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Internalization or exclusion of inducers Regulation of transcription initiation Regulation of transcription elongation and termination Regulation of mRNA stability or processing of primary transcripts Regulatlion of translation Regulation of protein activity and stability
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Internalization or exclusion of inducers Regulation of transcription initiation Regulation of transcription elongation and termination Regulation of mRNA stability or processing of primary transcripts Regulatlion of translation Regulation of protein activity and stability Regulation of protein secretion or localization Signals Signals Signals Signals Signals Signals Gene expression is controlled at many levels in prokaryotes
Transcription and its control in prokaryotes 1. The DNA substrate A. Packaging and availability of the DNA B. Number and arrangement of the genes. C. Transcription control sequences. 1. Promoters 2. Sites for activators and repressors. 3. Sites for the regulation of DNA topology. 4. Sites for the termination of transcription. 2. The General Transcription apparatus. A. RNA Polymerase core enzyme. B. RNA Polymerase holoenzymes. C. NusA, GreA, GreB 3. The process of transcription initiation. A. Identifiable kinetic intermediates B. Biochemical characterization of the process. 4. Transcription elongation. A. Pausing, Sliding,Termination, and Arrest. B. Role of the RNA-DNA hybrid. 5. Transcription termination. A. Rho factor. B. Attenuation. C. Other mechanisms. 6. Examples of mechanisms of activation and repression of transcription initiation. A. The lac operon B. Regulation of nitrogen assimilation (ntr) genes.
Packaging of prokaryotic DNA DNA is highly condensed in all living organisms, including phage. The packaging ratio for E. coli is ~1000. Ruptured phage, showing DNA released from a single phage.
Packaging of prokaryotic DNA The DNA is condensed and localized to a structure known as the nucleoid.
Packaging of prokaryotic DNA Nucleoids can be visualized in intact cells by a variety of staining techniques.
Structure of the bacterial nucleoid Nucleoids are associated with the cell membrane, and may be isolated by various techniques. In isolated nucleoids, the DNA appears to form many loops attached to a central structure.
Both protein and RNA are thought to be involved in DNA packaging “Domain” hypothesis for chromosomal organization.
Nucleoids can be isolated by sucrose density gradient centrifugation
Developmental regulation of nucleoid structure in Caulobacter crescentus Caulobacter crescentus is a dimorphic bacterium with asymmetric cell division Change in nucleoid part of a program of cell development. Nucleoid very dense Nucleoid less dense
Much remains to be learned about the components and regulation of nucleoid structure Proteins involved-unknown RNA involved-unknown Regulation of structure-unknown Problem-The DNA must be packaged so as to fit within the cell, yet it appears to all be able to be expressed at any time.
Number and arrangement of genes in bacteria Smallest genome-Mycoplasma genitalium ~580,000 bp ~470 genes parasitic, primate respiratory and genital tracts. Escherichia coli- ~4,500,000 bp. ~4.000 genes Average gene 0.8-1 Kbp Genes close together, ~80% of coding capacity is genes mRNA multiple genes may form part of the same transcription unit, termed an operon
Arrangement of genes in prokaryotes Genes may overlap. The same strand of DNA serves as the template strand, but it is read in two different reading frames. Many examples. Note: translational control signals for downstream gene are within the upstream gene. Genes may be located entirely within another gene, the reading frame may be the same or different. Example-phage FC174. Genes may overlap, and be encoded by the complementary strands of the DNA duplex. Example-phage FC174.
Non-random distrubution of bacterial genes with respect to direction of DNA replication. leading strand lagging strand oriC Origin of replication termination site leading strand genes lagging strand E. coli-About 55% of genes are oriented as shown. Certain other bacteria show a more pronounced bias. Head-on collisions of DNA polymerase and RNA polymerase are avoided.
The transcription of adjacent genes affects gene expression Transcription of upstream genes may increase or decrease expression of downstream genes. Transcription of convergent genes may increase or decrease expression of adjacent gene.