Gene Expression Control Cell Biology Lecture 9 Dr. Frances Santiago-Schwarz ECB Chapter 8(cont) Gene Expression Control And selected topics from ECB Chapter 9

SUMMARY : Differences in the initiation of transcription between eukaryotes and prokaryotes. These differences are fundamental and you should be able to describe them!!!

Assembly of Eukaryotic Transcriptional Initiation Complex Eukaryotic RNA polymerase II requires general transcription factors. The general transcription factors assemble in the promoter region (on the TATA box) and help RNA polymerase II to bind and start transcription. ECB Fig 8-10

of RNA pol II allows RNA processing Phosphorylation of RNA pol II allows RNA processing proteins to assemble on its tail ECB Fig 8-12

Eukaryotic genes can be controlled from a distance Eukaryotic genes can be controlled from a distance. The DNA sequences bound by gene activators are called enhancers. These enhancers sites can be located thousands of bps away from the gene. They can also be located down stream of the gene. ECB Fig 8-13

Hormones Signaling molecules Stimulate or inhibit activity in target cells Mechanism of action varies May bind to cell surface May enter cell and bind to regulatory proteins May bind with enhancers in DNA Some have widespread effects Somatotropin (growth hormone) Others signal only certain cells at certain times Prolactin stimulates milk production

A single gene regulatory protein can coordinate the expression of many different genes. EXAMPLE: Glucocorticoid receptor protein -Group of genes regulated by the binding of the hormone-Glucocorticoid receptor protein complex to a regulatory site in the DNA of the gene -Response to stress

A single gene regulatory protein can coordinate the expression of many different genes. Activator Proteins bound Regulatory region ECB Fig 8-20

Nucleosomes are able to inhibit or activate the initiation phase of transcription when they are positioned at the promoter region. Nucleosome Methylation (-CH3) of DNA can inactivate genes Acetylation of histones allows activation of genes by DNA unpacking and transcription Starr Figure 15.2 Page 240

Covalent modification of core histone tails. Figure 4-35 Mol Bio Cell

Nucleosomes are able to activate the initiation phase of transcription when they are positioned at the promoter region. ECB Fig 8-14

Gene regulatory proteins and the location of their binding sites relative to the promoters are different for different genes Same for all RNA Pol genes ECB Fig 8-15

DNA methylation The addition of methyl groups to the 5’ carbon of cytosine occurs at a frequency of up to 1 in every 100 bases This method of control is found in vertebrates & plants, but not in invertebrates such as flies or round worms Methylation occurs through the action of enzymes called methyltransferases Methylation is concentrated into GC-rich islands of DNA which are found in promoters Methylation of promoters is strongly correlated with gene repression

Methylation of Cytosine 5-methyl cytidine is referred to as the 5th deoxynucleoside

- Changes in the methylation state of a promoter alters the level of expression of a gene - Important during development (genes that do not contribute to particular path become permanently shut down through methylation)

Methylation is the mechanism by which X-inactivation occurs Recall that women are XX while men are XY In order to even out the dose of the genes on the X between the sexes women have one X randomly shut down in each cell The mechanism of this dosage compensation is through methylation as well as permanent chromosome compaction This results in the Barr body found in female cells Once either Xp or Xm has been inactivated, it remains silent throughout all subsequent cell divisions of that cell and its progeny, indicating that the inactive state is faithfully maintained through many cycles of DNA replication and mitosis.

X-inactivation (64 cell embryo)

Gene Expression Control: Emerging Concepts double-stranded RNA molecules (dsRNA) efficiently and specifically inhibit gene products on a posttranscriptional level (gene silencing) RNA interference is the phenomenon in which double-stranded RNA (dsRNA) can specifically suppress expression of a target gene, and was originally discovered in C. elegans Host cells can detect the presence of dsRNA (frequently of viral origin) and initiate a program of drastic responses in attempt to eliminate it. dsRNA-directed ssRNA destruction is the basis of the technique of RNA interference (RNAi) that is used by researchers to block specific gene expression

RNAi is a two-step mechanism. -First, long dsRNAs are cleaved by an enzyme known as Dicer in 21-23 nucleotides (nt) fragments, called siRNAs for small interfering RNAs. -Second, siRNAs are recruited to a ribonuclease complex (termed RISC for RNA Induced Silencing Complex) which in turn mediates the cleavage of the target mRNA.

Double-stranded RNA-binding proteins could suppress RNA interference-mediated antiviral defences Zsuzsanna Lichner, Dániel Silhavy and József Burgyán Agricultural Biotechnology Center, Plant Biology Institute, P.O. Box 411, H-2101, Gödöllö, Hungary RNA interference (RNAi) is a double-stranded (ds)RNA-inducible, sequence-specific RNA-degradation mechanism that operates as a natural antiviral system in plants and animals. Successful virus infection requires evasion or suppression of RNAi. Indeed, RNAi suppressor proteins have been identified in plant and animal viruses, although the molecular mechanism of silencing inhibition is still poorly understood. Because many RNA viruses encode dsRNA-binding proteins (dsRBPs) and as RNAi is triggered by the accumulation of dsRNAs, dsRBPs were examined to see if they inhibit RNAi. Here, it is shown that heterologous dsRBPs suppressed RNAi in plants, indicating that in natural host-virus interactions, pathogen-encoded dsRBPs could inactivate RNAi-mediated host defences.

Curr Opin Chem Biol. 2003 Aug;7(4):516-23.   MicroRNAs: key participants in gene regulatory networks. Ke XS, Liu CM, Liu DP, Liang CC. National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Science & Peking Union Medical College, Beijing 100005, China. microRNAs (miRNAs) are a newly identified and surprisingly large class of endogenous tiny regulatory RNAs. They exhibit various expressional patterns and are highly conserved across species. Recently, several regulatory targets of miRNAs have been predicted. Functional analysis of the potential targets indicated that miRNAs may be involved in a wide range of pivotally biological events. The nature of miRNAs and their intersection with small interfering RNAs endow them with many regulatory advantages over proteins and make them a potent and novel means to regulate gene expression at almost all levels. Here we argue that miRNAs are key participants in gene regulatory network.

Altering of Genes and Genomes ECB Fig 9-3

www.pitt.edu/ ~heh1/research.html                                       Horizontal gene transfer is any process in which an organism transfers genetic material (i.e. DNA) to another cell that is not its offspring. By contrast, vertical transfer occurs when an organism receives genetic material from its ancestor, e.g. its parent or a species from which it evolved.     www.pitt.edu/ ~heh1/research.html

Horizontal gene transfer and evolution