Transcription factor dynamics

Abstract

Gene expression is a fundamental process that is highly conserved from humans to bacteria. The first step in gene expression, transcription, is performed by structurally conserved DNAdependent RNA polymerases (RNAPs), which results in the synthesis of an RNA molecule from a DNA template. In bacteria, a single species of RNAP is responsible for transcribing both stable RNA (i.e. t-and rRNA) and protein-encoding genes (i.e. mRNA), unlike eukaryotic systems, which use three distinct RNAP species to transcribe the different gene classes (RNAP I transcribes most rRNA, RNAP II transcribes mRNA, and RNAP III transcribes tRNA and 5S rRNA). The versatility of bacterial RNAP is dependent on both dynamic interactions with co-factors and the coding sequence of the template DNA, which allows RNAP to respond appropriately to the transcriptional needs of the cell. Although the majority of the research on gene expression has focused on the initiation stage, regulation of the elongation phase is essential for cell viability and represents an important topic for study. The elongation factors that associate with RNAP are unique and highly conserved among prokaryotes, making disruption of their interactions a potentially important target for antibiotic development. One of the most significant advances in molecular biology over the last decade has been the use of green fluorescent protein (GFP) and its spectral variants to observe the subcellular localization of proteins in live intact cells. This review discusses transcription dynamics with respect to RNAP and its associated transcription elongation factors in the two best-studied prokaryotes, Escherichia coli and Bacillus subtilis. RNAP and the transcription machineryProkaryotic RNA polymerase (RNAP) is a large (~400 kDa) multi-subunit enzyme comprising a 2 bb9v subunits which form a crab-claw-like structure. Although little sequence homology exists between eubacterial RNAP, archaeal RNAP and eukaryotic RNAPII, the crab-claw structure is remarkably conserved (Zhang et al., 1999;Cramer et al., 2001;Hirata et al., 2008). The two a subunits act as a scaffold to hold the catalytic b and b9 subunits together, forming the crab-claw-like structure (Zhang et al., 1999). The exact role of the v subunit is unclear but it is related in both structure and sequence to the eukaryotic polymerase subunit Rpb6 (Minakhin et al., 2001). It appears to be responsible for controlling transcription in response to nutrient shifts, correct folding of the b9 subunit and its assembly into the core multi-subunit enzyme (Mukherjee et al., 1999;Vrentas et al., 2005;Chatterji et al., 2007). The channel formed by b and b9 is referred to as the primary channel, which contains a deep positively charged cleft housing the enzyme's active site. During transcription, downstream double-stranded DNA separates into a singlestranded DNA template, which enters the primary channel and contacts the active site to allow polymerization of RNA (Borukhov & Nudler, 2008). Due to the crowding of the primary channel by the DNA : R...

Essential Biological Processes of an Emerging Pathogen: DNA Replication, Transcription, and Cell Division in Acinetobacter spp

“…A wide disparity is observed in the number of factors encoded by bacteria; e.g., E. coli has 7 different factors, B. subtilis has 17, and Streptomyces coelicolor has 63 (152 (80,175). Of these, only homologs of 32 and 54 are identifiable in Acinetobacter spp., along with one to five ECF factors (Table 4).…”

“…Targeting these protein-nucleic acid interactions may also be a valid approach for the development of new antibiotics, as preventing antitermination complex formation would severely disrupt cell proliferation. Indeed rRNA gene dosage is closely related to the rate of cell proliferation, with fast-growing species such as B. subtilis, E. coli, and Vibrio natriegens containing 7 to 13 rRNA operons per chromosome, while the slow-growing organism Mycobacterium tuberculosis contains a single rRNA gene (152). Acinetobacter species contain five to seven rRNA operons and are known to grow rapidly, suggesting that rRNA synthesis may be a valid target for bacteriostatic or bactericidal agent development.…”

“…In vivo, the RNAP core forms a series of transient complexes with other protein cofactors: 70 and other factors, antitermination factors (NusA, -B, -E, and -G), transcript cleavage factors (GreA and -B), and regulatory factors (e.g., DksA) (152). Ultimately, these interactions underpin the regulation of gene expression and the transcription of rRNA and thus play important roles in cell proliferation.…”

“…To initiate transcription (152), the ϳ400-kDa core must first associate with a factor to form the RNAP holoenzyme (␣ 2 ␤␤Ј). The architecture of the RNAP complex has been elucidated through extensive biochemical and biophysical analyses in E. coli (reviewed in reference 24) and crystal structures of the Thermus aquaticus RNAP core (276) and Thermus thermophilus holoenzyme (251).…”

Bacterial Transcription as a Target for Antibacterial Drug Development

“…TCs involved in rRNA synthesis are resistant to pausing signals and transcribe at higher rates than those involved in mRNA synthesis. The difference in responses of RNAP to pause signals, etc., during mRNA and rRNA synthesis is because of its association with different suites of elongation factors (157,159,160). Many of these transcription factors are essential for bacterial viability and are not highly conserved in eukaryotic cells.…”