Dataset Information

Direct activator/co-activator interaction is essential for bacteriophage T4 middle gene expression.

ABSTRACT: The bacteriophage T4 AsiA protein is a bifunctional regulator that inhibits transcription from the major class of bacterial promoters and also serves as an essential co-activator of transcription from T4 middle promoters. AsiA binds the primary s factor in Escherichia coli, sigma(70), and modifies the promoter recognition properties of the sigma(70)-containing RNA polymerase(RNAP) holoenzyme. In its role as co-activator, AsiA directs RNAP to T4 middle promoters in the presence of the T4-encoded activator MotA. According to the current model for T4 middle promoter activation, AsiA plays an indirect role in stabilizing the activation complex by facilitating interaction between DNA-bound MotA and sigma(70). Here we show that AsiA also plays a direct role in T4 middle promoter activation by contacting the MotA activation domain. Furthermore,we show that interaction between AsiA and the beta-flap domain of RNAP is important for co-activation. Based on our findings, we propose a revised model for T4 middle promoter activation, with AsiA organizing the activation complex via three distinct protein-protein interactions.

SUBMITTER: Yuan AH

PROVIDER: S-EPMC5673250 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Direct activator/co-activator interaction is essential for bacteriophage T4 middle gene expression.

Yuan Andy H AH Hochschild Ann A

Molecular microbiology 20091015 4

The bacteriophage T4 AsiA protein is a bifunctional regulator that inhibits transcription from the major class of bacterial promoters and also serves as an essential co-activator of transcription from T4 middle promoters. AsiA binds the primary s factor in Escherichia coli, sigma(70), and modifies the promoter recognition properties of the sigma(70)-containing RNA polymerase(RNAP) holoenzyme. In its role as co-activator, AsiA directs RNAP to T4 middle promoters in the presence of the T4-encoded ...[more]

PMID: 19843221

Similar Datasets

Project description:Phage T4 has provided countless contributions to the paradigms of genetics and biochemistry. Its complete genome sequence of 168,903 bp encodes about 300 gene products. T4 biology and its genomic sequence provide the best-understood model for modern functional genomics and proteomics. Variations on gene expression, including overlapping genes, internal translation initiation, spliced genes, translational bypassing, and RNA processing, alert us to the caveats of purely computational methods. The T4 transcriptional pattern reflects its dependence on the host RNA polymerase and the use of phage-encoded proteins that sequentially modify RNA polymerase; transcriptional activator proteins, a phage sigma factor, anti-sigma, and sigma decoy proteins also act to specify early, middle, and late promoter recognition. Posttranscriptional controls by T4 provide excellent systems for the study of RNA-dependent processes, particularly at the structural level. The redundancy of DNA replication and recombination systems of T4 reveals how phage and other genomes are stably replicated and repaired in different environments, providing insight into genome evolution and adaptations to new hosts and growth environments. Moreover, genomic sequence analysis has provided new insights into tail fiber variation, lysis, gene duplications, and membrane localization of proteins, while high-resolution structural determination of the "cell-puncturing device," combined with the three-dimensional image reconstruction of the baseplate, has revealed the mechanism of penetration during infection. Despite these advances, nearly 130 potential T4 genes remain uncharacterized. Current phage-sequencing initiatives are now revealing the similarities and differences among members of the T4 family, including those that infect bacteria other than Escherichia coli. T4 functional genomics will aid in the interpretation of these newly sequenced T4-related genomes and in broadening our understanding of the complex evolution and ecology of phages-the most abundant and among the most ancient biological entities on Earth.

Project description:The bacteriophage T4 encodes 10 proteins, known collectively as the replisome, that are responsible for the replication of the phage genome. The replisomal proteins can be subdivided into three activities; the replicase, responsible for duplicating DNA, the primosomal proteins, responsible for unwinding and Okazaki fragment initiation, and the Okazaki repair proteins. The replicase includes the gp43 DNA polymerase, the gp45 processivity clamp, the gp44/62 clamp loader complex, and the gp32 single-stranded DNA binding protein. The primosomal proteins include the gp41 hexameric helicase, the gp61 primase, and the gp59 helicase loading protein. The RNaseH, a 5' to 3' exonuclease and T4 DNA ligase comprise the activities necessary for Okazaki repair. The T4 provides a model system for DNA replication. As a consequence, significant effort has been put forth to solve the crystallographic structures of these replisomal proteins. In this review, we discuss the structures that are available and provide comparison to related proteins when the T4 structures are unavailable. Three of the ten full-length T4 replisomal proteins have been determined; the gp59 helicase loading protein, the RNase H, and the gp45 processivity clamp. The core of T4 gp32 and two proteins from the T4 related phage RB69, the gp43 polymerase and the gp45 clamp are also solved. The T4 gp44/62 clamp loader has not been crystallized but a comparison to the E. coli gamma complex is provided. The structures of T4 gp41 helicase, gp61 primase, and T4 DNA ligase are unknown, structures from bacteriophage T7 proteins are discussed instead. To better understand the functionality of T4 DNA replication, in depth structural analysis will require complexes between proteins and DNA substrates. A DNA primer template bound by gp43 polymerase, a fork DNA substrate bound by RNase H, gp43 polymerase bound to gp32 protein, and RNase H bound to gp32 have been crystallographically determined. The preparation and crystallization of complexes is a significant challenge. We discuss alternate approaches, such as small angle X-ray and neutron scattering to generate molecular envelopes for modeling macromolecular assemblies.

Dataset Information

Direct activator/co-activator interaction is essential for bacteriophage T4 middle gene expression.

Publications

Direct activator/co-activator interaction is essential for bacteriophage T4 middle gene expression.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets