient organization of the yeast genome. It is hypothesized that the subunit genes are controlled by their upstream neighbors.

We have identified a conditional mutation in the shared Rpb6 subunit, assembled in RNA polymerases I, II, and III, that illuminated a new role that is independent of its assembly function. RNA polymerase II and Ill activities were significantly reduced in mutant cells before and after the shift to nonpermissive temperature. In contrast, RNA polymerase I was marginally affected. Although the Rpb6 mutant strain contained two mutations (P75S and Q100R), the majority of growth and transcription defects originated from substitution of an amino acid nearly identical in all eukaryotic counterparts as well as bacterial ω subunits (Q100R). Purification of mutant RNA polymerase II revealed that two subunits, Rpb4 and Rpb7, are selectively lost in mutant cells. Rpb4 and Rpb7 are present at substoichiometric levels, form a dissociable subcomplex, are required for RNA polymerase II activity at high temperatures, and have been implicated in the regulation of enzyme activity. Interaction experiments support a direct association between the Rpb6 and Rpb4 subunits, indicating that Rpb6 is one point of contact between the Rpb4/Rpb7 subcomplex and RNA polymerase II. The association of Rpb4/Rpb7 with Rpb6 suggests that analogous subunits of each RNA polymerase impart class-specific functions through a conserved core subunit. [ABSTRACT FROM AUTHOR]